Luis de Lecea

All Publications

• Sleep and vigilance states: Embracing spatiotemporal dynamics. Neuron Nir, Y., de Lecea, L. 2023

  Abstract

  The classic view of sleep and vigilance states is a global stationary perspective driven by the interaction betweenneuromodulators and thalamocortical systems. However, recent data are challenging this view by demonstrating that vigilance states are highly dynamic and regionally complex. Spatially, sleep- and wake-like states often co-occur across distinct brain regions, as in unihemispheric sleep, local sleep in wakefulness, and during development. Temporally, dynamic switching prevails around state transitions, during extended wakefulness, and in fragmented sleep. This knowledge, together with methods monitoring brain activity across multiple regions simultaneously at millisecond resolution with cell-type specificity, is rapidly shifting how we consider vigilance states. A new perspective incorporating multiple spatial and temporal scales may have important implications for considering the governing neuromodulatory mechanisms, the functional roles of vigilance states, and their behavioral manifestations. A modular and dynamic view highlights novel avenues for finer spatiotemporal interventions to improve sleep function.

  View details for DOI 10.1016/j.neuron.2023.04.012

  View details for PubMedID 37148873

• A tool for monitoring cell type-specific focused ultrasound neuromodulation and control of chronic epilepsy. Proceedings of the National Academy of Sciences of the United States of America Murphy, K. R., Farrell, J. S., Gomez, J. L., Stedman, Q. G., Li, N., Leung, S. A., Good, C. H., Qiu, Z., Firouzi, K., Butts Pauly, K., Khuri-Yakub, B. P., Michaelides, M., Soltesz, I., de Lecea, L. 2022; 119 (46): e2206828119

  Abstract

  Focused ultrasound (FUS) is a powerful tool for noninvasive modulation of deep brain activity with promising therapeutic potential for refractory epilepsy; however, tools for examining FUS effects on specific cell types within the deep brain do not yet exist. Consequently, how cell types within heterogeneous networks can be modulated and whether parameters can be identified to bias these networks in the context of complex behaviors remains unknown. To address this, we developed a fiber Photometry Coupled focused Ultrasound System (PhoCUS) for simultaneously monitoring FUS effects on neural activity of subcortical genetically targeted cell types in freely behaving animals. We identified a parameter set that selectively increases activity of parvalbumin interneurons while suppressing excitatory neurons in the hippocampus. A net inhibitory effect localized to the hippocampus was further confirmed through whole brain metabolic imaging. Finally, these inhibitory selective parameters achieved significant spike suppression in the kainate model of chronic temporal lobe epilepsy, opening the door for future noninvasive therapies.

  View details for DOI 10.1073/pnas.2206828119

  View details for PubMedID 36343238

• Adolescent sleep shapes social novelty preference in mice. Nature neuroscience Bian, W. J., Brewer, C. L., Kauer, J. A., de Lecea, L. 2022

  Abstract

  Sleep disturbances frequently occur in neurodevelopmental disorders such as autism, but the developmental role of sleep is largely unexplored, and a causal relationship between developmental sleep defects and behavioral consequences in adulthood remains elusive. Here, we show that in mice, sleep disruption (SD) in adolescence, but not in adulthood, causes long-lasting impairment in social novelty preference. Furthermore, adolescent SD alters the activation and release patterns of dopaminergic neurons in the ventral tegmental area (VTA) in response to social novelty. This developmental sleep function is mediated by balanced VTA activity during adolescence; chemogenetic excitation mimics, whereas silencing rescues, the social deficits of adolescent SD. Finally, we show that in Shank3-mutant mice, improving sleep or rectifying VTA activity during adolescence ameliorates adult social deficits. Together, our results identify a critical role of sleep and dopaminergic activity in the development of social interaction behavior.

  View details for DOI 10.1038/s41593-022-01076-8

  View details for PubMedID 35618950

• Hyperexcitable arousal circuits drive sleep instability during aging. Science (New York, N.Y.) Li, S. B., Damonte, V. M., Chen, C., Wang, G. X., Kebschull, J. M., Yamaguchi, H., Bian, W. J., Purmann, C., Pattni, R., Urban, A. E., Mourrain, P., Kauer, J. A., Scherrer, G., de Lecea, L. 2022; 375 (6583): eabh3021

  Abstract

  Sleep quality declines with age; however, the underlying mechanisms remain elusive. We found that hyperexcitable hypocretin/orexin (Hcrt/OX) neurons drive sleep fragmentation during aging. In aged mice, Hcrt neurons exhibited more frequent neuronal activity epochs driving wake bouts, and optogenetic activation of Hcrt neurons elicited more prolonged wakefulness. Aged Hcrt neurons showed hyperexcitability with lower KCNQ2 expression and impaired M-current, mediated by KCNQ2/3 channels. Single-nucleus RNA-sequencing revealed adaptive changes to Hcrt neuron loss in the aging brain. Disruption of Kcnq2/3 genes in Hcrt neurons of young mice destabilized sleep, mimicking aging-associated sleep fragmentation, whereas the KCNQ-selective activator flupirtine hyperpolarized Hcrt neurons and rejuvenated sleep architecture in aged mice. Our findings demonstrate a mechanism underlying sleep instability during aging and a strategy to improve sleep continuity.

  View details for DOI 10.1126/science.abh3021

  View details for PubMedID 35201886

• Parallel circuits from the bed nuclei of stria terminalis to the lateral hypothalamus drive opposing emotional states NATURE NEUROSCIENCE Giardino, W. J., Eban-Rothschild, A., Christoffel, D. J., Li, S., Malenka, R. C., de Lecea, L. 2018; 21 (8): 1084-+

  View details for DOI 10.1038/s41593-018-0198-x

  View details for Web of Science ID 000439859000011

• VTA dopaminergic neurons regulate ethologically relevant sleep-wake behaviors. Nature neuroscience Eban-Rothschild, A., Rothschild, G., Giardino, W. J., Jones, J. R., de Lecea, L. 2016; 19 (10): 1356-1366

  Abstract

  Dopaminergic ventral tegmental area (VTA) neurons are critically involved in a variety of behaviors that rely on heightened arousal, but whether they directly and causally control the generation and maintenance of wakefulness is unknown. We recorded calcium activity using fiber photometry in freely behaving mice and found arousal-state-dependent alterations in VTA dopaminergic neurons. We used chemogenetic and optogenetic manipulations together with polysomnographic recordings to demonstrate that VTA dopaminergic neurons are necessary for arousal and that their inhibition suppresses wakefulness, even in the face of ethologically relevant salient stimuli. Nevertheless, before inducing sleep, inhibition of VTA dopaminergic neurons promoted goal-directed and sleep-related nesting behavior. Optogenetic stimulation, in contrast, initiated and maintained wakefulness and suppressed sleep and sleep-related nesting behavior. We further found that different projections of VTA dopaminergic neurons differentially modulate arousal. Collectively, our findings uncover a fundamental role for VTA dopaminergic circuitry in the maintenance of the awake state and ethologically relevant sleep-related behaviors.

  View details for DOI 10.1038/nn.4377

  View details for PubMedID 27595385

• Tuning arousal with optogenetic modulation of locus coeruleus neurons NATURE NEUROSCIENCE Carter, M. E., Yizhar, O., Chikahisa, S., Nguyen, H., Adamantidis, A., Nishino, S., Deisseroth, K., de Lecea, L. 2010; 13 (12): 1526-U117

  Abstract

  Neural activity in the noradrenergic locus coeruleus correlates with periods of wakefulness and arousal. However, it is unclear whether tonic or phasic activity in these neurons is necessary or sufficient to induce transitions between behavioral states and to promote long-term arousal. Using optogenetic tools in mice, we found that there is a frequency-dependent, causal relationship among locus coeruleus firing, cortical activity, sleep-to-wake transitions and general locomotor arousal. We also found that sustained, high-frequency stimulation of the locus coeruleus at frequencies of 5 Hz and above caused reversible behavioral arrests. These results suggest that the locus coeruleus is finely tuned to regulate organismal arousal and that bursts of noradrenergic overexcitation cause behavioral attacks that resemble those seen in people with neuropsychiatric disorders.

  View details for DOI 10.1038/nn.2682

  View details for Web of Science ID 000284525800018

  View details for PubMedID 21037585

  View details for PubMedCentralID PMC3174240

• Phasic Firing in Dopaminergic Neurons Is Sufficient for Behavioral Conditioning SCIENCE Tsai, H., Zhang, F., Adamantidis, A., Stuber, G. D., Bonci, A., de Lecea, L., Deisseroth, K. 2009; 324 (5930): 1080-1084

  Abstract

  Natural rewards and drugs of abuse can alter dopamine signaling, and ventral tegmental area (VTA) dopaminergic neurons are known to fire action potentials tonically or phasically under different behavioral conditions. However, without technology to control specific neurons with appropriate temporal precision in freely behaving mammals, the causal role of these action potential patterns in driving behavioral changes has been unclear. We used optogenetic tools to selectively stimulate VTA dopaminergic neuron action potential firing in freely behaving mammals. We found that phasic activation of these neurons was sufficient to drive behavioral conditioning and elicited dopamine transients with magnitudes not achieved by longer, lower-frequency spiking. These results demonstrate that phasic dopaminergic activity is sufficient to mediate mammalian behavioral conditioning.

  View details for DOI 10.1126/science.1168878

  View details for Web of Science ID 000266246700044

  View details for PubMedID 19389999

• Neural substrates of awakening probed with optogenetic control of hypocretin neurons NATURE Adamantidis, A. R., Zhang, F., Aravanis, A. M., Deisseroth, K., de Lecea, L. 2007; 450 (7168): 420-U9

  Abstract

  The neural underpinnings of sleep involve interactions between sleep-promoting areas such as the anterior hypothalamus, and arousal systems located in the posterior hypothalamus, the basal forebrain and the brainstem. Hypocretin (Hcrt, also known as orexin)-producing neurons in the lateral hypothalamus are important for arousal stability, and loss of Hcrt function has been linked to narcolepsy. However, it is unknown whether electrical activity arising from Hcrt neurons is sufficient to drive awakening from sleep states or is simply correlated with it. Here we directly probed the impact of Hcrt neuron activity on sleep state transitions with in vivo neural photostimulation, genetically targeting channelrhodopsin-2 to Hcrt cells and using an optical fibre to deliver light deep in the brain, directly into the lateral hypothalamus, of freely moving mice. We found that direct, selective, optogenetic photostimulation of Hcrt neurons increased the probability of transition to wakefulness from either slow wave sleep or rapid eye movement sleep. Notably, photostimulation using 5-30 Hz light pulse trains reduced latency to wakefulness, whereas 1 Hz trains did not. This study establishes a causal relationship between frequency-dependent activity of a genetically defined neural cell type and a specific mammalian behaviour central to clinical conditions and neurobehavioural physiology.

  View details for DOI 10.1038/nature06310

  View details for Web of Science ID 000250918600055

  View details for PubMedID 17943086

• The murine meninges acquire lymphoid tissue properties and harbour autoreactive B cells during chronic Trypanosoma brucei infection. PLoS biology Quintana, J. F., Sinton, M. C., Chandrasegaran, P., Kumar Dubey, L., Ogunsola, J., Al Samman, M., Haley, M., McConnell, G., Kuispond Swar, N. R., Ngoyi, D. M., Bending, D., de Lecea, L., MacLeod, A., Mabbott, N. A. 2023; 21 (11): e3002389

  Abstract

  The meningeal space is a critical brain structure providing immunosurveillance for the central nervous system (CNS), but the impact of infections on the meningeal immune landscape is far from being fully understood. The extracellular protozoan parasite Trypanosoma brucei, which causes human African trypanosomiasis (HAT) or sleeping sickness, accumulates in the meningeal spaces, ultimately inducing severe meningitis and resulting in death if left untreated. Thus, sleeping sickness represents an attractive model to study immunological dynamics in the meninges during infection. Here, by combining single-cell transcriptomics and mass cytometry by time-of-flight (CyTOF) with in vivo interventions, we found that chronic T. brucei infection triggers the development of ectopic lymphoid aggregates (ELAs) in the murine meninges. These infection-induced ELAs were defined by the presence of ER-TR7+ fibroblastic reticular cells, CD21/35+ follicular dendritic cells (FDCs), CXCR5+ PD1+ T follicular helper-like phenotype, GL7+ CD95+ GC-like B cells, and plasmablasts/plasma cells. Furthermore, the B cells found in the infected meninges produced high-affinity autoantibodies able to recognise mouse brain antigens, in a process dependent on LTβ signalling. A mid-throughput screening identified several host factors recognised by these autoantibodies, including myelin basic protein (MBP), coinciding with cortical demyelination and brain pathology. In humans, we identified the presence of autoreactive IgG antibodies in the cerebrospinal fluid (CSF) of second stage HAT patients that recognised human brain lysates and MBP, consistent with our findings in experimental infections. Lastly, we found that the pathological B cell responses we observed in the meninges required the presence of T. brucei in the CNS, as suramin treatment before the onset of the CNS stage prevented the accumulation of GL7+ CD95+ GC-like B cells and brain-specific autoantibody deposition. Taken together, our data provide evidence that the meningeal immune response during chronic T. brucei infection results in the acquisition of lymphoid tissue-like properties, broadening our understanding of meningeal immunity in the context of chronic infections. These findings have wider implications for understanding the mechanisms underlying the formation ELAs during chronic inflammation resulting in autoimmunity in mice and humans, as observed in other autoimmune neurodegenerative disorders, including neuropsychiatric lupus and multiple sclerosis.

  View details for DOI 10.1371/journal.pbio.3002389

  View details for PubMedID 37983289

• Dorsomedial and preoptic hypothalamic circuits control torpor. Current biology : CB Yamaguchi, H., Murphy, K. R., Fukatsu, N., Sato, K., Yamanaka, A., de Lecea, L. 2023

  Abstract

  Endotherms can survive low temperatures and food shortage by actively entering a hypometabolic state known as torpor. Although the decrease in metabolic rate and body temperature (Tb) during torpor is controlled by the brain, the specific neural circuits underlying these processes have not been comprehensively elucidated. In this study, we identify the neural circuits involved in torpor regulation by combining whole-brain mapping of torpor-activated neurons, cell-type-specific manipulation of neural activity, and viral tracing-based circuit mapping. We find that Trpm2-positive neurons in the preoptic area and Vgat-positive neurons in the dorsal medial hypothalamus are activated during torpor. Genetic silencing shows that the activity of either cell type is necessary to enter the torpor state. Finally, we show that these cells receive projections from the arcuate and suprachiasmatic nucleus and send projections to brain regions involved in thermoregulation. Our results demonstrate an essential role of hypothalamic neurons in the regulation of Tb and metabolic rate during torpor and identify critical nodes of the torpor regulatory network.

  View details for DOI 10.1016/j.cub.2023.10.076

  View details for PubMedID 37992720

• Sleep and the hypothalamus. Science (New York, N.Y.) Adamantidis, A. R., de Lecea, L. 2023; 382 (6669): 405-412

  Abstract

  Neural substrates of wakefulness, rapid eye movement sleep (REMS), and non-REMS (NREMS) in the mammalian hypothalamus overlap both anatomically and functionally with cellular networks that support physiological and behavioral homeostasis. Here, we review the roles of sleep neurons of the hypothalamus in the homeostatic control of thermoregulation or goal-oriented behaviors during wakefulness. We address how hypothalamic circuits involved in opposing behaviors such as core body temperature and sleep compute conflicting information and provide a coherent vigilance state. Finally, we highlight some of the key unresolved questions and challenges, and the promise of a more granular view of the cellular and molecular diversity underlying the integrative role of the hypothalamus in physiological and behavioral homeostasis.

  View details for DOI 10.1126/science.adh8285

  View details for PubMedID 37883555

• Cell type specific focused ultrasound neuromodulation in preclinical models of sleep and psychiatric disorders. Neuropsychopharmacology : official publication of the American College of Neuropsychopharmacology Murphy, K. R., de Lecea, L. 2023

  View details for DOI 10.1038/s41386-023-01662-9

  View details for PubMedID 37463978

• The Effect of Food Motivational Status and Hypocretin (Orexin) 1 Receptor Antagonism on Decision Processes in the Murine Go/No-Go Behavioural Task Jacobson, L., Ji, M., Metha, J., Murawski, C., Nicholson, J., De Lecea, L., Hoyer, D. WILEY. 2023: 1025-1026

  View details for Web of Science ID 001043027400729

• Adolescent sleep defects and dopaminergic hyperactivity in mice with a schizophrenia-linked Shank3 mutation. Sleep Bian, W. J., González, O. C., de Lecea, L. 2023

  Abstract

  Shank3 is a shared risk gene for autism spectrum disorders and schizophrenia. Sleep defects have been characterized for autism models with Shank3 mutations, however, evidence has been lacking for the potential sleep defects caused by Shank3 mutation associated with schizophrenia and how early in development these defects may occur. Here we characterized the sleep architecture of adolescent mice carrying a schizophrenia-linked, R1117X mutation in Shank3. We further employed GRABDA dopamine sensor and fiber photometry to record dopamine release in the nucleus accumbens during sleep/wake states. Our results show that homozygous mutant R1117X mice have significantly reduced sleep in the dark phase during adolescence, altered electroencephalogram power especially during the rapid-eye-movement sleep, and dopamine hyperactivity during sleep but not during wakefulness. Further analyses suggest that these adolescent defects in sleep architecture and dopaminergic neuromodulation tightly correlate with the social novelty preference later in adulthood and predict adult social performance during same-sex social interactions. Our results provide novel insights to the sleep phenotypes in mouse models of schizophrenia and the potential use of developmental sleep as a predictive metric for adult social symptoms. Together with recent studies in other Shank3 models, our work underscores the idea that Shank3-involved circuit disruptions may be one of the shared pathologies in certain types of schizophrenia and autism. Future research is needed to establish the causal relationship among adolescent sleep defects, dopaminergic dysregulation, and adult behavioral changes in Shank3 mutation animals and other models.

  View details for DOI 10.1093/sleep/zsad131

  View details for PubMedID 37144901

• Myeloid deficiency of the intrinsic clock protein BMAL1 accelerates cognitive aging by disrupting microglial synaptic pruning. Journal of neuroinflammation Iweka, C. A., Seigneur, E., Hernandez, A. L., Paredes, S. H., Cabrera, M., Blacher, E., Pasternak, C. T., Longo, F. M., de Lecea, L., Andreasson, K. I. 2023; 20 (1): 48

  Abstract

  Aging is associated with loss of circadian immune responses and circadian gene transcription in peripheral macrophages. Microglia, the resident macrophages of the brain, also show diurnal rhythmicity in regulating local immune responses and synaptic remodeling. To investigate the interaction between aging and microglial circadian rhythmicity, we examined mice deficient in the core clock transcription factor, BMAL1. Aging Cd11bcre;Bmallox/lox mice demonstrated accelerated cognitive decline in association with suppressed hippocampal long-term potentiation and increases in immature dendritic spines. C1q deposition at synapses and synaptic engulfment were significantly decreased in aging Bmal1-deficient microglia, suggesting that BMAL1 plays a role in regulating synaptic pruning in aging. In addition to accelerated age-associated hippocampal deficits, Cd11bcre;Bmallox/lox mice also showed deficits in the sleep-wake cycle with increased wakefulness across light and dark phases. These results highlight an essential role of microglial BMAL1 in maintenance of synapse homeostasis in the aging brain.

  View details for DOI 10.1186/s12974-023-02727-8

  View details for PubMedID 36829230

  View details for PubMedCentralID 8157517

• Automated Sleep Deprivation Setup Using a Shaking Platform in Mice. Bio-protocol Bian, W., De Lecea, L. 2023; 13 (4): e4620

  Abstract

  The functions of sleep remain largely unclear, and even less is known about its role in development. A general strategy to tackle these questions is to disrupt sleep and measure the outcomes. However, some existing sleep deprivation methods may not be suitable for studying the effects of chronic sleep disruption, due to their lack of effectiveness and/or robustness, substantial stress caused by the deprivation method, or consuming a large quantity of time and manpower. More problems may be encountered when applying these existing protocols to young, developing animals, because of their likely heightened vulnerability to stressors, and difficulties in precisely monitoring sleep at young ages. Here, we report a protocol of automated sleep disruption in mice using a commercially available, shaking platform-based deprivation system. We show that this protocol effectively and robustly deprives both non-rapid-eye-movement (NREM) sleep and rapid-eye-movement (REM) sleep without causing a significant stress response, and does not require human supervision. This protocol uses adolescent mice, but the method also works with adult mice. Graphical abstract Automated sleep deprivation system. The platform of the deprivation chamber was programmed to shake in a given frequency and intensity to keep the animal awake while its brain and muscle activities were continuously monitored by electroencephalography and electromyography.

  View details for DOI 10.21769/BioProtoc.4620

  View details for PubMedID 36845529

• Optogenetic and pharmacological interventions link hypocretin neurons to impulsivity in mice. Communications biology Tyree, S. M., Jennings, K. J., Gonzalez, O. C., Li, S., Nicholson, J. R., von Heimendahl, M., de Lecea, L. 2023; 6 (1): 74

  Abstract

  Neurons in the lateral hypothalamus expressing the neuropeptide Hypocretin, also known as orexin, are known critical modulators of arousal stability. However, their role in the different components of the arousal construct such as attention and decision making is poorly understood. Here we study Hypocretin neuronal circuit dynamics during stop action impulsivity in a Go/NoGo task in mice. We show that Hypocretin neuronal activity correlates with anticipation of reward. We then assessed the causal role of Hypocretin neuronal activity using optogenetics in a Go/NoGo task. We show that stimulation of Hypocretin neurons during the cue period dramatically increases the number of premature responses. These effects are mimicked by amphetamine, reduced by atomoxetine, a norepinephrine uptake inhibitor, and blocked by a Hypocretin receptor 1 selective antagonist. We conclude that Hypocretin neurons have a key role in the integration of salient stimuli during wakefulness to produce appropriate and timely responses to rewarding and aversive cues.

  View details for DOI 10.1038/s42003-023-04409-w

  View details for PubMedID 36658362

• Single cell and spatial transcriptomic analyses reveal microglia-plasma cell crosstalk in the brain during Trypanosoma brucei infection. Nature communications Quintana, J. F., Chandrasegaran, P., Sinton, M. C., Briggs, E. M., Otto, T. D., Heslop, R., Bentley-Abbot, C., Loney, C., de Lecea, L., Mabbott, N. A., MacLeod, A. 2022; 13 (1): 5752

  Abstract

  Human African trypanosomiasis, or sleeping sickness, is caused by the protozoan parasite Trypanosoma brucei and induces profound reactivity of glial cells and neuroinflammation when the parasites colonise the central nervous system. However, the transcriptional and functional responses of the brain to chronic T. brucei infection remain poorly understood. By integrating single cell and spatial transcriptomics of the mouse brain, we identify that glial responses triggered by infection are readily detected in the proximity to the circumventricular organs, including the lateral and 3rd ventricle. This coincides with the spatial localisation of both slender and stumpy forms of T. brucei. Furthermore, in silico predictions and functional validations led us to identify a previously unknown crosstalk between homeostatic microglia and Cd138+ plasma cells mediated by IL-10 and B cell activating factor (BAFF) signalling. This study provides important insights and resources to improve understanding of the molecular and cellular responses in the brain during infection with African trypanosomes.

  View details for DOI 10.1038/s41467-022-33542-z

  View details for PubMedID 36180478

• The brake matters: Hyperexcitable arousal circuits in sleep fragmentation with age. Clinical and translational medicine Li, S., de Lecea, L. 2022; 12 (6): e900

  View details for DOI 10.1002/ctm2.900

  View details for PubMedID 35696605

• Lateral hypothalamic galanin neurons are activated by stress and blunt anxiety-like behavior in mice. Behavioural brain research Owens-French, J., Li, S., Francois, M., Leigh Townsend, R., Daniel, M., Soulier, H., Turner, A., de Lecea, L., Munzberg, H., Morrison, C., Qualls-Creekmore, E. 1800: 113773

  Abstract

  Despite the prevalence of anxiety disorders, the molecular identity of neural circuits underlying anxiety remains unclear. The lateral hypothalamus (LH) is one brain region implicated in the regulation of anxiety, and our recent data found that chemogenetic activation of LH galanin neurons attenuated the stress response to a novel environment as measured by the marble burying test. Thus, we hypothesize that LH galanin neurons may contribute to anxiety-related behavior. We used chemogenetics and fiber photometry to test the ability of LH galanin neurons to influence anxiety and stress-related behavior. Chemogenetic activation of LH galanin neurons significantly decreased anxiety-like behavior in the elevated plus maze, open field test, and light dark test. However, LH galanin activation did not alter restraint stress induced HPA activation or freezing behavior in the fear conditioning paradigm. In vivo calcium monitoring by fiber photometry indicated that LH galanin neurons were activated by anxiogenic and/or stressful stimuli including tail suspension, novel mouse interaction, and predator odor. Further, in a fear conditioning task, calcium transients strongly increased during foot shock, but were not affected by the unconditioned stimulus tone. These data indicate that LH galanin neurons both respond to and modulate anxiety, with no influence on stress induced HPA activation or fear behaviors. Further investigation of LH galanin circuitry and functional mediators of behavioral output may offer a more refined pharmacological target as an alternative to first-line broad pharmacotherapies such as benzodiazepines.

  View details for DOI 10.1016/j.bbr.2022.113773

  View details for PubMedID 35101456

• Hypocretins (orexins): The ultimate translational neuropeptides. Journal of internal medicine Jacobson, L. H., Hoyer, D., de Lecea, L. 1800

  Abstract

  The hypocretins (Hcrts), also known as orexins, are two neuropeptides produced exclusively in the lateral hypothalamus. They act on two specific receptors that are widely distributed across the brain and involved in a myriad of neurophysiological functions that include sleep, arousal, feeding, reward, fear, anxiety and cognition. Hcrt cell loss in humans leads to narcolepsy with cataplexy (narcolepsy type 1), a disorder characterized by intrusions of sleep into wakefulness, demonstrating that the Hcrt system is nonredundant and essential for sleep/wake stability. The causal link between Hcrts and arousal/wakefulness stabilisation has led to the development of a new class of drugs, Hcrt receptor antagonists to treat insomnia, based on the assumption that blocking orexin-induced arousal will facilitate sleep. This has been clinically validated: currently, two Hcrt receptor antagonists are approved to treat insomnia (suvorexant and lemborexant), with a New Drug Application recently submitted to the US Food and Drug Administration for a third drug (daridorexant). Other therapeutic applications under investigation include reduction of cravings in substance-use disorders and prevention of neurodegenerative disorders such as Alzheimer's disease, given the apparent bidirectional relationship between poor sleep and worsening of the disease. Circuit neuroscience findings suggest that the Hcrt system is a hub that integrates diverse inputs modulating arousal (e.g., circadian rhythms, metabolic status, positive and negative emotions) and conveys this information to multiple output regions. This neuronal architecture explains the wealth of physiological functions associated with Hcrts and highlights the potential of the Hcrt system as a therapeutic target for a number of disorders. We discuss present and future possible applications of drugs targeting the Hcrt system for the treatment of circuit-related neuropsychiatric and neurodegenerative conditions.

  View details for DOI 10.1111/joim.13406

  View details for PubMedID 35043499

• Peripheral Lipopolyssacharide Rapidly Silences REM-Active LHGABA Neurons. Frontiers in behavioral neuroscience Borniger, J. C., de Lecea, L. 2021; 15: 649428

  Abstract

  Immune factors (e.g., cytokines, chemokines) can alter the activity of neuronal circuits to promote "sickness behavior," a suite of adaptive actions that organisms exhibit in response to infection/injury in order to maximize their chances of recovery (i.e., return to homeostasis). This includes drastic alterations in sleep/wake states, locomotor activity, and food intake, among other behaviors. Despite the ample evidence highlighting interactions between the brain and systemic immunity, studies on how immune challenges alter the activity of genetically defined cell populations controlling arousal states are scarce. As the lateral hypothalamus (LH) serves a major integrative function in behavioral arousal, food intake, and monitoring and responding to changes in systemic physiology, we investigated how GABAergic neurons within this brain region alter their activity across normal sleep/wake states and in response to a peripheral immune challenge with bacterial endotoxin [lipopolysaccharides (LPS)]. Using fiber photometry (GCaMP6s Ca2+ signal) in tandem with electroencephalogram (EEG)/EMG recordings to determine arousal states, we observed that population activity of GABAergic neurons in the lateral hypothalamus (LHGABA) is highest during rapid-eye-movement sleep (REM), and this activity changes drastically across spontaneous arousal state transitions, with the lowest activity observed during non-REM sleep. Upon intraperitoneal LPS challenge, LHGABA neurons rapidly decrease their activity in tandem with elimination of REM sleep behavior (characteristic of cytokine-induced sickness). Together, these data suggest that peripheral immune challenges can rapidly (in < 40 min) alter subcortical neuronal circuits controlling arousal states. Additionally, we demonstrate that fiber photometry offers a sensitive and cell-type specific tool that can be applied to study the neuronal substrates of sickness behavior.

  View details for DOI 10.3389/fnbeh.2021.649428

  View details for PubMedID 33716686

• Twenty-Three Years of Hypocretins: The "Rosetta Stone" of Sleep/Arousal Circuits. Frontiers of neurology and neuroscience de Lecea, L. 2021; 45: 1-10

  Abstract

  The discovery of the hypocretins/orexins (HCRTs) has revolutionized sleep science in the last two decades. A combination of anatomical tracing methods, optogenetics, and pharmacology is delineating a blueprint of functional inputs and outputs of the HCRT system. Here, we discuss several models of HCRT action that involve the integration between physiological variables, circadian output, and sleep homeostasis. Generation of activity maps during the sleep-wake cycle at the cellular level will allow investigators to decipher computational frameworks modeling operations of HCRT networks.

  View details for DOI 10.1159/000514961

  View details for PubMedID 34052811

• Heterogeneity of Hypocretin/Orexin Neurons. Frontiers of neurology and neuroscience Sagi, D., de Lecea, L., Appelbaum, L. 2021; 45: 61-74

  Abstract

  The multifunctional, hypothalamic hypocretin/orexin (HCRT)-producing neurons regulate an array of physiological and behavioral states including arousal, sleep, feeding, emotions, stress, and reward. How a presumably uniform HCRT neuron population regulates such a diverse set of functions is not clear. The role of the HCRT neuropeptides may vary depending on the timing and localization of secretion and neuronal activity. Moreover, HCRT neuropeptides may not mediate all functions ascribed to HCRT neurons. Some could be orchestrated by additional neurotransmitters and neuropeptides that are expressed in HCRT neurons. We hypothesize that HCRT neurons are segregated into genetically, anatomically and functionally distinct subpopulations. We discuss accumulating data that suggest the existence of such HCRT neuron subpopulations that may effectuate the diverse functions of these neurons in mammals and fish.

  View details for DOI 10.1159/000514964

  View details for PubMedID 34052814

• Hypocretin/Orexins and Hyperarousal de Lecea, L. SPRINGERNATURE. 2020: 31

  View details for Web of Science ID 000596371000065

• Neural and Hormonal Control of Sexual Behavior. Endocrinology Jennings, K. J., de Lecea, L. 2020

  Abstract

  Gonadal hormones contribute to the sexual differentiation of brain and behavior throughout the lifespan, from initial neural patterning to 'activation' of adult circuits. Sexual behavior is an ideal system in which to investigate the mechanisms underlying hormonal activation of neural circuits. Sexual behavior is a hormonally regulated, innate social behavior found across species. Although both sexes seek out and engage in sexual behavior, the specific actions involved in mating are sexually dimorphic. Thus, the neural circuits mediating sexual motivation and behavior in males and females are overlapping yet distinct. Furthermore, sexual behavior is strongly dependent on circulating gonadal hormones in both sexes. There has been significant recent progress on elucidating how gonadal hormones modulate physiological properties within sexual behavior circuits with consequences for behavior. Therefore, in this mini-review we review the neural circuits of male and female sexual motivation and behavior from initial sensory detection of pheromones to the extended amygdala and on to medial hypothalamic nuclei and reward systems. We also discuss how gonadal hormones impact the physiology and functioning of each node within these circuits. By better understanding the myriad of ways in which gonadal hormones impact sexual behavior circuits, we can gain a richer and more complete appreciation for the neural substrates of complex behavior.

  View details for DOI 10.1210/endocr/bqaa150

  View details for PubMedID 32845294

• Impaired hypocretin/orexin system alters responses to salient stimuli in obese male mice. The Journal of clinical investigation Tan, Y., Hang, F., Liu, Z., Stoiljkovic, M., Wu, M., Tu, Y., Han, W., Lee, A. M., Kelley, C., Hajos, M., Lu, L., de Lecea, L., de Araujo, I., Picciotto, M., Horvath, T. L., Gao, X. 2020

  Abstract

  The brain has evolved in an environment where food sources are scarce and foraging for food is one of the major challenges for survival of the individual and species. Basic and clinical studies show that obesity/overnutrition leads to overwhelming changes in the brain in animals and humans. However, the exact mechanisms underlying the consequences of excessive energy intake are not well understood. Neurons expressing the neuropeptide hypocretin/orexin (Hcrt) in the lateral/perifonical hypothalamus (LH) are critical for homeostatic regulation, reward seeking, stress response, and cognitive functions. In this study, we examined adaptations in Hcrt cells regulating behavioral responses to salient stimuli in diet-induced obese mice. Our results demonstrated changes in primary cilia, synaptic transmission and plasticity, cellular responses to neurotransmitters necessary for reward seeking and stress responses in Hcrt neurons from obese mice. Activities of neuronal networks in the LH and hippocampus were impaired as a result of decreased hypocretinergic function. The weakened Hcrt system decreased reward seeking while altering responses to acute stress (stress coping strategy), which were reversed by selectively activating Hcrt cells with chemogenetics. Taken together, our data suggest that a deficiency in the Hcrt signaling may be a common cause of behavioral changes (such as lowered arousal, weakened reward seek and altered stress response) in obese animals.

  View details for DOI 10.1172/JCI130889

  View details for PubMedID 32516139

• Sleep and neuropsychiatric illness NEUROPSYCHOPHARMACOLOGY Winkelman, J. W., de Lecea, L. 2020; 45 (1): 1–2

  View details for DOI 10.1038/s41386-019-0514-5

  View details for Web of Science ID 000499141800001

  View details for PubMedID 31486776

  View details for PubMedCentralID PMC6879507

• Neurobiological and Hormonal Mechanisms Regulating Women's Sleep. Frontiers in neuroscience Dorsey, A., de Lecea, L., Jennings, K. J. 2020; 14: 625397

  Abstract

  Sleep is crucial for optimal well-being, and sex differences in sleep quality have significant implications for women's health. We review the current literature on sex differences in sleep, such as differences in objective and subjective sleep measures and their relationship with aging. We then discuss the convincing evidence for the role of ovarian hormones in regulating female sleep, and survey how these hormones act on a multitude of brain regions and neurochemicals to impact sleep. Lastly, we identify several important areas in need of future research to narrow the knowledge gap and improve the health of women and other understudied populations.

  View details for DOI 10.3389/fnins.2020.625397

  View details for PubMedID 33519372

• Hypothalamic circuitry underlying stress-induced insomnia and peripheral immunosuppression Science Advances Li, S. B., Borniger, J. C., Yamaguchi, H., Hédou, J., Gaudilliere, B., de Lecea, L. 2020; 6 (37)

  View details for DOI 10.1126/sciadv.abc2590

• Editorial: Hypocretins/Orexins. Frontiers in endocrinology Lopez, M., de Lecea, L., Dieguez, C. 2020; 11: 357

  View details for DOI 10.3389/fendo.2020.00357

  View details for PubMedID 32582031

• Brain Circuit of Claustrophobia-like Behavior in Mice Identified by Upstream Tracing of Sighing. Cell reports Li, P. n., Li, S. B., Wang, X. n., Phillips, C. D., Schwarz, L. A., Luo, L. n., de Lecea, L. n., Krasnow, M. A. 2020; 31 (11): 107779

  Abstract

  Emotions are distinct patterns of behavioral and physiological responses triggered by stimuli that induce different brain states. Elucidating the circuits is difficult because of challenges in interrogating emotional brain states and their complex outputs. Here, we leverage the recent discovery in mice of a neural circuit for sighing, a simple, quantifiable output of various emotions. We show that mouse confinement triggers sighing, and this "claustrophobic" sighing, but not accompanying tachypnea, requires the same medullary neuromedin B (Nmb)-expressing neurons as physiological sighing. Retrograde tracing from the Nmb neurons identified 12 forebrain centers providing presynaptic input, including hypocretin (Hcrt)-expressing lateral hypothalamic neurons. Confinement activates Hcrt neurons, and optogenetic activation induces sighing and tachypnea whereas pharmacologic inhibition suppresses both responses. The effect on sighing is mediated by HCRT directly on Nmbneurons. We propose that this HCRT-NMB neuropeptide relay circuit mediates claustrophobic sighing and that activated Hcrt neurons are a claustrophobia brain state that directly controls claustrophobic outputs.

  View details for DOI 10.1016/j.celrep.2020.107779

  View details for PubMedID 32553161

• The hypocretin (orexin) system: from a neural circuitry perspective. Neuropharmacology Li, S. B., de Lecea, L. n. 2020; 167: 107993

  Abstract

  Hypocretin/orexin neurons are distributed restrictively in the hypothalamus, a brain region known to orchestrate diverse functions including sleep, reward processing, food intake, thermogenesis, and mood. Since the hypocretins/orexins were discovered more than two decades ago, extensive studies have accumulated concrete evidence showing the pivotal role of hypocretin/orexin in diverse neural modulation. New method of viral-mediated tracing system offers the possibility to map the monosynaptic inputs and detailed anatomical connectivity of Hcrt neurons. With the development of powerful research techniques including optogenetics, fiber-photometry, cell-type/pathway specific manipulation and neuronal activity monitoring, as well as single-cell RNA sequencing, the details of how hypocretinergic system execute functional modulation of various behaviors are coming to light. In this review, we focus on the function of neural pathways from hypocretin neurons to target brain regions. Anatomical and functional inputs to hypocretin neurons are also discussed. We further briefly summarize the development of pharmaceutical compounds targeting hypocretin signaling. This article is part of the special issue on Neuropeptides.

  View details for DOI 10.1016/j.neuropharm.2020.107993

  View details for PubMedID 32135427

• Arousal-state dependent alterations in VTA-GABAergic neuronal activity. eNeuro Eban-Rothschild, A. n., Borniger, J. C., Rothschild, G. n., Giardino, W. J., Morrow, J. G., de Lecea, L. n. 2020

  Abstract

  Decades of research have implicated the ventral tegmental area (VTA) in motivation, learning and reward processing. We and others recently demonstrated that it also serves as an important node in sleep/wake regulation. Specifically, VTA-dopaminergic neuron activation is sufficient to drive wakefulness and necessary for the maintenance of wakefulness. However, the role of VTA-GABAergic neurons in arousal regulation is not fully understood. It is still unclear whether VTA-GABAergic neurons predictably alter their activity across arousal states, what is the nature of interactions between VTA-GABAergic activity and cortical oscillations, and how activity in VTA-GABAergic neurons relates to VTA-dopaminergic neurons in the context of sleep/wake regulation. To address these, we simultaneously recorded population activity from VTA-subpopulations and EEG/EMG signals during spontaneous sleep/wake states and in the presence of salient stimuli in freely-behaving mice. We found that VTA-GABAergic neurons exhibit robust arousal-state-dependent alterations in population activity, with high activity and transients during wakefulness and REM sleep. During wakefulness, population activity of VTA-GABAergic neurons, but not VTA-dopaminergic neurons, was positively correlated with EEG gamma power and negatively correlated with theta power. During NREM sleep, population activity in both VTA-GABAergic and VTA-dopaminergic neurons negatively correlated with delta, theta, and sigma power bands. Salient stimuli, with both positive and negative valence, activated VTA-GABAergic neurons. Together, our data indicate that VTA-GABAergic neurons, like their dopaminergic counterparts, drastically alter their activity across sleep-wake states. Changes in their activity predicts cortical oscillatory patterns reflected in the EEG, which are distinct from EEG spectra associated with dopaminergic neural activity.Statement of Significance Little is known about how ventral tegmental area (VTA) neural ensembles couple arousal to motivated behaviors. Using cell-type specific genetic tools, we investigated the population activity of GABAergic and dopaminergic neurons within the VTA across sleep/wake states and in the presence of salient stimuli. We demonstrate that coordinated neural activity within VTA-GABAergic neurons peaks during wakefulness and REM sleep. Furthermore, neuronal activity in VTA-GABAergic neurons is correlated with high frequency, low amplitude cortical oscillations during waking, but negatively correlated with high amplitude slower frequency oscillations during NREM sleep. Our results demonstrate that VTA-GABAergic neuronal activity is tightly linked to cortical arousal and highlight this population as a potential important node in sleep/wake regulation.

  View details for DOI 10.1523/ENEURO.0356-19.2020

  View details for PubMedID 32054621

• Multisensory modulation of body ownership in mice. Neuroscience of consciousness Buckmaster, C. L., Rathmann-Bloch, J. E., de Lecea, L. n., Schatzberg, A. F., Lyons, D. M. 2020; 2020 (1): niz019

  Abstract

  Body ownership is a fundamental aspect of self-consciousness that reflects more than the presence of physical body parts. As demonstrated by the rubber hand illusion (RHI), human brains construct body ownership experiences using available multisensory information. Experimental conditions similar to those that induce the RHI in humans have been recently adapted to induce the rubber tail illusion (RTI) in mice. Here, we show that the RTI is enhanced in both sexes of mice by repetitive synchronous stroking comprised of correlated visual and tactile stimulation of real and rubber tails compared to visual-only mimicked stroking conducted without tactile stimulation. The RTI also appears to be enhanced in female but not male mice by slow compared to fast stroking that reflects an interoceptive manipulation associated with affective touch in humans. Sex differences in slow stroking effects are exploratory and require replication in mice. Sex differences have not been reported for the RHI in healthy humans, but women rate slow stroking as more affectively pleasant compared to the ratings of men. Results suggest that the RHI in humans resembles aspects of the RTI in mice. Studies of mice may therefore provide neurobiological insights on evolutionarily conserved mechanisms of bodily self-consciousness in humans.

  View details for DOI 10.1093/nc/niz019

  View details for PubMedID 31988796

  View details for PubMedCentralID PMC6977007

• CRISPR/Cas9 Editing of Neuropeptide Receptor Signaling Reveals an Extended Amygdala Circuit Mechanism Modulating Alcohol Drinking, Anxiety, and Avoidance Giardino, W., Yamaguchi, H., de Lecea, L. NATURE PUBLISHING GROUP. 2019: 505

  View details for Web of Science ID 000509665600913

• Construction of Viral Vectors for Cell Type-specific CRISPR Gene Editing in the Adult Mouse Brain. Bio-protocol Yamaguchi, H., de Lecea, L. 2019; 9 (16): e3334

  Abstract

  Recently developed gene editing technologies based on engineered CRISPR/Cas9 systems enables researchers to disrupt genes in a cell type-specific manner in the adult mouse brain. Using these technologies, we recently showed that the dopamine beta-hydroxylase gene in Locus Coeruleus (LC) norepinephrine neurons plays a vital role in the maintenance of wakefulness. Our method consists of four steps, (1) crossing Cre-dependent spCas9 knockin mice with a Cre-driver mouse line to express spCas9 in the target neural populations, (2) cloning of sgRNA, (3) construction of an AAV (adeno associated virus) vector expressing dual sgRNA, and (4) virus packaging and stereotaxic injection of the virus into the target brain area. Here, we describe a detailed protocol of AAV vector construction for cell type-specific CRISPR gene editing in the adult mouse brain. The method adopts a dual-sgRNA strategy for efficient disruption of the target gene. At first, a few different sgRNAs targeting the same gene are cloned into a plasmid expressing spCas9. After evaluation of the sgRNAs by a T7 endonuclease assay, the two most efficient sgRNAs are cloned in tandem into an AAV vector using the Gibson Assembly method.

  View details for DOI 10.21769/BioProtoc.3334

  View details for PubMedID 33654839

  View details for PubMedCentralID PMC7854134

• Construction of Viral Vectors for Cell Type-specific CRISPR Gene Editing in the Adult Mouse Brain BIO-PROTOCOL Yamaguchi, H., de Lecea, L. 2019; 9 (16)

  View details for DOI 10.21769/BioProtoc.3334

  View details for Web of Science ID 000482206700005

• In vivo cell type-specific CRISPR gene editing for sleep research JOURNAL OF NEUROSCIENCE METHODS Yamaguchi, H., de Lecea, L. 2019; 316: 99–102

  View details for DOI 10.1016/j.jneumeth.2018.10.016

  View details for Web of Science ID 000460717600010

• Hypocretin and the Regulation of Sleep-Wake Transitions HANDBOOK OF SLEEP RESEARCH, VOL 30 Nevarez, N., de Lecea, L., Dringenberg, H. C. 2019; 30: 89–99

  View details for DOI 10.1016/B978-0-12-813743-7.00006-2

  View details for Web of Science ID 000558272300007

• In vivo cell type-specific CRISPR knockdown of dopamine beta hydroxylase reduces locus coeruleus evoked wakefulness. Nature communications Yamaguchi, H., Hopf, F. W., Li, S. B., de Lecea, L. 2018; 9 (1): 5211

  Abstract

  Locus coeruleus (LC) neurons in the brainstem have long been associated with attention and arousal. Optogenetic stimulation of LC-NE neurons induces immediate sleep-to-wake transitions. However, LC neurons also secrete other neurotransmitters in addition to NE. To interrogate the role of NE derived from the LC in regulating wakefulness, we applied in vivo cell type-specific CRISPR/Cas9 technology to disrupt the dopamine beta hydroxylase (dbh) gene selectively in adult LC-NE neurons. Unilateral dbh gene disruption abolished immediate arousal following optogenetic stimulation of LC. Bilateral LC-specific dbh disruption significantly reduced NE concentration in LC projection areas and reduced wake length even in the presence of salient stimuli. These results suggest that NE may be crucial for the awakening effect of LC stimulation and serve as proof-of-principle that CRISPR gene editing in adult neurons can be used to interrogate gene function within genetically-defined neuronal circuitry associated with complex behaviors.

  View details for DOI 10.1038/s41467-018-07566-3

  View details for PubMedID 30523254

  View details for PubMedCentralID PMC6283864

• Parallel Circuits From the Bed Nuclei of Stria Terminalis to the Lateral Hypothalamus Drive Opposing Emotional States Giardino, W., Eban-Rothschild, A., Christoffel, D., Li, S., Malenka, R., de Lecea, L. NATURE PUBLISHING GROUP. 2018: S234

  View details for Web of Science ID 000509546600441

• In vivo cell type-specific CRISPR gene editing for sleep research. Journal of neuroscience methods Yamaguchi, H., de Lecea, L. 2018

  Abstract

  Sleep is an innate behavior conserved in all animals and, in vertebrates, is regulated by neuronal circuits in the brain. The conventional techniques of forward and reverse genetics have enabled researchers to investigate the molecular mechanisms that regulate sleep and arousal. However, functional interrogation of genes in specific cell subtypes in the brain remains a challenge. Here, we review the background of newly developed gene-editing technologies using engineered CRISPR/Cas9 system and describe the application to interrogate gene functions within genetically-defined brain cell populations in sleep research.

  View details for PubMedID 30439390

• Parallel circuits from the bed nuclei of stria terminalis to the lateral hypothalamus drive opposing emotional states. Nature neuroscience Giardino, W. J., Eban-Rothschild, A., Christoffel, D. J., Li, S., Malenka, R. C., de Lecea, L. 2018

  Abstract

  Lateral hypothalamus (LH) neurons containing the neuropeptide hypocretin (HCRT; orexin) modulate affective components of arousal, but their relevant synaptic inputs remain poorly defined. Here we identified inputs onto LH neurons that originate from neuronal populations in the bed nuclei of stria terminalis (BNST; a heterogeneous region of extended amygdala). We characterized two non-overlapping LH-projecting GABAergic BNST subpopulations that express distinct neuropeptides (corticotropin-releasing factor, CRF, and cholecystokinin, CCK). To functionally interrogate BNSTLH circuitry, we used tools for monitoring and manipulating neural activity with cell-type-specific resolution in freely behaving mice. We found that Crf-BNST and Cck-BNST neurons respectively provide abundant and sparse inputs onto Hcrt-LH neurons, display discrete physiological responses to salient stimuli, drive opposite emotionally valenced behaviors, and receive different proportions of inputs from upstream networks. Together, our data provide an advanced model for how parallel BNSTLH pathways promote divergent emotional states via connectivity patterns of genetically defined, circuit-specific neuronal subpopulations.

  View details for PubMedID 30038273

• Hypocretin as a Hub for Arousal and Motivation FRONTIERS IN NEUROLOGY Tyree, S. M., Bomiger, J. C., de Lecea, L. 2018; 9: 413

  Abstract

  The lateral hypothalamus is comprised of a heterogeneous mix of neurons that serve to integrate and regulate sleep, feeding, stress, energy balance, reward, and motivated behavior. Within these populations, the hypocretin/orexin neurons are among the most well studied. Here, we provide an overview on how these neurons act as a central hub integrating sensory and physiological information to tune arousal and motivated behavior accordingly. We give special attention to their role in sleep-wake states and conditions of hyper-arousal, as is the case with stress-induced anxiety. We further discuss their roles in feeding, drug-seeking, and sexual behavior, which are all dependent on the motivational state of the animal. We further emphasize the application of powerful techniques, such as optogenetics, chemogenetics, and fiber photometry, to delineate the role these neurons play in lateral hypothalamic functions.

  View details for PubMedID 29928253

• Hypocretin/orexin deficiency decreases cocaine abuse liability NEUROPHARMACOLOGY Steiner, N., Rossetti, C., Sakurai, T., Yanagisawa, M., de Lecea, L., Magistretti, P. J., Halfon, O., Boutrel, B. 2018; 133: 395–403

  Abstract

  Compelling evidence indicates that hypocretin/orexin signaling regulates arousal, stress and reward-seeking behaviors. However, most studies on drug reward-related processes have so far described the effects of pharmacological blockers disrupting hypocretin/orexin transmission. We report here an extensive study on cocaine-related behaviors in hypocretin/orexin-deficient mice (KO) and their heterozygous (HET) and wildtype (WT) littermates. We evaluated behavioral sensitization following repeated administrations and preference for an environment repeatedly paired with cocaine injections (15 mg/kg). Mice were also trained to self-administer cocaine (0.5-1.5 mg/kg/infusion). Our observations show that whereas all mice exhibited quite similar responses to acute administration of cocaine, only Hcrt KO mice exhibited reduced cocaine-seeking behaviors following a period of abstinence or extinction, and reduced cocaine incubation craving. Further, if the present findings confirm that Hcrt deficient mice may display a hypoactive phenotype, possibly linked to a reduced alertness concomitant to a decreased exploration of their environment, hypocretin/orexin defiency did not cause any attentional deficit. We thus report that innate disruption of hypocretin/orexin signaling moderately alters cocaine reward but significantly reduces long-term affective dependence that may explain the lack of relapse for cocaine seeking seen in Hcrt KO mice. Overall, with blunted cocaine intake at the highest concentration and reduced responsiveness to cocaine cues after prolonged abstinence, our findings suggest that hypocretin deficient mice may display signs of resilience to cocaine addiction.

  View details for PubMedID 29454841

• Neuronal Mechanisms for Sleep/Wake Regulation and Modulatory Drive NEUROPSYCHOPHARMACOLOGY Eban-Rothschild, A., Appelbaum, L., de Lecea, L. 2018; 43 (5): 937–52

  Abstract

  Humans have been fascinated by sleep for millennia. After almost a century of scientific interrogation, significant progress has been made in understanding the neuronal regulation and functions of sleep. The application of new methods in neuroscience that enable the analysis of genetically defined neuronal circuits with unprecedented specificity and precision has been paramount in this endeavor. In this review, we first discuss electrophysiological and behavioral features of sleep/wake states and the principal neuronal populations involved in their regulation. Next, we describe the main modulatory drives of sleep and wakefulness, including homeostatic, circadian, and motivational processes. Finally, we describe a revised integrative model for sleep/wake regulation.

  View details for PubMedID 29206811

  View details for PubMedCentralID PMC5854814

• Optical Probing of Orexin/Hypocretin Receptor Antagonists Sleep Li, S., Nevárez, N., Giardino, W. J., de Lecea, L. 2018

  View details for DOI 10.1093/sleep/zsy141

• In vivo cell type-specific CRISPR knockdown of dopamine beta hydroxylase reduces locus coeruleus evoked wakefulness Nature Communications Yamaguchi, H., Hopf, F., Li, S., de Lecea, L. 2018; 9

  View details for DOI 10.1038/s41467-018-07566-3

• Recent advances in understanding the roles of hypocretin/orexin in arousal, affect, and motivation. F1000Research Nevarez, N., de Lecea, L. 2018; 7

  Abstract

  The hypocretins (Hcrts) are two alternatively spliced neuropeptides (Hcrt1/Ox-A and Hcrt2/Ox-B) that are synthesized exclusively in the hypothalamus. Data collected in the 20 years since their discovery have supported the view that the Hcrts play a broad role in the control of arousal with a particularly important role in the maintenance of wakefulness and sleep-to-wake transitions. While this latter point has received an overwhelming amount of research attention, a growing literature has begun to broaden our understanding of the many diverse roles that the Hcrts play in physiology and behavior. Here, we review recent advances in the neurobiology of Hcrt in three sections. We begin by surveying findings on Hcrt function within normal sleep/wake states as well as situations of aberrant sleep (that is, narcolepsy). In the second section, we discuss research establishing a role for Hcrt in mood and affect (that is, anxiety, stress, and motivation). Finally, in the third section, we briefly discuss future directions for the field and place an emphasis on analytical modeling of Hcrt neural activity. We hope that the data discussed here provide a broad overview of recent progress in the field and make clear the diversity of roles played by these neuromodulators.

  View details for PubMedID 30254737

• Optical Probing of Orexin/Hypocretin Receptor Antagonists. Sleep Li, S. B., Nevárez, N. n., Giardino, W. J., de Lecea, L. n. 2018

  Abstract

  The present study investigated the function of Hypocretin (Hcrt or Orexin/OX) receptor antagonists in sleep modulation and memory function with optical methods in transgenic mice.We used Hcrt-IRES-Cre knock-in mice and AAV vectors expressing channelrhodopsin-2 (ChR2) to render Hcrt neurons sensitive to blue light stimulation. We optogenetically stimulated Hcrt neurons and measured latencies to wakefulness in the presence or absence of OX1/2R antagonists and Zolpidem. We also examined endogenous Hcrt neuronal activity with fiber photometry. Changes in memory after optogenetic sleep disruption were evaluated by the novel object recognition test (NOR) and compared for groups treated with vehicle, OX1/2R antagonists, or Zolpidem. We also analyzed EEG power spectra of wakefulness, rapid eye movement (REM) sleep, and non-REM (NREM) sleep following the injections of vehicle, OX1/2R antagonists, and Zolpidem in young adult mice.Acute optogenetic stimulation of Hcrt neurons at different frequencies resulted in wakefulness. Treatment with dual OX1/2R antagonists (DORAs) DORA12 and MK6096, as well as selective OX2R antagonist MK1064 and Zolpidem, but not selective OX1R antagonist 1SORA1, significantly reduced the bout length of optogenetic stimulation-evoked wakefulness episode. Fiber photometry recordings of GCaMP6f signals showed that Hcrt neurons are active during wakefulness, even in the presence of OXR antagonists. Treatment with dual OX1/2R antagonists improved memory function despite optogenetic sleep fragmentation caused impaired memory function in a NOR test.Our results show DORAs and selective OX2R antagonists stabilize sleep and improve sleep-dependent cognitive processes even when challenged by optogenetic stimulation mimicking highly arousing stimuli.

  View details for PubMedID 30060151

• Hypothalamic Tuberomammillary Nucleus Neurons: Electrophysiological Diversity and Essential Role in Arousal Stability JOURNAL OF NEUROSCIENCE Fujita, A., Bonnavion, P., Wilson, M. H., Mickelsen, L. E., Bloit, J., de Lecea, L., Jackson, A. C. 2017; 37 (39): 9574–92

  Abstract

  Histaminergic (HA) neurons, found in the posterior hypothalamic tuberomammillary nucleus (TMN), extend fibers throughout the brain and exert modulatory influence over numerous physiological systems. Multiple lines of evidence suggest that the activity of HA neurons is important in the regulation of vigilance despite the lack of direct, causal evidence demonstrating its requirement for the maintenance of arousal during wakefulness. Given the strong correlation between HA neuron excitability and behavioral arousal, we investigated both the electrophysiological diversity of HA neurons in brain slices and the effect of their acute silencing in vivo in male mice. For this purpose, we first validated a transgenic mouse line expressing cre recombinase in histidine decarboxylase-expressing neurons (Hdc-Cre) followed by a systematic census of the membrane properties of both HA and non-HA neurons in the ventral TMN (TMNv) region. Through unsupervised hierarchical cluster analysis, we found electrophysiological diversity both between TMNv HA and non-HA neurons, and among HA neurons. To directly determine the impact of acute cessation of HA neuron activity on sleep-wake states in awake and behaving mice, we examined the effects of optogenetic silencing of TMNv HA neurons in vivo We found that acute silencing of HA neurons during wakefulness promotes slow-wave sleep, but not rapid eye movement sleep, during a period of low sleep pressure. Together, these data suggest that the tonic firing of HA neurons is necessary for the maintenance of wakefulness, and their silencing not only impairs arousal but is sufficient to rapidly and selectively induce slow-wave sleep.SIGNIFICANCE STATEMENT The function of monoaminergic systems and circuits that regulate sleep and wakefulness is often disrupted as part of the pathophysiology of many neuropsychiatric disorders. One such circuit is the posterior hypothalamic histamine (HA) system, implicated in supporting wakefulness and higher brain function, but has been difficult to selectively manipulate owing to cellular heterogeneity in this region. Here we use a transgenic mouse to interrogate both the characteristic firing properties of HA neurons and their specific role in maintaining wakefulness. Our results demonstrate that the acute, cell type-specific silencing of HA neurons during wakefulness is sufficient to not only impair arousal but to rapidly and selectively induce slow-wave sleep. This work furthers our understanding of HA-mediated mechanisms that regulate behavioral arousal.

  View details for PubMedID 28874450

• To sleep or not to sleep: neuronal and ecological insights. Current opinion in neurobiology Eban-Rothschild, A., Giardino, W. J., de Lecea, L. 2017; 44: 132-138

  Abstract

  Daily, animals need to decide when to stop engaging in cognitive processes and behavioral responses to the environment, and go to sleep. The main processes regulating the daily organization of sleep and wakefulness are circadian rhythms and homeostatic sleep pressure. In addition, motivational processes such as food seeking and predator evasion can modulate sleep/wake behaviors. Here, we discuss the principal processes regulating the propensity to stay awake or go to sleep-focusing on neuronal and behavioral aspects. We first introduce the neuronal populations involved in sleep/wake regulation. Next, we describe the circadian and homeostatic drives for sleep. Then, we highlight studies demonstrating various effects of motivational processes on sleep/wake behaviors, and discuss possible neuronal mechanisms underlying their control.

  View details for DOI 10.1016/j.conb.2017.04.010

  View details for PubMedID 28500869

• Rat intersubjective decisions are encoded by frequency-specific oscillatory contexts. Brain and behavior Schaich Borg, J. n., Srivastava, S. n., Lin, L. n., Heffner, J. n., Dunson, D. n., Dzirasa, K. n., de Lecea, L. n. 2017; 7 (6): e00710

  Abstract

  It is unknown how the brain coordinates decisions to withstand personal costs in order to prevent other individuals' distress. Here we test whether local field potential (LFP) oscillations between brain regions create "neural contexts" that select specific brain functions and encode the outcomes of these types of intersubjective decisions.Rats participated in an "Intersubjective Avoidance Test" (IAT) that tested rats' willingness to enter an innately aversive chamber to prevent another rat from getting shocked. c-Fos immunoreactivity was used to screen for brain regions involved in IAT performance. Multi-site local field potential (LFP) recordings were collected simultaneously and bilaterally from five brain regions implicated in the c-Fos studies while rats made decisions in the IAT. Local field potential recordings were analyzed using an elastic net penalized regression framework.Rats voluntarily entered an innately aversive chamber to prevent another rat from getting shocked, and c-Fos immunoreactivity in brain regions known to be involved in human empathy-including the anterior cingulate, insula, orbital frontal cortex, and amygdala-correlated with the magnitude of "intersubjective avoidance" each rat displayed. Local field potential recordings revealed that optimal accounts of rats' performance in the task require specific frequencies of LFP oscillations between brain regions in addition to specific frequencies of LFP oscillations within brain regions. Alpha and low gamma coherence between spatially distributed brain regions predicts more intersubjective avoidance, while theta and high gamma coherence between a separate subset of brain regions predicts less intersubjective avoidance. Phase relationship analyses indicated that choice-relevant coherence in the alpha range reflects information passed from the amygdala to cortical structures, while coherence in the theta range reflects information passed in the reverse direction.These results indicate that the frequency-specific "neural context" surrounding brain regions involved in social cognition encodes outcomes of decisions that affect others, above and beyond signals from any set of brain regions in isolation.

  View details for PubMedID 28638715

  View details for PubMedCentralID PMC5474713

• Stress Coping and Resilience Modeled in Mice ANIMAL MODELS FOR THE STUDY OF HUMAN DISEASE, 2ND EDITION Lyons, D. M., de Lecea, L., Schatzberg, A. F., Conn, P. M. 2017: 1145-1153

  View details for Web of Science ID 000479185800045

• Behavioral Neuroscience of Orexin/Hypocretin Preface BEHAVIORAL NEUROSCIENCE OF OREXIN/HYPOCRETIN Lawrence, A. J., de Lecea, L., Lawrence, A. J., DeLecea, L. 2017; 33: V-VI

  View details for Web of Science ID 000528635600001

• Optogenetic Investigation of Arousal Circuits. International journal of molecular sciences Tyree, S. M., de Lecea, L. n. 2017; 18 (8)

  Abstract

  Modulation between sleep and wake states is controlled by a number of heterogeneous neuron populations. Due to the topological proximity and genetic co-localization of the neurons underlying sleep-wake state modulation optogenetic methods offer a significant improvement in the ability to benefit from both the precision of genetic targeting and millisecond temporal control. Beginning with an overview of the neuron populations mediating arousal, this review outlines the progress that has been made in the investigation of arousal circuits since the incorporation of optogenetic techniques and the first in vivo application of optogenetic stimulation in hypocretin neurons in the lateral hypothalamus. This overview is followed by a discussion of the future progress that can be made by incorporating more recent technological developments into the research of neural circuits.

  View details for PubMedID 28809797

• Lateral Hypothalamic Control of the Ventral Tegmental Area: Reward Evaluation and the Driving of Motivated Behavior. Frontiers in systems neuroscience Tyree, S. M., de Lecea, L. n. 2017; 11: 50

  Abstract

  The lateral hypothalamus (LH) plays an important role in many motivated behaviors, sleep-wake states, food intake, drug-seeking, energy balance, etc. It is also home to a heterogeneous population of neurons that express and co-express multiple neuropeptides including hypocretin (Hcrt), melanin-concentrating hormone (MCH), cocaine- and amphetamine-regulated transcript (CART) and neurotensin (NT). These neurons project widely throughout the brain to areas such as the locus coeruleus, the bed nucleus of the stria terminalis, the amygdala and the ventral tegmental area (VTA). Lateral hypothalamic projections to the VTA are believed to be important for driving behavior due to the involvement of dopaminergic reward circuitry. The purpose of this article is to review current knowledge regarding the lateral hypothalamic connections to the VTA and the role they play in driving these behaviors.

  View details for DOI 10.3389/fnsys.2017.00050

  View details for PubMedID 28729827

  View details for PubMedCentralID PMC5498520

• Neuronal substrates for initiation, maintenance, and structural organization of sleep/wake states. F1000Research Eban-Rothschild, A., de Lecea, L. 2017; 6: 212-?

  Abstract

  Animals continuously alternate between sleep and wake states throughout their life. The daily organization of sleep and wakefulness is orchestrated by circadian, homeostatic, and motivational processes. Over the last decades, much progress has been made toward determining the neuronal populations involved in sleep/wake regulation. Here, we will discuss how the application of advanced in vivo tools for cell type-specific manipulations now permits the functional interrogation of different features of sleep/wake state regulation: initiation, maintenance, and structural organization. We will specifically focus on recent studies examining the roles of wake-promoting neuronal populations.

  View details for DOI 10.12688/f1000research.9677.1

  View details for PubMedID 28357049

• Hypocretins and Arousal. Current topics in behavioral neurosciences Li, S., Giardino, W. J., de Lecea, L. 2016

  Abstract

  How the brain controls vigilance state transitions remains to be fully understood. The discovery of hypocretins, also known as orexins, and their link to narcolepsy has undoubtedly allowed us to advance our knowledge on key mechanisms controlling the boundaries and transitions between sleep and wakefulness. Lack of function of hypocretin neurons (a relatively simple and non-redundant neuronal system) results in inappropriate control of sleep states without affecting the total amount of sleep or homeostatic mechanisms. Anatomical and functional evidence shows that the hypothalamic neurons that produce hypocretins/orexins project widely throughout the entire brain and interact with major neuromodulator systems in order to regulate physiological processes underlying wakefulness, attention, and emotions. Here, we review the role of hypocretins/orexins in arousal state transitions, and discuss possible mechanisms by which such a relatively small population of neurons controls fundamental brain state dynamics.

  View details for DOI 10.1007/7854_2016_58

  View details for PubMedID 28012091

• VTA Dopaminergic Neurons Regulate Ethologically Relevant Sleep-Wake Behaviors Eban-Rothschild, A., Rothschild, G., Giardino, W. J., Jones, J. R., de Lecea, L. NATURE PUBLISHING GROUP. 2016: S259

  View details for Web of Science ID 000440365600441

• Obesity- and gender-dependent role of endogenous somatostatin and cortistatin in the regulation of endocrine and metabolic homeostasis in mice SCIENTIFIC REPORTS Luque, R. M., Cordoba-Chacon, J., Pozo-Salas, A. I., Porteiro, B., de Lecea, L., Nogueiras, R., Gahete, M. D., Castano, J. P. 2016; 6

  Abstract

  Somatostatin (SST) and cortistatin (CORT) regulate numerous endocrine secretions and their absence [knockout (KO)-models] causes important endocrine-metabolic alterations, including pituitary dysregulations. We have demonstrated that the metabolic phenotype of single or combined SST/CORT KO-models is not drastically altered under normal conditions. However, the biological actions of SST/CORT are conditioned by the metabolic-status (e.g. obesity). Therefore, we used male/female SST- and CORT-KO mice fed low-fat (LF) or high-fat (HF) diet to explore the interplay between SST/CORT and obesity in the control of relevant pituitary-axes and whole-body metabolism. Our results showed that the SST/CORT role in the control of GH/prolactin secretions is maintained under LF- and HF-diet conditions as SST-KOs presented higher GH/prolactin-levels, while CORT-KOs displayed higher GH- and lower prolactin-levels than controls under both diets. Moreover, the impact of lack of SST/CORT on the metabolic-function was gender- and diet-dependent. Particularly, SST-KOs were more sensitive to HF-diet, exhibiting altered growth and body-composition (fat/lean percentage) and impaired glucose/insulin-metabolism, especially in males. Conversely, only males CORT-KO under LF-diet conditions exhibited significant alterations, displaying higher glucose-levels and insulin-resistance. Altogether, these data demonstrate a tight interplay between SST/CORT-axis and the metabolic status in the control of endocrine/metabolic functions and unveil a clear dissociation of SST/CORT roles.

  View details for DOI 10.1038/srep37992

  View details for Web of Science ID 000389199600001

  View details for PubMedID 27901064

  View details for PubMedCentralID PMC5128804

• Hubs and spokes of the lateral hypothalamus: cell types, circuits and behaviour JOURNAL OF PHYSIOLOGY-LONDON Bonnavion, P., Mickelsen, L. E., Fujita, A., de Lecea, L., Jackson, A. C. 2016; 594 (22): 6443-6462

  Abstract

  The hypothalamus is among the most phylogenetically conserved regions in the vertebrate brain, reflecting its critical role in maintaining physiological and behavioural homeostasis. By integrating signals arising from both the brain and periphery, it governs a litany of behaviourally important functions essential for survival. In particular, the lateral hypothalamic area (LHA) is central to the orchestration of sleep-wake states, feeding, energy balance and motivated behaviour. Underlying these diverse functions is a heterogeneous assembly of cell populations typically defined by neurochemical markers, such as the well-described neuropeptides hypocretin/orexin and melanin-concentrating hormone. However, anatomical and functional evidence suggests a rich diversity of other cell populations with complex neurochemical profiles that include neuropeptides, receptors and components of fast neurotransmission. Collectively, the LHA acts as a hub for the integration of diverse central and peripheral signals and, through complex local and long-range output circuits, coordinates adaptive behavioural responses to the environment. Despite tremendous progress in our understanding of the LHA, defining the identity of functionally discrete LHA cell types, and their roles in driving complex behaviour, remain significant challenges in the field. In this review, we discuss advances in our understanding of the neurochemical and cellular heterogeneity of LHA neurons and the recent application of powerful new techniques, such as opto- and chemogenetics, in defining the role of LHA circuits in feeding, reward, arousal and stress. From pioneering work to recent developments, we review how the interrogation of LHA cells and circuits is contributing to a mechanistic understanding of how the LHA coordinates complex behaviour.

  View details for DOI 10.1113/JP271946

  View details for Web of Science ID 000389029900003

  View details for PubMedID 27302606

  View details for PubMedCentralID PMC5108896

• Fasting modulates GH/IGF-I axis and its regulatory systems in the mammary gland of female mice: Influence of endogenous cortistatin. Molecular and cellular endocrinology Villa-Osaba, A., Gahete, M. D., Cordoba-Chacon, J., de Lecea, L., Castaño, J. P., Luque, R. M. 2016; 434: 14-24

  Abstract

  Growth hormone (GH) and insulin-like growth factor-I (IGF-I) are essential factors in mammary-gland (MG) development and are altered during fasting. However, no studies have investigated the alterations in the expression of GH/IGF-I and its regulatory systems (somatostatin/cortistatin and ghrelin) in MG during fasting. Therefore, this study was aimed at characterizing the regulation of GH/IGF-I/somatostatin/cortistatin/ghrelin-systems expression in MG of fasted female-mice (compared to fed-controls) and the influence of endogenous-cortistatin (using cortistatin-knockouts). Fasting decreased IGF-I while increased IGF-I/Insulin-receptors expression in MGs. Fasting provoked an increase in GH expression that might be associated to enhanced ghrelin-variants/ghrelin-O-acyl-transferase enzyme expression, while an upregulation of somatostatin-receptors was observed. However, cortistatin-knockouts mice showed a decrease in GH and somatostatin receptor-subtypes expression. Altogether, we demonstrate that GH/IGF-I, somatostatin/cortistatin and ghrelin systems expression is altered in MG during fasting, suggesting a relevant role in coordinating its response to metabolic stress, wherein endogenous cortistatin might be essential for an appropriate response.

  View details for DOI 10.1016/j.mce.2016.06.014

  View details for PubMedID 27291340

• In vivo assessment of behavioral recovery and circulatory exchange in the peritoneal parabiosis model SCIENTIFIC REPORTS Castellano, J. M., Palner, M., Li, S., Freeman, G. M., Andy Nguyen, A., Shen, B., Stan, T., Mosher, K. I., Chin, F. T., de Lecea, L., Luo, J., Wyss-Coray, T. 2016; 6

  Abstract

  The sharing of circulation between two animals using a surgical procedure known as parabiosis has created a wealth of information towards our understanding of physiology, most recently in the neuroscience arena. The systemic milieu is a complex reservoir of tissues, immune cells, and circulating molecules that is surprisingly not well understood in terms of its communication across organ systems. While the model has been used to probe complex physiological questions for many years, critical parameters of recovery and exchange kinetics remain incompletely characterized, limiting the ability to design experiments and interpret results for complex questions. Here we provide evidence that mice joined by parabiosis gradually recover much physiology relevant to the study of brain function. Specifically, we describe the timecourse for a variety of recovery parameters, including those for general health and metabolism, motor coordination, activity, and sleep behavior. Finally, we describe the kinetics of chimerism for several lymphocyte populations as well as the uptake of small molecules into the brains of mice following parabiosis. Our characterization provides an important resource to those attempting to understand the complex interplay between the immune system and the brain as well as other organ systems.

  View details for DOI 10.1038/srep29015

  View details for Web of Science ID 000378851500002

  View details for PubMedID 27364522

  View details for PubMedCentralID PMC4929497

• Cortistatin Is a Key Factor Regulating the Sex-Dependent Response of the GH and Stress Axes to Fasting in Mice ENDOCRINOLOGY Cordoba-Chacon, J., Gahete, M. D., Pozo-Salas, A. I., de Lecea, L., Castano, J. P., Luque, R. M. 2016; 157 (7): 2810-2823

  Abstract

  Cortistatin (CORT) shares high structural and functional similarities with somatostatin (SST) but displays unique sex-dependent pituitary actions. Indeed, although female CORT-knockout (CORT-KO) mice exhibit enhanced GH expression/secretion, Proopiomelanocortin expression, and circulating ACTH/corticosterone/ghrelin levels, male CORT-KO mice only display increased plasma GH/corticosterone levels. Changes in peripheral ghrelin and SST (rather than hypothalamic levels) seem to regulate GH/ACTH axes in CORT-KOs under fed conditions. Because changes in GH/ACTH axes during fasting provide important adaptive mechanisms, we sought to determine whether CORT absence influences GH/ACTH axes during fasting. Accordingly, fed and fasted male/female CORT-KO were compared with littermate controls. Fasting increased circulating GH levels in male/female controls but not in CORT-KO, suggesting that CORT can be a relevant regulator of GH secretion during fasting. However, GH levels were already higher in CORT-KO than in controls in fed state, which might preclude a further elevation in GH levels. Interestingly, although fasting-induced pituitary GH expression was elevated in both male/female controls, GH expression only increased in fasted female CORT-KOs, likely owing to specific changes observed in key factors controlling somatotrope responsiveness (ie, circulating ghrelin and IGF-1, and pituitary GHRH and ghrelin receptor expression). Fasting increased corticosterone levels in control and, most prominently, in CORT-KO mice, which might be associated with a desensitization to SST signaling and to an augmentation in CRH and ghrelin-signaling regulating corticotrope function. Altogether, these results provide compelling evidence that CORT plays a key, sex-dependent role in the regulation of the GH/ACTH axes in response to fasting.

  View details for DOI 10.1210/en.2016-1195

  View details for Web of Science ID 000378877200022

  View details for PubMedID 27175972

• Hypocretin (Orexin) System and its Functions in Arousal/sleep de Lecea, L. ELSEVIER SCIENCE INC. 2016: 170S-171S

  View details for Web of Science ID 000432440802081

• Superficial Layer-Specific Histaminergic Modulation of Medial Entorhinal Cortex Required for Spatial Learning CEREBRAL CORTEX He, C., Luo, F., Chen, X., Chen, F., Li, C., Ren, S., Qiao, Q., Zhang, J., de Lecea, L., Gao, D., Hu, Z. 2016; 26 (4): 1590-1608

  Abstract

  The medial entorhinal cortex (MEC) plays a crucial role in spatial learning and memory. Whereas the MEC receives a dense histaminergic innervation from the tuberomamillary nucleus of the hypothalamus, the functions of histamine in this brain region remain unclear. Here, we show that histamine acts via H1Rs to directly depolarize the principal neurons in the superficial, but not deep, layers of the MEC when recording at somata. Moreover, histamine decreases the spontaneous GABA, but not glutamate, release onto principal neurons in the superficial layers by acting at presynaptic H3Rs without effect on synaptic release in the deep layers. Histamine-induced depolarization is mediated via inhibition of Kir channels and requires the activation of protein kinase C, whereas the inhibition of spontaneous GABA release by histamine depends on voltage-gated Ca(2+) channels and extracellular Ca(2+). Furthermore, microinjection of the H1R or H3R, but not H2R, antagonist respectively into the superficial, but not deep, layers of MEC impairs rat spatial learning as assessed by water maze tasks but does not affect the motor function and exploratory activity in an open field. Together, our study indicates that histamine plays an essential role in spatial learning by selectively regulating neuronal excitability and synaptic transmission in the superficial layers of the MEC.

  View details for DOI 10.1093/cercor/bhu322

  View details for Web of Science ID 000374246700021

  View details for PubMedID 25595181

• Resting easy with a sleep regulator ELIFE Giardino, W. J., de Lecea, L. 2015; 4

  View details for DOI 10.7554/eLife.12093

  View details for Web of Science ID 000367511500001

  View details for PubMedID 26651823

  View details for PubMedCentralID PMC4744186

• Sleep disruption impairs haematopoietic stem cell transplantation in mice NATURE COMMUNICATIONS Rolls, A., Pang, W. W., Ibarra, I., Colas, D., Bonnavion, P., Korin, B., Heller, H. C., Weissman, I. L., de Lecea, L. 2015; 6

  Abstract

  Many of the factors affecting the success of haematopoietic cell transplantation are still unknown. Here we show in mice that donor sleep deprivation reduces the ability of its haematopoietic stem cells (HSCs) to engraft and reconstitute the blood and bone marrow of an irradiated recipient by more than 50%. We demonstrate that sleep deprivation downregulates the expression of microRNA (miR)-19b, a negative regulator of the suppressor of cytokine signalling (SOCS) genes, which inhibit HSC migration and homing. Accordingly, HSCs from sleep-deprived mice have higher levels of SOCS genes expression, lower migration capacity in vitro and reduced homing to the bone marrow in vivo. Recovery of sleep after sleep deprivation restored the reconstitution potential of the HSCs. Taken together, this study provides insights into cellular and molecular mechanisms underlying the effects of sleep deprivation on HSCs, emphasizing the potentially critical role of donor sleep in the success of bone marrow transplantation.

  View details for DOI 10.1038/ncomms9516

  View details for Web of Science ID 000364930800001

  View details for PubMedID 26465715

  View details for PubMedCentralID PMC4621781

• Not So Giants: Mice Lacking Both Somatostatin and Cortistatin Have High GH Levels but Show No Changes in Growth Rate or IGF-1 Levels ENDOCRINOLOGY Pedraza-Arevalo, S., Cordoba-Chacon, J., Pozo-Salas, A. I., Lopez, F. L., De Lecea, L., Gahete, M. D., Castano, J. P., Luque, R. M. 2015; 156 (6): 1958-1964

  View details for DOI 10.1210/en.2015-1132

  View details for Web of Science ID 000358435400006

• Obesity Alters Gene Expression for GH/IGF-I Axis in Mouse Mammary Fat Pads: Differential Role of Cortistatin and Somatostatin PLOS ONE Villa-Osaba, A., Gahete, M. D., Cordoba-Chacon, J., de Lecea, L., Pozo-Salas, A. I., Javier Delgado-Lista, F., Alvarez-Benito, M., Lopez-Miranda, J., Luque, R. M., Castano, J. P. 2015; 10 (3)

  Abstract

  Locally produced growth hormone (GH) and IGF-I are key factors in the regulation of mammary gland (MG) development and may be important in breast cancer development/progression. Somatostatin (SST) and cortistatin (CORT) regulate GH/IGF-I axis at various levels, but their role in regulating GH/IGF-I in MGs remains unknown. Since obesity alters the expression of these systems in different tissues and is associated to MG (patho) physiology, we sought to investigate the role of SST/CORT in regulating GH/IGF-I system in the MGs of lean and obese mice. Therefore, we analyzed GH/IGF-I as well as SST/CORT and ghrelin systems expression in the mammary fat pads (MFPs) of SST- or CORT-knockout (KO) mice and their respective littermate-controls fed a low-fat (LF) or a high-fat (HF) diet for 16wks. Our results demonstrate that the majority of the components of GH/IGF-I, SST/CORT and ghrelin systems are locally expressed in mouse MFP. Expression of elements of the GH/IGF-I axis was significantly increased in MFPs of HF-fed control mice while lack of endogenous SST partially suppressed, and lack of CORT completely blunted, the up-regulation observed in obese WT-controls. Since SST/CORT are known to exert an inhibitory role on the GH/IGFI axis, the increase in SST/CORT-receptor sst2 expression in MFPs of HF-fed CORT- and SST-KOs together with an elevation on circulating SST in CORT-KOs could explain the differences observed. These results offer new information on the factors (GH/IGF-I axis) involved in the endocrine/metabolic dysregulation of MFPs in obesity, and suggest that CORT is not a mere SST sibling in regulating MG physiology.

  View details for DOI 10.1371/journal.pone.0120955

  View details for Web of Science ID 000351880000116

  View details for PubMedID 25806796

  View details for PubMedCentralID PMC4373840

• Antagonistic interplay between hypocretin and leptin in the lateral hypothalamus regulates stress responses NATURE COMMUNICATIONS Bonnavion, P., Jackson, A. C., Carter, M. E., de Lecea, L. 2015; 6

  Abstract

  The hypothalamic-pituitary-adrenal (HPA) axis functions to coordinate behavioural and physiological responses to stress in a manner that depends on the behavioural state of the organism. However, the mechanisms through which arousal and metabolic states influence the HPA axis are poorly understood. Here using optogenetic approaches in mice, we show that neurons that produce hypocretin (Hcrt)/orexin in the lateral hypothalamic area (LHA) regulate corticosterone release and a variety of behaviours and physiological hallmarks of the stress response. Interestingly, we found that Hcrt neuronal activity and Hcrt-mediated stress responses were inhibited by the satiety hormone leptin, which acts, in part, through a network of leptin-sensitive neurons in the LHA. These data demonstrate how peripheral metabolic signals interact with hypothalamic neurons to coordinate stress and arousal and suggest one mechanism through which hyperarousal or altered metabolic states may be linked with abnormal stress responses.

  View details for DOI 10.1038/ncomms7266

  View details for Web of Science ID 000350202800016

  View details for PubMedID 25695914

  View details for PubMedCentralID PMC4335349

• Optogenetics in Freely Moving Mammals: Dopamine and Reward. Cold Spring Harbor protocols Zhang, F., Tsai, H., Airan, R. D., Stuber, G. D., Adamantidis, A. R., de Lecea, L., Bonci, A., Deisseroth, K. 2015; 2015 (8): pdb top086330-?

  Abstract

  Brain reward systems play a central role in the cognitive and hedonic behaviors of mammals. Multiple neuron types and brain regions are involved in reward processing, posing fascinating scientific questions, and major experimental challenges. Using diverse approaches including genetics, electrophysiology, imaging, and behavioral analysis, a large body of research has focused on both normal functioning of the reward circuitry and on its potential significance in neuropsychiatric diseases. In this introduction, we illustrate a real-world application of optogenetics to mammalian behavior and physiology, delineating procedures and technologies for optogenetic control of individual components of the reward circuitry. We describe the experimental setup and protocol for integrating optogenetic modulation of dopamine neurons with fast-scan cyclic voltammetry, conditioned place preference, and operant conditioning to assess the causal role of well-defined electrical and biochemical signals in reward-related behavior.

  View details for DOI 10.1101/pdb.top086330

  View details for PubMedID 26240415

• Optogenetic control of hypocretin (orexin) neurons and arousal circuits. Current topics in behavioral neurosciences de Lecea, L. n. 2015; 25: 367–78

  Abstract

  In 1998, our group discovered a cDNA that encoded the precursor of two putative neuropeptides that we called hypocretins for their hypothalamic expression and their similarity to the secretin family of neuropeptides. In the last 16 years, numerous studies have placed the hypocretin system as an integrator of homeostatic functions with a crucial, non-redundant function as arousal stabilizer. We recently applied optogenetic methods to interrogate the role of individual neuronal circuits in sleep-to-wake transitions. The neuronal connections between the hypocretin system and the locus coeruleus (LC) seem to be crucial in establishing the appropriate dynamic of spontaneous awakenings.

  View details for PubMedID 25502546

  View details for PubMedCentralID PMC5047484

• Optogenetic Dissection of Neural Circuit Function in Behaving Animals NEURAL TRACING METHODS: TRACING NEURONS AND THEIR CONNECTIONS Herrera, C., Adamantidis, A., Zhang, F., Deisseroth, K., de Lecea, L., Arenkiel, B. R. 2015; 92: 143–60

  View details for DOI 10.1007/978-1-4939-1963-5_7

  View details for Web of Science ID 000362844900009

• A Framework for Quantitative Modeling of Neural Circuits Involved in Sleep-to-Wake Transition. Frontiers in neurology Sorooshyari, S., Huerta, R., de Lecea, L. 2015; 6: 32-?

  Abstract

  Identifying the neuronal circuits and dynamics of sleep-to-wake transition is essential to understanding brain regulation of behavioral states, including sleep-wake cycles, arousal, and hyperarousal. Recent work by different laboratories has used optogenetics to determine the role of individual neuromodulators in state transitions. The optogenetically driven data do not yet provide a multi-dimensional schematic of the mechanisms underlying changes in vigilance states. This work presents a modeling framework to interpret, assist, and drive research on the sleep-regulatory network. We identify feedback, redundancy, and gating hierarchy as three fundamental aspects of this model. The presented model is expected to expand as additional data on the contribution of each transmitter to a vigilance state becomes available. Incorporation of conductance-based models of neuronal ensembles into this model and existing models of cortical excitability will provide more comprehensive insight into sleep dynamics as well as sleep and arousal-related disorders.

  View details for DOI 10.3389/fneur.2015.00032

  View details for PubMedID 25767461

  View details for PubMedCentralID PMC4341569

• The Hypocretin/Orexin System: An Increasingly Important Role in Neuropsychiatry MEDICINAL RESEARCH REVIEWS Chen, Q., de Lecea, L., Hu, Z., Gao, D. 2015; 35 (1): 152-197

  Abstract

  Hypocretins, also named as orexins, are excitatory neuropeptides secreted by neurons specifically located in lateral hypothalamus and perifornical areas. Orexinergic fibers are extensively distributed in various brain regions and involved in a number of physiological functions, such as arousal, cognition, stress, appetite, and metabolism. Arousal is the most important function of orexin system as dysfunction of orexin signaling leads to narcolepsy. In addition to narcolepsy, orexin dysfunction is associated with serious neural disorders, including addiction, depression, and anxiety. However, some results linking orexin with these disorders are still contradictory, which may result from differences of detection methods or the precision of tools used in measurements; strategies targeted to orexin system (e.g., antagonists to orexin receptors, gene delivery, and cell transplantation) are promising new tools for treatment of neuropsychiatric disorders, though studies are still in a stage of preclinical or clinical research.

  View details for DOI 10.1002/med.21326

  View details for Web of Science ID 000346080200005

  View details for PubMedID 25044006

• Potential role of orexin and sleep modulation in the pathogenesis of Alzheimer's disease JOURNAL OF EXPERIMENTAL MEDICINE Roh, J. H., Jiang, H., Finn, M. B., Stewart, F. R., Mahan, T. E., Cirrito, J. R., Heda, A., Snider, B. J., Li, M., Yanagisawa, M., de Lecea, L., Holtzman, D. M. 2014; 211 (13): 2487-2496

  Abstract

  Age-related aggregation of amyloid-β (Aβ) is an upstream pathological event in Alzheimer's disease (AD) pathogenesis, and it disrupts the sleep-wake cycle. The amount of sleep declines with aging and to a greater extent in AD. Poor sleep quality and insufficient amounts of sleep have been noted in humans with preclinical evidence of AD. However, how the amount and quality of sleep affects Aβ aggregation is not yet well understood. Orexins (hypocretins) initiate and maintain wakefulness, and loss of orexin-producing neurons causes narcolepsy. We tried to determine whether orexin release or secondary changes in sleep via orexin modulation affect Aβ pathology. Amyloid precursor protein (APP)/Presenilin 1 (PS1) transgenic mice, in which the orexin gene is knocked out, showed a marked decrease in the amount of Aβ pathology in the brain with an increase in sleep time. Focal overexpression of orexin in the hippocampus in APP/PS1 mice did not alter the total amount of sleep/wakefulness and the amount of Aβ pathology. In contrast, sleep deprivation or increasing wakefulness by rescue of orexinergic neurons in APP/PS1 mice lacking orexin increased the amount of Aβ pathology in the brain. Collectively, modulation of orexin and its effects on sleep appear to modulate Aβ pathology in the brain.

  View details for DOI 10.1084/jem.20141788

  View details for Web of Science ID 000346366100001

  View details for PubMedID 25422493

  View details for PubMedCentralID PMC4267230

• Hypocretin (orexin) neuromodulation of stress and reward pathways CURRENT OPINION IN NEUROBIOLOGY Giardino, W. J., de Lecea, L. 2014; 29: 103-108

  Abstract

  Hypocretin (also known as orexin) is a peptide neuromodulator that is expressed exclusively in the lateral hypothalamic area and plays a fundamental role in wakefulness and arousal. Chronic stress and compulsive drug-seeking are two examples of dysregulated states of hyperarousal that are influenced by hypocretin transmission throughout hypothalamic, extended amygdala, brainstem, and mesolimbic pathways. Here, we review current advances in the understanding of hypocretin's modulatory actions underlying conditions of negative and positive emotional valence, focusing particularly on mechanisms that facilitate adaptive (and maladaptive) responses to stressful or rewarding environmental stimuli. We conclude by discussing progress toward integrated theories for hypocretin modulation of divergent behavioral domains.

  View details for DOI 10.1016/j.conb.2014.07.006

  View details for Web of Science ID 000347128200015

  View details for PubMedID 25050887

  View details for PubMedCentralID PMC4267967

• Basal Forebrain Cholinergic Modulation of Sleep Transitions SLEEP Irmak, S. O., de Lecea, L. 2014; 37 (12): 1941-U104

  Abstract

  The basal forebrain cholinergic system is involved in cognitive processes that require an attentive state, an increased level of arousal, and/ or cortical activation associated with low amplitude fast EEG activity. The activity of most neurons in the basal forebrain cholinergic space is tightly correlated with the cortical EEG and the activity state. While most cholinergic neurons fire maximally during waking and REM sleep, the activity of other types of basal forebrain neurons vastly differs across different arousal and sleep states. Numerous studies have suggested a role for the basal forebrain cholinergic neurons in eliciting cortical activation and arousal. However, the intricate local connectivity within the region requires the use of cell-specific manipulation methods to demonstrate such a causal relationship.Here we have combined optogenetics with surface EEG recordings in freely moving mice in order to investigate the effects of acute cholinergic activation on the dynamics of sleep-to-wake transitions. We recorded from naturally sleeping animals and analyzed transitions from NREM sleep to REM sleep and/ or wakefulness in response to photo-stimulation of cholinergic neurons in substantia innominata.Our results show that optogenetic activation of BF cholinergic neurons during NREM sleep is sufficient to elicit cortical activation and facilitate state transitions, particularly transitions to wakefulness and arousal, at a time scale similar to the activation induced by other subcortical systems. Our results provide in vivo cell-specific demonstration for the role of basal forebrain cholinergic system in induction of wakefulness and arousal.

  View details for DOI 10.5665/sleep.4246

  View details for Web of Science ID 000345827600011

  View details for PubMedID 25325504

• Control of sleep-to-wake transitions via fast amino acid and slow neuropeptide transmission NEW JOURNAL OF PHYSICS Mosqueiro, T., de Lecea, L., Huerta, R. 2014; 16

  View details for DOI 10.1088/1367-2630/16/11/115010

  View details for Web of Science ID 000346761400001

• Light and chemical control of neuronal circuits: possible applications in neurotherapy. Expert review of neurotherapeutics Whittle, A. J., Walsh, J., de Lecea, L. 2014; 14 (9): 1007-1017

  Abstract

  Millions of people worldwide suffer from diseases that result from a failure of central pathways to regulate behavioral and physiological processes. Advances in genetics and pharmacology have already allowed us to appreciate that rather than this dysregulation being systemic throughout the brain, it is usually rooted in specific subsets of dysfunctional cells within discrete neurological circuits. This article discusses the advent of opto- and chemogenetic tools and how they are providing the means to dissect these circuits with a degree of temporal and spatial sensitivity not previously possible. We also highlight the potential applications for treating disease and the key developments likely to have the greatest impact over the next 5 years.

  View details for DOI 10.1586/14737175.2014.948850

  View details for PubMedID 25115180

• Optogenetic control of hypocretin/orexin neurons de Lecea, L. AMER CHEMICAL SOC. 2014

  View details for Web of Science ID 000349167402135

• Lack of Endogenous Cortistatin but Not Somatostatin Exacerbates Carcinogen-Induced Mammary Gland Tumorigenesis in Mice Gahete, M. D., Villa-Osaba, A., Cordoba-Chacon, J., de Lecea, L., Gracia-Navarro, F., Gonzalez-Rey, E., Luque, R. M., Casta, J. P. ENDOCRINE SOC. 2014

  View details for Web of Science ID 000209805108106

• Cortistatin Is a Key Factor Regulating the Gender-Dependent Response of the Growth Hormone (GH) and Adrenocorticotropin (ACTH) Axes to Fasting in Mice Cordoba-Chacon, J., Gahete, M. D., Pozo-Salas, A., de Lecea, L., Castaao, J. P., Luque, R. M. ENDOCRINE SOC. 2014

  View details for Web of Science ID 000209805101298

• Hypocretin (orexin) regulation of sleep-to-wake transitions FRONTIERS IN PHARMACOLOGY de Lecea, L., Huerta, R. 2014; 5

  Abstract

  The hypocretin (Hcrt), also known as orexin, peptides are essential for arousal stability. Here we discuss background information about the interaction of Hcrt with other neuromodulators, including norepinephrine and acetylcholine probed with optogenetics. We conclude that Hcrt neurons integrate metabolic, circadian and limbic inputs and convey this information to a network of neuromodulators, each of which has a different role on the dynamic of sleep-to-wake transitions. This model may prove useful to predict the effects of orexin receptor antagonists in sleep disorders and other conditions.

  View details for DOI 10.3389/fphar.2014.00016

  View details for Web of Science ID 000347042700001

  View details for PubMedID 24575043

  View details for PubMedCentralID PMC3921570

• Establishing a fiber-optic-based optical neural interface. Cold Spring Harbor protocols Adamantidis, A. R., Zhang, F., de Lecea, L., Deisseroth, K. 2014; 2014 (8): pdb prot083337-?

  Abstract

  Selective expression of opsins in genetically defined neurons makes it possible to control a subset of neurons without affecting nearby cells and processes in the intact brain, but light must still be delivered to the target brain structure. Light scattering limits the delivery of light from the surface of the brain. For this reason, we have developed a fiber-optic-based optical neural interface (ONI), which allows optical access to any brain structure in freely moving mammals. The ONI system is constructed by modifying the small animal cannula system from PlasticsOne. The system for bilateral stimulation consists of a bilateral cannula guide that has been stereotactically implanted over the target brain region, a screw cap for securing the optical fiber to the animal's head, a fiber guard modified from the internal cannula adapter, and a bare fiber whose length is customized based on the depth of the target region. For unilateral stimulation, a single-fiber system can be constructed using unilateral cannula parts from PlasticsOne. We describe here the preparation of the bilateral ONI system and its use in optical stimulation of the mouse or rat brain. Delivery of opsin-expressing virus and implantation of the ONI may be conducted in the same surgical session; alternatively, with a transgenic animal no opsin virus is delivered during the surgery. Similar procedures are useful for deep or superficial injections (even for neocortical targets, although in some cases surface light-emitting diodes or cortex-apposed fibers can be used for the most superficial cortical targets).

  View details for DOI 10.1101/pdb.prot083337

  View details for PubMedID 25086020

• Control of sleep-to-wake transitions via fast aminoacid and slow neuropeptide transmission. New journal of physics Mosqueiro, T. n., de Lecea, L. n., Huerta, R. n. 2014; 16

  Abstract

  The Locus Coeruleus (LC) modulates cortical, subcortical, cerebellar, brainstem and spinal cord circuits and it expresses receptors for neuromodulators that operate in a time scale of several seconds. Evidences from anatomical, electrophysiological and optogenetic experiments have shown that LC neurons receive input from a group of neurons called Hypocretins (HCRTs) that release a neuropeptide called hypocretin. It is less known how these two groups of neurons can be coregulated using GABAergic neurons. Since the time scales of GABA A inhibition is several orders of magnitude faster than the hypocretin neuropeptide effect, we investigate the limits of circuit activity regulation using a realistic model of neurons. Our investigation shows that GABA A inhibition is insufficient to control the activity levels of the LCs. Despite slower forms of GABA A can in principle work, there is not much plausibility due to the low probability of the presence of slow GABA A and lack of robust stability at the maximum firing frequencies. The best possible control mechanism predicted by our modeling analysis is the presence of inhibitory neuropeptides that exert effects in a similar time scale as the hypocretin/orexin. Although the nature of these inhibitory neuropeptides has not been identified yet, it provides the most efficient mechanism in the modeling analysis. Finally, we present a reduced mean-field model that perfectly captures the dynamics and the phenomena generated by this circuit. This investigation shows that brain communication involving multiple time scales can be better controlled by employing orthogonal mechanisms of neural transmission to decrease interference between cognitive processes and hypothalamic functions.

  View details for PubMedID 25598695

  View details for PubMedCentralID PMC4292803

• The hypocretins/orexins: integrators of multiple physiological functions BRITISH JOURNAL OF PHARMACOLOGY Li, J., Hu, Z., de Lecea, L. 2014; 171 (2): 332-350

  Abstract

  The hypocretins (Hcrts), also known as orexins, are two peptides derived from a single precursor produced in the posterior lateral hypothalamus. Over the past decade, the orexin system has been associated with numerous physiological functions, including sleep/arousal, energy homeostasis, endocrine, visceral functions and pathological states, such as narcolepsy and drug abuse. Here, we review the discovery of Hcrt/orexins and their receptors and propose a hypothesis as to how the orexin system orchestrates these multifaceted physiological functions.

  View details for DOI 10.1111/bph.12415

  View details for Web of Science ID 000328712000005

  View details for PubMedID 24102345

• Optogenetics: opsins and optical interfaces in neuroscience. Cold Spring Harbor protocols Adamantidis, A. R., Zhang, F., de Lecea, L., Deisseroth, K. 2014; 2014 (8): pdb top083329-?

  Abstract

  Optogenetics is defined as the integration of optics and genetics to control well-defined events within specified cells of living tissue. In this introduction, we focus on the basic techniques necessary for employing microbial opsins as optogenetic tools in mammalian brains. We provide a guide for the fundamentals of optogenetic application-selecting an opsin, implementing expression of opsins based on the neuroscientific experimental requirements, and adapting the corresponding optical hardware for delivery of light into mammalian brains.

  View details for DOI 10.1101/pdb.top083329

  View details for PubMedID 25086025

• Orexin/hypocretin system modulates amygdala-dependent threat learning through the locus coeruleus PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Sears, R. M., Fink, A. E., Wigestrand, M. B., Farb, C. R., de Lecea, L., LeDoux, J. E. 2013; 110 (50): 20260-20265

  Abstract

  Survival in a dangerous environment requires learning about stimuli that predict harm. Although recent work has focused on the amygdala as the locus of aversive memory formation, the hypothalamus has long been implicated in emotional regulation, and the hypothalamic neuropeptide orexin (hypocretin) is involved in anxiety states and arousal. Nevertheless, little is known about the role of orexin in aversive memory formation. Using a combination of behavioral pharmacology, slice physiology, and optogenetic techniques, we show that orexin acts upstream of the amygdala via the noradrenergic locus coeruleus to enable threat (fear) learning, specifically during the aversive event. Our results are consistent with clinical studies linking orexin levels to aversive learning and anxiety in humans and dysregulation of the orexin system may contribute to the etiology of fear and anxiety disorders.

  View details for DOI 10.1073/pnas.1320325110

  View details for Web of Science ID 000328061700069

  View details for PubMedID 24277819

  View details for PubMedCentralID PMC3864341

• Sleep to forget: interference of fear memories during sleep. Molecular psychiatry Rolls, A., Makam, M., Kroeger, D., Colas, D., De Lecea, L., Heller, H. C. 2013; 18 (11): 1166-1170

  Abstract

  Memories are consolidated and strengthened during sleep. Here we show that memories can also be weakened during sleep. We used a fear-conditioning paradigm in mice to condition footshock to an odor (conditioned stimulus (CS)). Twenty-four hours later, presentation of the CS odor during sleep resulted in an enhanced fear response when tested during subsequent wake. However, if the re-exposure of the CS odor during sleep was preceded by bilateral microinjections of a protein synthesis inhibitor into the basolateral amygdala, the subsequent fear response was attenuated. These findings demonstrate that specific fear memories can be selectively reactivated and either strengthened or attenuated during sleep, suggesting the potential for developing sleep therapies for emotional disorders.

  View details for DOI 10.1038/mp.2013.121

  View details for PubMedID 24081009

  View details for PubMedCentralID PMC5036945

• Paradoxical Effect of Cortistatin Treatment and Its Deficiency on Experimental Autoimmune Encephalomyelitis JOURNAL OF IMMUNOLOGY Souza-Moreira, L., Morell, M., Delgado-Maroto, V., Pedreno, M., Martinez-Escudero, L., Caro, M., O'Valle, F., Luque, R., Gallo, M., de Lecea, L., Castano, J. P., Gonzalez-Rey, E. 2013; 191 (5): 2144-2154

  Abstract

  Cortistatin is a cyclic-neuropeptide produced by brain cortex and immune cells that shows potent anti-inflammatory activity. In this article, we investigated the effect of cortistatin in two models of experimental autoimmune encephalomyelitis (EAE) that mirror chronic and relapsing-remitting multiple sclerosis. A short-term systemic treatment with cortistatin reduced clinical severity and incidence of EAE, the appearance of inflammatory infiltrates in spinal cord, and the subsequent demyelination and axonal damage. This effect was associated with a reduction of the two deleterious components of the disease, namely, the autoimmune and inflammatory response. Cortistatin decreased the presence/activation of encephalitogenic Th1 and Th17 cells in periphery and nervous system, and downregulated various inflammatory mediators, whereas it increased the number of regulatory T cells with suppressive effects on the encephalitogenic response. Moreover, cortistatin regulated glial activity and favored an active program of neuroprotection/regeneration. We further used cortistatin-deficient mice to investigate the role of endogenous cortistatin in the control of immune responses. Surprisingly, cortistatin-deficient mice were partially resistant to EAE and other inflammatory disorders, despite showing competent inflammatory/autoreactive responses. This unexpected phenotype was associated with elevated circulating glucocorticoids and an anxiety-like behavior. Our findings provide a powerful rationale for the assessment of the efficacy of cortistatin as a novel multimodal therapeutic approach to treat multiple sclerosis and identify cortistatin as a key endogenous component of neuroimmune system.

  View details for DOI 10.4049/jimmunol.1300384

  View details for Web of Science ID 000323393300015

  View details for PubMedID 23918980

• Optogenetics in psychiatric diseases. Current opinion in neurobiology Touriño, C., Eban-Rothschild, A., de Lecea, L. 2013; 23 (3): 430-435

  Abstract

  Optogenetic tools have revolutionized the field of neuroscience, and brought the study of neural circuits to a higher level. Optogenetics has significantly improved our understanding not only of the neuronal connections and function of the healthy brain, but also of the neuronal changes that lead to psychiatric disorders. In this review, we summarize recent optogenetic studies that explored different brain circuits involved in natural behaviors, such as sleep and arousal, reward, fear, and social and aggressive behavior. In addition, we describe how alterations in these circuits may lead to psychiatric disorders such as addiction, anxiety, depression, or schizophrenia.

  View details for DOI 10.1016/j.conb.2013.03.007

  View details for PubMedID 23642859

• Cortistatin Inhibits Migration and Proliferation of Human Vascular Smooth Muscle Cells and Decreases Neointimal Formation on Carotid Artery Ligation CIRCULATION RESEARCH Duran-Prado, M., Morell, M., Delgado-Maroto, V., Castano, J. P., Aneiros-Fernandez, J., de Lecea, L., Culler, M. D., Hernandez-Cortes, P., O'Valle, F., Delgado, M. 2013; 112 (11): 1444-?

  Abstract

  Proliferation and migration of smooth muscle cells (SMCs) are key steps for the progression of atherosclerosis and restenosis. Cortistatin is a multifunctional neuropeptide belonging to the somatostatin family that exerts unique functions in the nervous and immune systems. Cortistatin is elevated in plasma of patients experiencing coronary heart disease and attenuates vascular calcification.To investigate the occurrence of vascular cortistatin and its effects on the proliferation and migration of SMCs in vitro and in vivo and to delimitate the receptors and signal transduction pathways governing its actions.SMCs from mouse carotid and human aortic arteries and from human atherosclerotic plaques highly expressed cortistatin. Cortistatin expression positively correlated with the progression of arterial intima hyperplasia. Cortistatin inhibited platelet-derived growth factor-stimulated proliferation of human aortic SMCs via binding to somatostatin receptors (sst2 and sst5) and ghrelin receptor, induction of cAMP and p38-mitogen-activated protein kinase, and inhibition of Akt activity. Moreover, cortistatin impaired lamellipodia formation and migration of human aortic SMCs toward platelet-derived growth factor by inhibiting, in a ghrelin-receptor-dependent manner, Rac1 activation and cytosolic calcium increases. These effects on SMC proliferation and migration correlated with an inhibitory action of cortistatin on the neointimal formation in 2 models of carotid arterial ligation. Endogenous cortistatin seems to play a critical role in regulating SMC function because cortistatin-deficient mice developed higher neointimal hyperplasic lesions than wild-type mice.Cortistatin emerges as a natural endogenous regulator of SMCs under pathological conditions and an attractive candidate for the pharmacological management of vascular diseases that course with neointimal lesion formation.

  View details for DOI 10.1161/CIRCRESAHA.112.300695

  View details for Web of Science ID 000319448900013

  View details for PubMedID 23595952

• Functional wiring of hypocretin and LC-NE neurons: implications for arousal FRONTIERS IN BEHAVIORAL NEUROSCIENCE Carter, M. E., de Lecea, L., Adamantidis, A. 2013; 7

  Abstract

  To survive in a rapidly changing environment, animals must sense their external world and internal physiological state and properly regulate levels of arousal. Levels of arousal that are abnormally high may result in inefficient use of internal energy stores and unfocused attention to salient environmental stimuli. Alternatively, levels of arousal that are abnormally low may result in the inability to properly seek food, water, sexual partners, and other factors necessary for life. In the brain, neurons that express hypocretin neuropeptides may be uniquely posed to sense the external and internal state of the animal and tune arousal state according to behavioral needs. In recent years, we have applied temporally precise optogenetic techniques to study the role of these neurons and their downstream connections in regulating arousal. In particular, we have found that noradrenergic neurons in the brainstem locus coeruleus (LC) are particularly important for mediating the effects of hypocretin neurons on arousal. Here, we discuss our recent results and consider the implications of the anatomical connectivity of these neurons in regulating the arousal state of an organism across various states of sleep and wakefulness.

  View details for DOI 10.3389/fnbeh.2013.00043

  View details for Web of Science ID 000319055000001

  View details for PubMedID 23730276

  View details for PubMedCentralID PMC3657625

• Hypothalamic Neurotensin Projections Promote Reward by Enhancing Glutamate Transmission in the VTA JOURNAL OF NEUROSCIENCE Kempadoo, K. A., Tourino, C., Cho, S. L., Magnani, F., Leinninger, G., Stuber, G. D., Zhang, F., Myers, M. G., Deisseroth, K., de Lecea, L., Bonci, A. 2013; 33 (18): 7618-?

  Abstract

  The lateral hypothalamus (LH) sends a dense glutamatergic and peptidergic projection to dopamine neurons in the ventral tegmental area (VTA), a cell group known to promote reinforcement and aspects of reward. The role of the LH to VTA projection in reward-seeking behavior can be informed by using optogenetic techniques to dissociate the actions of LH neurons from those of other descending forebrain inputs to the VTA. In the present study, we identify the effect of neurotensin (NT), one of the most abundant peptides in the LH to VTA projection, on excitatory synaptic transmission in the VTA and reward-seeking behavior. Mice displayed robust intracranial self-stimulation of LH to VTA fibers, an operant behavior mediated by NT 1 receptors (Nts1) and NMDA receptors. Whole-cell patch-clamp recordings of VTA dopamine neurons demonstrated that NT (10 nm) potentiated NMDA-mediated EPSCs via Nts1. Results suggest that NT release from the LH into the VTA activates Nts1, thereby potentiating NMDA-mediated EPSCs and promoting reward. The striking behavioral and electrophysiological effects of NT and glutamate highlight the LH to VTA pathway as an important component of reward.

  View details for DOI 10.1523/JNEUROSCI.2588-12.2013

  View details for Web of Science ID 000318420400002

  View details for PubMedID 23637156

• Repeated in vivo exposure of cocaine induces long-lasting synaptic plasticity in hypocretin/orexin-producing neurons in the lateral hypothalamus in mice JOURNAL OF PHYSIOLOGY-LONDON Rao, Y., Mineur, Y. S., Gan, G., Wang, A. H., Liu, Z., Wu, X., Suyama, S., de Lecea, L., Horvath, T. L., Picciotto, M. R., Gao, X. 2013; 591 (7): 1951-1966

  Abstract

  Hypocretin (orexin), a neuropeptide synthesized exclusively in the perifornical/lateral hypothalamus, is critical for drug seeking and relapse, but it is not clear how the circuitry centred on hypocretin-producing neurons (hypocretin neurons) is modified by drugs of abuse and how changes in this circuit might alter behaviours related to drug addiction. In this study, we show that repeated, but not single, in vivo cocaine administration leads to a long-lasting, experience-dependent potentiation of glutamatergic synapses on hypocretin neurons in mice following a cocaine-conditioned place preference (CPP) protocol. The synaptic potentiation occurs postsynaptically and probably involves up-regulation of AMPA-type glutamate receptors on hypocretin neurons. Phosphorylation of cAMP response element-binding protein (CREB) is also significantly increased in hypocretin neurons in cocaine-treated animals, suggesting that CREB-mediated pathways may contribute to synaptic potentiation in these cells. Furthermore, the potentiation of synaptic efficacy in hypocretin neurons persists during cocaine withdrawal, but reverses to baseline levels after prolonged abstinence. Finally, the induction of long-term potentiation (LTP) triggered by a high-frequency stimulation is facilitated in hypocretin neurons in cocaine-treated mice, suggesting that long-lasting changes in synapses onto hypocretin neurons would probably be further potentiated by other stimuli (such as concurrent environmental cues) paired with the drug. In summary, we show here that hypocretin neurons undergo experience-dependent synaptic potentiation that is distinct from that reported in other reward systems, such as the ventral tegmental area, following exposure to cocaine. These findings support the idea that the hypocretin system is important for behavioural changes associated with cocaine administration in animals and humans.

  View details for DOI 10.1113/jphysiol.2012.246983

  View details for Web of Science ID 000316918300030

  View details for PubMedID 23318871

  View details for PubMedCentralID PMC3624862

• Optogenetic control of arousal neurons GENETIC BASIS OF SLEEP AND SLEEP DISORDERS Adamantidis, A., Carter, M. E., de Lecea, L., Shaw, P., Tafti, M., Thorpy, M. 2013: 66–72

  View details for Web of Science ID 000325766000009

• Mechanism for Hypocretin-mediated sleep-to-wake transitions PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Carter, M. E., Brill, J., Bonnavion, P., Huguenard, J. R., Huerta, R., de Lecea, L. 2012; 109 (39): E2635-E2644

  Abstract

  Current models of sleep/wake regulation posit that Hypocretin (Hcrt)-expressing neurons in the lateral hypothalamus promote and stabilize wakefulness by projecting to subcortical arousal centers. However, the critical downstream effectors of Hcrt neurons are unknown. Here we use optogenetic, pharmacological, and computational tools to investigate the functional connectivity between Hcrt neurons and downstream noradrenergic neurons in the locus coeruleus (LC) during nonrapid eye movement (NREM) sleep. We found that photoinhibiting LC neurons during Hcrt stimulation blocked Hcrt-mediated sleep-to-wake transitions. In contrast, when LC neurons were optically stimulated to increase membrane excitability, concomitant photostimulation of Hcrt neurons significantly increased the probability of sleep-to-wake transitions compared with Hcrt stimulation alone. We also built a conductance-based computational model of Hcrt-LC circuitry that recapitulates our behavioral results using LC neurons as the main effectors of Hcrt signaling. These results establish the Hcrt-LC connection as a critical integrator-effector circuit that regulates NREM sleep/wake behavior during the inactive period. This coupling of distinct neuronal systems can be generalized to other hypothalamic integrator nuclei with downstream effector/output populations in the brain.

  View details for DOI 10.1073/pnas.1202526109

  View details for Web of Science ID 000309604500009

  View details for PubMedID 22955882

  View details for PubMedCentralID PMC3465396

• Shining Light on Wakefulness and Arousal BIOLOGICAL PSYCHIATRY de Lecea, L., Carter, M. E., Adamantidis, A. 2012; 71 (12): 1046-1052

  Abstract

  Alterations in arousal states are associated with multiple neuropsychiatric disorders, including generalized anxiety disorders, addiction, schizophrenia, and depression. Therefore, elucidating the neurobiological mechanisms controlling the boundaries between arousal, hyperarousal, and hypoarousal is a crucial endeavor in biological psychiatry. Substantial research over several decades has identified distinct arousal-promoting neural populations in the brain; however, how these nuclei act individually and collectively to promote and maintain wakefulness and various arousal states is unknown. We have recently applied optogenetic technology to the repertoire of techniques used to study arousal. Here, we discuss the recent results of these experiments and propose future use of this approach as a way to understand the complex dynamics of neural circuits controlling arousal and arousal-related behaviors.

  View details for DOI 10.1016/j.biopsych.2012.01.032

  View details for Web of Science ID 000304471300005

  View details for PubMedID 22440618

• Hypocretins and the neurobiology of sleep-wake mechanisms OREXIN/HYPOCRETIN SYSTEM de Lecea, L. 2012; 198: 15-24

  Abstract

  In 1998, our group discovered a cDNA that encoded the precursor of two putative neuropeptides that we called hypocretins for their hypothalamic expression and their similarity to the secretin family of neuropeptides. In the past 15 years, numerous studies have placed the hypocretin system as an integrator of homeostatic functions with a crucial, nonredundant function as an arousal stabilizer. Here, we discuss some of the data that have accumulated over the years on the integrating capacity of these hypothalamic neurons and their role on sleep-to-wake transitions.

  View details for DOI 10.1016/B978-0-444-59489-1.00003-3

  View details for Web of Science ID 000311106700004

  View details for PubMedID 22813967

• Relationships between sleep and addiction ADICCIONES Canellas, F., de Lecea, L. 2012; 24 (4): 287-290

  Abstract

  While it is well known that there is an interaction between sleep disorders and substance abuse, it is certainly more complex than was previously thought. There is a positive relationship both between having a substance use disorder and suffering from a sleep disorder, and vice versa. The effects on sleep depend on the substance used, but it has been shown that both during use and in withdrawal periods consumers have various sleep problems, and basically more fragmented sleep. We know that sleep problems must be taken into account to prevent addiction relapses. Recent research shows that the hypocretinergic system defined by neuropeptide hypocretin / orexin (Hcrt / ox), located in the lateral hypothalamus and involved in, among other things, the regulation of the sleep-wake cycle, may play an important role in addictive behaviors. Different studies have demonstrated interactions between the hypocretinergic system, acute response to stress circuits and reward systems. We also know that selective optogenetic activation of the hypocretinergic system increases the probability of transition from sleep to wakefulness, and is sufficient for initiating an addictive compulsive behavior relapse. Hypocretinergic system activation could explain the hyperarousal associated with stress and addiction. Improved knowledge of this interaction would help us to understand better the mechanisms of addiction and find new strategies for the treatment of addictions.

  View details for DOI 10.20882/adicciones.78

  View details for Web of Science ID 000311766800002

  View details for PubMedID 23241715

  View details for PubMedCentralID PMC5047372

• Cortistatin Is Not a Somatostatin Analogue but Stimulates Prolactin Release and Inhibits GH and ACTH in a Gender-Dependent Fashion: Potential Role of Ghrelin ENDOCRINOLOGY Cordoba-Chacon, J., Gahete, M. D., Pozo-Salas, A. I., Martinez-Fuentes, A. J., de Lecea, L., Gracia-Navarro, F., Kineman, R. D., Castano, J. P., Luque, R. M. 2011; 152 (12): 4800-4812

  Abstract

  Cortistatin (CST) and somatostatin (SST) evolve from a common ancestral gene and share remarkable structural, pharmacological, and functional homologies. Although CST has been considered as a natural SST-analogue acting through their shared receptors (SST receptors 1-5), emerging evidence indicates that these peptides might in fact exert unique roles via selective receptors [e.g. CST, not SST, binds ghrelin receptor growth hormone secretagogue receptor type 1a (GHS-R1a)]. To determine whether the role of endogenous CST is different from SST, we characterized the endocrine-metabolic phenotype of male/female CST null mice (cort-/-) at hypothalamic-pituitary-systemic (pancreas-stomach-adrenal-liver) levels. Also, CST effects on hormone expression/secretion were evaluated in primary pituitary cell cultures from male/female mice and female primates (baboons). Specifically, CST exerted an unexpected stimulatory role on prolactin (PRL) secretion, because both male/female cort-/- mice had reduced PRL levels, and CST treatment (in vivo and in vitro) increased PRL secretion, which could be blocked by a GHS-R1a antagonist in vitro and likely relates to the decreased success of female cort-/- in first-litter pup care at weaning. In contrast, CST inhibited GH and adrenocorticotropin-hormone axes in a gender-dependent fashion. In addition, a rise in acylated ghrelin levels was observed in female cort-/- mice, which were associated with an increase in stomach ghrelin/ghrelin O-acyl transferase expression. Finally, CST deficit uncovered a gender-dependent role of this peptide in the regulation of glucose-insulin homeostasis, because male, but not female, cort-/- mice developed insulin resistance. The fact that these actions are not mimicked by SST and are strongly gender dependent offers new grounds to investigate the hitherto underestimated physiological relevance of CST in the regulation of physiological/metabolic processes.

  View details for DOI 10.1210/en.2011-1542

  View details for Web of Science ID 000297376500034

  View details for PubMedID 21971153

  View details for PubMedCentralID PMC3230064

• Plasma levels of neuropeptides and metabolic hormones, and sleepiness in obstructive sleep apnea RESPIRATORY MEDICINE Sanchez-de-la-Torre, M., Barcelo, A., Pierola, J., Esquinas, C., de la Pena, M., Duran-Cantolla, J., Capote, F., Masa, J. F., Marin, J. M., Vila, M., Cao, G., Martinez, M., de Lecea, L., Gozal, D., Montserrat, J. M., Barbe, F. 2011; 105 (12): 1954-1960

  Abstract

  Obstructive sleep apnea (OSA) is related to obesity and metabolic disorders. The main clinical symptoms are excessive daytime sleepiness (EDS) and snoring. However, not all patients with OSA manifest EDS. Hypocretin-1, neuropeptide Y, leptin, ghrelin and adiponectin are implicated in both metabolic and sleep regulation, two conditions affected by OSA. We hypothesized that levels of these peptides may be related to EDS in OSA patients.We included 132 patients with EDS, as defined by an Epworth Sleepiness Scale (ESS) score ≥ 13 (mean ± SD, 15.7 ± 2.3) and 132 patients without EDS as defined by an ESS score ≤ 9 (6.5 ± 1.9). All patients had an apnea-hypopnea index (AHI) ≥ 20 h(-1). Both groups were matched for gender (males; 83.3% vs. 85.6%), age (50.15 ± 11.2 yrs vs. 50.7 ± 9.9 yrs), body mass index (BMI) (31.8 ± 5.6 kg m(-2) vs. 32.1 ± 4.8 kg m(-2)), and apnea-hypopnea index (AHI) (45.5 ± 19.1 h(-1) vs. 43 ± 19.2 h(-1)).OSA patients with EDS showed significantly higher plasma hypocretin-1 levels (p < 0.001) and lower plasma ghrelin levels (p < 0.001) than OSA patients without EDS. There were no statistically significant differences in neuropeptide Y (p = 0.08), leptin (p = 0.07) and adiponectin (p = 0.72) between the two groups. In the multiple linear regression model ESS score was associated with plasma levels of hypocretin-1, ghrelin and total sleep time.Our study shows that EDS in patients with OSA is associated with increased circulating hypocretin-1 and decreased circulating ghrelin levels, two peptides involved in the regulation of body weight, energy balance, sympathetic tone and sleep-wake cycle. This relationship is independent of AHI and obesity (two key phenotypic features of OSA).

  View details for DOI 10.1016/j.rmed.2011.08.014

  View details for Web of Science ID 000297779900026

  View details for PubMedID 21889324

• Activation of Central Orexin/Hypocretin Neurons by Dietary Amino Acids NEURON Karnani, M. M., Apergis-Schoute, J., Adamantidis, A., Jensen, L. T., de Lecea, L., Fugger, L., Burdakov, D. 2011; 72 (4): 616-629

  Abstract

  Hypothalamic orexin/hypocretin (orx/hcrt) neurons regulate energy balance, wakefulness, and reward; their loss produces narcolepsy and weight gain. Glucose can lower the activity of orx/hcrt cells, but whether other dietary macronutrients have similar effects is unclear. We show that orx/hcrt cells are stimulated by nutritionally relevant mixtures of amino acids (AAs), both in brain slice patch-clamp experiments, and in c-Fos expression assays following central or peripheral administration of AAs to mice in vivo. Physiological mixtures of AAs electrically excited orx/hcrt cells through a dual mechanism involving inhibition of K(ATP) channels and activation of system-A amino acid transporters. Nonessential AAs were more potent in activating orx/hcrt cells than essential AAs. Moreover, the presence of physiological concentrations of AAs suppressed the glucose responses of orx/hcrt cells. These results suggest a new mechanism of hypothalamic integration of macronutrient signals and imply that orx/hcrt cells sense macronutrient balance, rather than net energy value, in extracellular fluid.

  View details for DOI 10.1016/j.neuron.2011.08.027

  View details for Web of Science ID 000297180100012

  View details for PubMedID 22099463

• Neural Integration of Reward, Arousal, and Feeding: Recruitment of VTA, Lateral Hypothalamus, and Ventral Striatal Neurons IUBMB LIFE Gutierrez, R., Kay Lobo, M., Zhang, F., de Lecea, L. 2011; 63 (10): 824-830

  Abstract

  The ability to control neuronal activity using light pulses and optogenetic tools has revealed new properties of neural circuits and established causal relationships between activation of a single genetically defined population of neurons and complex behaviors. Here, we briefly review the causal effect of activity of six genetically defined neural circuits on behavior, including the dopaminergic neurons DA in the ventral tegmental area (VTA); the two main populations of medium-sized spiny neurons (D1- and D2-positive) in the striatum; the giant Cholinergic interneurons in the ventral striatum; and the hypocretin- and MCH- expressing neurons in the lateral hypothalamus. We argue that selective spatiotemporal recruitment and coordinated spiking activity among these cell type-specific neural circuits may underlie the neural integration of reward, learning, arousal and feeding.

  View details for DOI 10.1002/iub.539

  View details for Web of Science ID 000295375800010

• Optogenetic disruption of sleep continuity impairs memory consolidation PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Rolls, A., Colas, D., Adamantidis, A., Carter, M., Lanre-Amos, T., Heller, H. C., de Lecea, L. 2011; 108 (32): 13305-13310

  Abstract

  Memory consolidation has been proposed as a function of sleep. However, sleep is a complex phenomenon characterized by several features including duration, intensity, and continuity. Sleep continuity is disrupted in different neurological and psychiatric conditions, many of which are accompanied by memory deficits. This finding has raised the question of whether the continuity of sleep is important for memory consolidation. However, current techniques used in sleep research cannot manipulate a single sleep feature while maintaining the others constant. Here, we introduce the use of optogenetics to investigate the role of sleep continuity in memory consolidation. We optogenetically targeted hypocretin/orexin neurons, which play a key role in arousal processes. We used optogenetics to activate these neurons at different intervals in behaving mice and were able to fragment sleep without affecting its overall amount or intensity. Fragmenting sleep after the learning phase of the novel object recognition (NOR) task significantly decreased the performance of mice on the subsequent day, but memory was unaffected if the average duration of sleep episodes was maintained at 62-73% of normal. These findings demonstrate the use of optogenetic activation of arousal-related nuclei as a way to systematically manipulate a specific feature of sleep. We conclude that regardless of the total amount of sleep or sleep intensity, a minimal unit of uninterrupted sleep is crucial for memory consolidation.

  View details for DOI 10.1073/pnas.1015633108

  View details for Web of Science ID 000293691400068

  View details for PubMedID 21788501

  View details for PubMedCentralID PMC3156195

• Optogenetic Interrogation of Dopaminergic Modulation of the Multiple Phases of Reward-Seeking Behavior JOURNAL OF NEUROSCIENCE Adamantidis, A. R., Tsai, H., Boutrel, B., Zhang, F., Stuber, G. D., Budygin, E. A., Tourino, C., Bonci, A., Deisseroth, K., de Lecea, L. 2011; 31 (30): 10829-10835

  Abstract

  Phasic activation of dopaminergic neurons is associated with reward-predicting cues and supports learning during behavioral adaptation. While noncontingent activation of dopaminergic neurons in the ventral tegmental are (VTA) is sufficient for passive behavioral conditioning, it remains unknown whether the phasic dopaminergic signal is truly reinforcing. In this study, we first targeted the expression of channelrhodopsin-2 to dopaminergic neurons of the VTA and optimized optogenetically evoked dopamine transients. Second, we showed that phasic activation of dopaminergic neurons in freely moving mice causally enhances positive reinforcing actions in a food-seeking operant task. Interestingly, such effect was not found in the absence of food reward. We further found that phasic activation of dopaminergic neurons is sufficient to reactivate previously extinguished food-seeking behavior in the absence of external cues. This was also confirmed using a single-session reversal paradigm. Collectively, these data suggest that activation of dopaminergic neurons facilitates the development of positive reinforcement during reward-seeking and behavioral flexibility.

  View details for DOI 10.1523/JNEUROSCI.2246-11.2011

  View details for Web of Science ID 000293171900010

  View details for PubMedID 21795535

  View details for PubMedCentralID PMC3171183

• Intraventricular administration of neuropeptide S has reward-like effects EUROPEAN JOURNAL OF PHARMACOLOGY Cao, J., de Lecea, L., Ikemoto, S. 2011; 658 (1): 16-21

  Abstract

  Neuropeptide S (NPS) is an endogenous brain peptide produced by neurons located in the lower brainstem, and functional studies suggest that NPS has arousing effects. Because its receptors are found in reward-associated regions throughout the brain, we evaluated whether intraventricular NPS injections elicit reward-related effects in rats. Rats increased lever presses that led to intraventricular administration of NPS (0.34-34 pmol per infusion) in a dose dependent manner, with a cue-assisted procedure. Cue-assisted self-administration of NPS was decreased by systemic administration of the dopamine receptor antagonist SCH 23390 (0.025 mg/kg, i.p.) or the hypocretin-1 (orexin-1) receptor antagonist SB 334867 (20 mg/kg, i.p.). In addition, intraventricular NPS injections (1000 pmol) induced conditioned place preference, whereas a lower dose (100 pmol) of NPS induced conditioned place aversion. Finally, NPS injections (100-1000 pmol) acutely facilitated locomotor activity, whereas repeated NPS injections did not lead to locomotor sensitization. Our data suggest that intraventricular NPS injections have reward-like effects in that NPS weakly facilitates seeking and induces positive reinforcement. These effects may depend on intact dopamine and hypocretin systems.

  View details for DOI 10.1016/j.ejphar.2011.02.009

  View details for Web of Science ID 000289394700003

  View details for PubMedID 21349268

  View details for PubMedCentralID PMC3066039

• Optogenetic investigation of neural circuits in vivo TRENDS IN MOLECULAR MEDICINE Carter, M. E., de Lecea, L. 2011; 17 (4): 197-206

  Abstract

  The recent development of light-activated optogenetic probes allows for the identification and manipulation of specific neural populations and their connections in awake animals with unprecedented spatial and temporal precision. This review describes the use of optogenetic tools to investigate neurons and neural circuits in vivo. We describe the current panel of optogenetic probes, methods of targeting these probes to specific cell types in the nervous system, and strategies of photostimulating cells in awake, behaving animals. Finally, we survey the application of optogenetic tools to studying functional neuroanatomy, behavior and the etiology and treatment of various neurological disorders.

  View details for DOI 10.1016/j.molmed.2010.12.005

  View details for Web of Science ID 000291135600005

  View details for PubMedID 21353638

  View details for PubMedCentralID PMC3148823

• The Hypocretins/Orexins: Master Regulators of Arousal and Hyperarousal Carter, M. E., Adamantidis, A., de Lecea, L., Baumann, C. R., Bassetti, C. L., Scammell, T. E. SPRINGER. 2011: 121–28

  View details for DOI 10.1007/978-1-4419-8390-9_11

  View details for Web of Science ID 000395493100011

• Neuropeptide S facilitates cue-induced relapse to cocaine seeking through activation of the hypothalamic hypocretin system PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Kallupi, M., Cannella, N., Economidou, D., Ubaldi, M., Ruggeri, B., Weiss, F., Massi, M., Marugan, J., Heilig, M., Bonnavion, P., de Lecea, L., Ciccocioppo, R. 2010; 107 (45): 19567-19572

  Abstract

  Drug addiction is a chronic relapsing disorder characterized by compulsive drug seeking and use. Environmental conditioning factors are among the major determinants of relapse in abstinent cocaine users. Here we describe a role of the neuropeptide S (NPS) system in regulating relapse. In rats with a history of cocaine self-administration, presentation of stimuli predictive of drug availability reinstates drug seeking, triggering relapse. Intracerebroventricular (ICV) injection of NPS increased conditioned reinstatement of cocaine seeking, whereas peripheral administration of the NPS receptor antagonist SHA 68 reduced it. Manipulation of the NPS receptor system did not modify cocaine self-administration. We also found that ICV NPS administration activates c-Fos expression in hypocretin-1/orexin-A (Hcrt-1/Ox-A) immunoreactive neurons in the lateral hypothalamus (LH) and in the perifornical area (PeF). Of note, intra-LH and intra-PeF administration of NPS increased conditioned reinstatement of cocaine responding, an effect that was selectively blocked with the Hcrt-1/Ox-A receptor selective antagonist SB334867. Finally, results showed that intra-LH injection of the NPS antagonist [D-Cys(tBu) (5)]NPS blocked cue-induced cocaine seeking, indicating a role for this system in the pathophysiology of drug relapse.

  View details for DOI 10.1073/pnas.1004100107

  View details for Web of Science ID 000283997800079

  View details for PubMedID 20974945

  View details for PubMedCentralID PMC2984152

• Sleep and metabolism: Role of hypothalamic neuronal circuitry BEST PRACTICE & RESEARCH CLINICAL ENDOCRINOLOGY & METABOLISM Rolls, A., Borg, J. S., de Lecea, L. 2010; 24 (5): 817-828

  Abstract

  Sleep and metabolism are intertwined physiologically and behaviorally, but the neural systems underlying their coordination are still poorly understood. The hypothalamus is likely to play a major role in the regulation sleep, metabolism, and their interaction. And increasing evidence suggests that hypocretin cells in the lateral hypothalamus may provide particularly important contributions. Here we review: 1) direct interactions between biological arousal and metabolic systems in the hypothalamus, and 2) indirect interactions between these two systems mediated by stress or reward, emphasizing the role of hypocretins. An increased understanding of the mechanisms underlying these interactions may provide novel approaches for the treatment of patients with sleep disorders and obesity, as well as suggest new therapeutic strategies for symptoms of aging, stress, or addiction.

  View details for DOI 10.1016/j.beem.2010.08.002

  View details for Web of Science ID 000285813600011

  View details for PubMedID 21112028

• Hypocretins in the Control of Sleep and Wakefulness CURRENT NEUROLOGY AND NEUROSCIENCE REPORTS Bonnavion, P., de Lecea, L. 2010; 10 (3): 174-179

  Abstract

  During the past 10 years since the discovery of hypocretins (Hcrt, also called orexins), the list of their physiologic implications has been growing, from their primary roles in the sleep-wake cycle and feeding to the control of the cardiovascular system, pain, locomotion, stress, and addiction as well as their involvement in psychiatric disorders such as panic, anxiety, and depression. This diverse set of functions is consistent with the localization of Hcrt neurons in the lateral hypothalamus, a major integrating center of sensory inputs and emotional processes, and their widespread excitatory projections throughout the brain. Newly developed optical tools allow us to manipulate the activity of genetically identified neurons with millisecond precision in vivo and to test specific hypotheses about the causal relationships between Hcrt cells and specific behaviors. Here, we review the basic roles of the Hcrt peptides and discuss how these new technologies increase our understanding of the underpinnings of alertness and arousal.

  View details for DOI 10.1007/s11910-010-0101-y

  View details for Web of Science ID 000278111700003

  View details for PubMedID 20425032

• Reelin Regulates Postnatal Neurogenesis and Enhances Spine Hypertrophy and Long-Term Potentiation JOURNAL OF NEUROSCIENCE Pujadas, L., Gruart, A., Bosch, C., Delgado, L., Teixeira, C. M., Rossi, D., de Lecea, L., Martinez, A., Delgado-Garcia, J. M., Soriano, E. 2010; 30 (13): 4636-4649

  Abstract

  Reelin, an extracellular protein essential for neural migration and lamination, is also expressed in the adult brain. To unravel the function of this protein in the adult forebrain, we generated transgenic mice that overexpress Reelin under the control of the CaMKIIalpha promoter. Overexpression of Reelin increased adult neurogenesis and impaired the migration and positioning of adult-generated neurons. In the hippocampus, the overexpression of Reelin resulted in an increase in synaptic contacts and hypertrophy of dendritic spines. Induction of long-term potentiation (LTP) in alert-behaving mice showed that Reelin overexpression evokes a dramatic increase in LTP responses. Hippocampal field EPSP during a classical conditioning paradigm was also increased in these mice. Our results indicate that Reelin levels in the adult brain regulate neurogenesis and migration, as well as the structural and functional properties of synapses. These observations suggest that Reelin controls developmental processes that remain active in the adult brain.

  View details for DOI 10.1523/JNEUROSCI.5284-09.2010

  View details for Web of Science ID 000276178000014

  View details for PubMedID 20357114

• A decade of hypocretins: past, present and future of the neurobiology of arousal ACTA PHYSIOLOGICA de Lecea, L. 2010; 198 (3): 203-208

  Abstract

  In 1998, two groups independently identified the hypocretins, also known as orexins, as two hypothalamic peptides derived from the same precursor expressed in a few thousand neurones restricted to the perifornical area. A decade later, an amazing set of discoveries has demonstrated a key role for this neurotransmitter system in arousal and beyond. Here I review some of the experiments that led to these discoveries and the implications in the neurobiology of the hypothalamus and our understanding of brain arousal.

  View details for DOI 10.1111/j.1748-1716.2009.02004.x

  View details for Web of Science ID 000274147900003

  View details for PubMedID 19473132

• The role of hypocretin in driving arousal and goal-oriented behaviors BRAIN RESEARCH Boutrel, B., Cannella, N., de Lecea, L. 2010; 1314: 103-111

  Abstract

  The hypocretins (Hcrts), also called orexins, are two neuropeptides secreted by a few thousand neurons restricted to the lateral hypothalamus. The Hcrt peptides bind to two receptors located in nuclei associated with diverse cognitive and physiological functions. Experimental evidence has demonstrated that the physiological roles of hypocretins extend far beyond its initial role in food consumption and has emerged as a key system in the fields of sleep disorders and drug addiction. Here, we discuss recent evidence demonstrating a key role of hypocretin in the motivation for reward seeking in general, and drug taking in particular, and we delineate a physiological framework for this peptidergic system in orchestrating the appropriate levels of alertness required for the elaboration and the execution of goal-oriented behaviors. We propose a general role for hypocretins in mediating arousal, especially when an organism must respond to unexpected stressors and environmental challenges, which serve to shape survival behaviors. We also discuss the limit of the current experimental paradigms to address the question of how a system normally involved in the regulation of vigilance states and hyperarousal may promote a pathological state that elicits compulsive craving and relapse to drug seeking.

  View details for DOI 10.1016/j.brainres.2009.11.054

  View details for Web of Science ID 000275312400010

  View details for PubMedID 19948148

• Hypocretins Regulate the Anxiogenic-Like Effects of Nicotine and Induce Reinstatement of Nicotine-Seeking Behavior JOURNAL OF NEUROSCIENCE Plaza-Zabala, A., Martin-Garcia, E., de Lecea, L., Maldonado, R., Berrendero, F. 2010; 30 (6): 2300-2310

  Abstract

  Emerging evidence suggests that the hypocretinergic system is involved in addictive behavior. In this study, we investigated the role of these hypothalamic neuropeptides in anxiety-like responses of nicotine and stress-induced reinstatement of nicotine-seeking behavior. Acute nicotine (0.8 mg/kg, s.c.) induced anxiogenic-like effects in the elevated plus-maze and activated the paraventricular nucleus of the hypothalamus (PVN) as revealed by c-Fos expression. Pretreatment with the hypocretin receptor 1 (Hcrtr-1) antagonist SB334867 or preprohypocretin gene deletion blocked both nicotine effects. In the PVN, SB334867 also prevented the activation of corticotrophin releasing factor (CRF) and arginine-vasopressin (AVP) neurons, which expressed Hcrtr-1. In addition, an increase of the percentage of c-Fos-positive hypocretin cells in the perifornical and dorsomedial hypothalamic (PFA/DMH) areas was found after nicotine (0.8 mg/kg, s.c.) administration. Intracerebroventricular infusion of hypocretin-1 (Hcrt-1) (0.75 nmol/1 mul) or footshock stress reinstated a previously extinguished nicotine-seeking behavior. The effects of Hcrt-1 were blocked by SB334867, but not by the CRF1 receptor antagonist antalarmin. Moreover, SB334867 did not block CRF-dependent footshock-induced reinstatement of nicotine-seeking while antalarmin was effective in preventing this nicotine motivational response. Therefore, the Hcrt system interacts with CRF and AVP neurons in the PVN and modulates the anxiogenic-like effects of nicotine whereas Hcrt and CRF play a different role in the reinstatement of nicotine-seeking. Indeed, Hcrt-1 reinstates nicotine-seeking through a mechanism independent of CRF activation whereas CRF mediates the reinstatement induced by stress.

  View details for DOI 10.1523/JNEUROSCI.5724-09.2010

  View details for Web of Science ID 000274398200030

  View details for PubMedID 20147556

  View details for PubMedCentralID PMC2829191

• Optogenetic interrogation of neural circuits: technology for probing mammalian brain structures NATURE PROTOCOLS Zhang, F., Gradinaru, V., Adamantidis, A. R., Durand, R., Airan, R. D., de Lecea, L., Deisseroth, K. 2010; 5 (3): 439-456

  Abstract

  Elucidation of the neural substrates underlying complex animal behaviors depends on precise activity control tools, as well as compatible readout methods. Recent developments in optogenetics have addressed this need, opening up new possibilities for systems neuroscience. Interrogation of even deep neural circuits can be conducted by directly probing the necessity and sufficiency of defined circuit elements with millisecond-scale, cell type-specific optical perturbations, coupled with suitable readouts such as electrophysiology, optical circuit dynamics measures and freely moving behavior in mammals. Here we collect in detail our strategies for delivering microbial opsin genes to deep mammalian brain structures in vivo, along with protocols for integrating the resulting optical control with compatible readouts (electrophysiological, optical and behavioral). The procedures described here, from initial virus preparation to systems-level functional readout, can be completed within 4-5 weeks. Together, these methods may help in providing circuit-level insight into the dynamics underlying complex mammalian behaviors in health and disease.

  View details for DOI 10.1038/nprot.2009.226

  View details for Web of Science ID 000275234900006

  View details for PubMedID 20203662

• Optogenetic deconstruction of sleep-wake circuitry in the brain. Frontiers in molecular neuroscience Adamantidis, A., Carter, M. C., de Lecea, L. 2010; 2: 31-?

  Abstract

  How does the brain regulate the sleep-wake cycle? What are the temporal codes of sleep and wake-promoting neural circuits? How do these circuits interact with each other across the light/dark cycle? Over the past few decades, many studies from a variety of disciplines have made substantial progress in answering these fundamental questions. For example, neurobiologists have identified multiple, redundant wake-promoting circuits in the brainstem, hypothalamus, and basal forebrain. Sleep-promoting circuits have been found in the preoptic area and hypothalamus. One of the greatest challenges in recent years has been to selectively record and manipulate these sleep-wake centers in vivo with high spatial and temporal resolution. Recent developments in microbial opsin-based neuromodulation tools, collectively referred to as "optogenetics," have provided a novel method to demonstrate causal links between neural activity and specific behaviors. Here, we propose to use optogenetics as a fundamental tool to probe the necessity, sufficiency, and connectivity of defined neural circuits in the regulation of sleep and wakefulness.

  View details for DOI 10.3389/neuro.02.031.2009

  View details for PubMedID 20126433

  View details for PubMedCentralID PMC2814554

• A role for Melanin-Concentrating Hormone in learning and memory PEPTIDES Adamantidis, A., de Lecea, L. 2009; 30 (11): 2066-2070

  Abstract

  The neurobiological substrate of learning process and persistent memory storage involves multiple brain areas. The neocortex and hippocampal formation are known as processing and storage sites for explicit memory, whereas the striatum, amygdala, neocortex and cerebellum support implicit memory. Synaptic plasticity, long-term changes in synaptic transmission efficacy and transient recruitment of intracellular signaling pathways in these brain areas have been proposed as possible mechanisms underlying short- and long-term memory retention. In addition to the classical neurotransmitters (glutamate, GABA), experimental evidence supports a role for neuropeptides in modulating memory processes. This review focuses on the role of the Melanin-Concentrating Hormone (MCH) and receptors on memory formation in animal studies. Possible mechanisms may involve direct MCH modulation of neural circuit activity that support memory storage and cognitive functions, as well as indirect effect on arousal.

  View details for DOI 10.1016/j.peptides.2009.06.024

  View details for Web of Science ID 000271733100017

  View details for PubMedID 19576257

• Sleep Homeostasis Modulates Hypocretin-Mediated Sleep-to-Wake Transitions JOURNAL OF NEUROSCIENCE Carter, M. E., Adamantidis, A., Ohtsu, H., Deisseroth, K., de Lecea, L. 2009; 29 (35): 10939-10949

  Abstract

  The hypocretins (Hcrts) (also called orexins) are two neuropeptides expressed in the lateral hypothalamus that play a crucial role in the stability of wakefulness. Previously, our laboratory demonstrated that in vivo photostimulation of Hcrt neurons genetically targeted with ChR2, a light-activated cation channel, was sufficient to increase the probability of an awakening event during both slow-wave sleep and rapid eye movement sleep. In the current study, we ask whether Hcrt-mediated sleep-to-wake transitions are affected by light/dark period and sleep pressure. We found that stimulation of Hcrt neurons increased the probability of an awakening event throughout the entire light/dark period but that this effect was diminished with sleep pressure induced by 2 or 4 h of sleep deprivation. Interestingly, photostimulation of Hcrt neurons was still sufficient to increase activity assessed by c-Fos expression in Hcrt neurons after sleep deprivation, although this stimulation did not cause an increase in transitions to wakefulness. In addition, we found that photostimulation of Hcrt neurons increases neural activity assessed by c-Fos expression in the downstream arousal-promoting locus ceruleus and tuberomammilary nucleus but not after 2 h of sleep deprivation. Finally, stimulation of Hcrt neurons was still sufficient to increase the probability of an awakening event in histidine decarboxylase-deficient knock-out animals. Collectively, these results suggest that the Hcrt system promotes wakefulness throughout the light/dark period by activating multiple downstream targets, which themselves are inhibited with increased sleep pressure.

  View details for DOI 10.1523/JNEUROSCI.1205-09.2009

  View details for Web of Science ID 000269518500018

  View details for PubMedID 19726652

• The Hypocretins and their Role in Narcolepsy CNS & NEUROLOGICAL DISORDERS-DRUG TARGETS Kroeger, D., de Lecea, L. 2009; 8 (4): 271-280

  Abstract

  A series of discoveries spanning the last decade have uncovered a new neurotransmitter - hypocretin - and its role in energy metabolism, arousal, and addiction. Also, notably, a lack of hypocretin function has been unequivocally associated with the sleep disorder narcolepsy. Here we review these findings and discuss how they will influence future treatments of narcolepsy and other arousal and hyperarousal disorders. We introduce the concept of the hypocretin peptides and receptors and discuss the neuroanatomy and neurophysiology of the hypocretin system. A gain of function through pharmacolological and optogenetic means is also addressed in the following text, as is the loss of function: specifically narcolepsy in dogs, mice and humans and the challenges currently faced in treatment.

  View details for Web of Science ID 000271343800006

  View details for PubMedID 19689309

• Hypocretins and Cocaine Addiction: An Update with Hypocretin-Deficient Mice Steiner, N., de Lecea, L., Halfon, O., Boutrel, B. ELSEVIER SCIENCE INC. 2009: 107S

  View details for Web of Science ID 000265144200348

• Neuropeptide S Reinstates Cocaine-Seeking Behavior and Increases Locomotor Activity through Corticotropin-Releasing Factor Receptor 1 in Mice JOURNAL OF NEUROSCIENCE Paneda, C., Huitron-Resendiz, S., Frago, L. M., Chowen, J. A., Picetti, R., de Lecea, L., Roberts, A. J. 2009; 29 (13): 4155-4161

  Abstract

  Neuropeptide S (NPS) is a recently discovered neuropeptide that increases arousal and wakefulness while decreasing anxiety-like behavior. Here, we used a self-administration paradigm to demonstrate that intracerebroventricular infusion of NPS reinstates extinguished cocaine-seeking behavior in a dose-dependent manner in mice. The highest dose of NPS (0.45 nM) increased active lever pressing in the absence of cocaine to levels that were equivalent to those observed during self-administration. In addition, we examined the role of the corticotropin-releasing factor receptor 1 (CRF(1)) in this behavior as well as locomotor stimulation and anxiolysis. CRF(1) knock-out mice did not respond to either the locomotor stimulant or cocaine reinstatement effects of NPS, but still responded to its anxiolytic effect. The CRF(1) antagonist antalarmin also blocked the increase in active lever responding in the reinstatement model and the locomotor activating properties of NPS without affecting its anxiolytic actions. Our results suggest that NPS receptors may be an important target for drug abuse research and treatment and that CRF(1) mediates the cocaine-seeking and locomotor stimulant effects of NPS, but not its effects on anxiety-like behavior.

  View details for DOI 10.1523/JNEUROSCI.5256-08.2009

  View details for Web of Science ID 000264767500019

  View details for PubMedID 19339610

• The brain hypocretins and their receptors: mediators of allostatic arousal CURRENT OPINION IN PHARMACOLOGY Carter, M. E., Borg, J. S., de Lecea, L. 2009; 9 (1): 39-45

  Abstract

  The hypocretins (abbreviated 'Hcrts' - also called 'orexins') are two neuropeptides secreted exclusively by a small population of neurons in the lateral hypothalamus. These peptides bind to two receptors located throughout the brain in nuclei associated with diverse cognitive and physiological functions. Initially, the brain Hcrt system was found to have a major role in the regulation of sleep/wake transitions. More recent studies indicate Hcrts may play a role in other physiological functions, including food intake, addiction, and stress. Taken together, these studies suggest a general role for Hcrts in mediating arousal, especially when an organism must respond to unexpected stressors and challenges in the environment.

  View details for DOI 10.1016/j.coph.2008.12.018

  View details for Web of Science ID 000263817700007

  View details for PubMedID 19185540

• The hypocretins as sensors for metabolism and arousal JOURNAL OF PHYSIOLOGY-LONDON Adamantidis, A., de Lecea, L. 2009; 587 (1): 33-40

  Abstract

  Sleep disturbances are associated with hormonal imbalances and may result in metabolic disorders including obesity and diabetes. Therefore, circuits controlling both sleep and metabolism are likely to play a role in these physiopathological conditions. The hypocretin (Hcrt) system is a strong candidate for mediating both sleep and metabolic imbalances because Hcrt neurons are sensitive to metabolic hormones, including leptin and ghrelin, and modulate arousal and goal-orientated behaviours. This review discusses the role of Hcrt neurons as a sensors of energy balance and arousal and proposes new ways of probing local hypothalamic circuits regulating sleep and metabolism with unprecedented cellular specificity and temporal resolution.

  View details for DOI 10.1113/jphysiol.2008.164400

  View details for Web of Science ID 000262151800012

  View details for PubMedID 19047201

  View details for PubMedCentralID PMC2670020

• A Consensus Definition of Cataplexy in Mouse Models of Narcolepsy SLEEP Scammell, T. E., Willie, J. T., Guilleminault, C., Siegel, J. M., Int Working Grp Rodent Models Nar 2009; 32 (1): 111–16

  Abstract

  People with narcolepsy often have episodes of cataplexy, brief periods of muscle weakness triggered by strong emotions. Many researchers are now studying mouse models of narcolepsy, but definitions of cataplexy-like behavior in mice differ across labs. To establish a common language, the International Working Group on Rodent Models of Narcolepsy reviewed the literature on cataplexy in people with narcolepsy and in dog and mouse models of narcolepsy and then developed a consensus definition of murine cataplexy. The group concluded that murine cataplexy is an abrupt episode of nuchal atonia lasting at least 10 seconds. In addition, theta activity dominates the EEG during the episode, and video recordings document immobility. To distinguish a cataplexy episode from REM sleep after a brief awakening, at least 40 seconds of wakefulness must precede the episode. Bouts of cataplexy fitting this definition are common in mice with disrupted orexin/hypocretin signaling, but these events almost never occur in wild type mice. It remains unclear whether murine cataplexy is triggered by strong emotions or whether mice remain conscious during the episodes as in people with narcolepsy. This working definition provides helpful insights into murine cataplexy and should allow objective and accurate comparisons of cataplexy in future studies using mouse models of narcolepsy.

  View details for Web of Science ID 000262075600018

  View details for PubMedID 19189786

  View details for PubMedCentralID PMC2625315

• Optogenetic probing of hypocretin neuronal network 19th Congress of the European-Sleep-Research-Society Adamantidis, A. R., Zhang, F., Aravanis, A., Deisseroth, K., De Lecea, L. WILEY-BLACKWELL. 2008: 88–88

  View details for Web of Science ID 000262850300220

• Sleep and metabolism: shared circuits, new connections TRENDS IN ENDOCRINOLOGY AND METABOLISM Adamantidis, A., de Lecea, L. 2008; 19 (10): 362-370

  Abstract

  Association between sleep disturbances and hormonal imbalances can result in metabolic disorders, including obesity and diabetes. The hypothalamus is likely to play a part in these pathophysiological conditions because it contains sleep-wake circuits that are sensitive to metabolic hormones, including leptin and ghrelin. Thus, shared hypothalamic circuits such as the hypocretin and melanin-concentrating hormone systems are strong candidates for mediating both sleep and metabolic imbalances. This review reveals new roles for these systems as sensors and effectors of sleep and wakefulness, and discusses their plasticity in regulating sleep and energy balance. New optical tools that remotely control neuronal circuit activity provide an effective means to understand the cooperativity of shared circuits in regulating hypothalamic functions such as sleep and metabolism.

  View details for DOI 10.1016/j.tem.2008.08.007

  View details for Web of Science ID 000261477600003

  View details for PubMedID 18938086

• Effect of cortistatin on tau phosphorylation at Ser262 site JOURNAL OF NEUROSCIENCE RESEARCH Rubio, A., Perez, M., de Lecea, L., Avila, J. 2008; 86 (11): 2462-2475

  Abstract

  The development of intraneuronal lesions as a result of the progressive deposition of hyperphosphorylated tau at specific brain regions (such as hippocampus and cortex) plays a key role in the pathological process of Alzheimer's disease. However, the mechanisms by which tau phosphorylation is regulated, mainly in the pathology found in the cortex, are still poorly understood. Here, we analyzed the effect of cortistatin, a cortical neuropeptide related to somatostatin, on tau phosphorylation at Ser262 in cultures of murine cortical neurons. Both somatostatin and cortistatin induce tau phosphorylation at Ser262, a site modified in Alzheimer's disease, although with different kinetics in cortex. The effect of cortistatin likely is mediated by heterodimeric receptors composed of somatostatin receptor subtypes 2 and 4 and also by protein kinase C signaling. Cortistatin-deficient mice show decreased tau phosphorylation at Ser262 in the cortex but not in other brain regions tested. Our results suggest an important role for cortistatin in the regulation of tau phosphorylation that may be associated with the pathophysiology of Alzheimer's disease in regions such as the cerebral cortex.

  View details for DOI 10.1002/jnr.21689

  View details for Web of Science ID 000258478100011

  View details for PubMedID 18438934

• Cortistatin - Functions in the central nervous system MOLECULAR AND CELLULAR ENDOCRINOLOGY de Lecea, L. 2008; 286 (1-2): 88-95

  Abstract

  Cortistatin (CST) is a neuropeptide from the somatostatin (SRIF)/urotensin (UII) family named after its predominantly cortical expression and ability to depress cortical activity, which was discovered a decade ago. In vitro assays show CST is able to bind all five cloned somatostatin receptors and shares many pharmacological and functional properties with SRIF. However, distinct from SRIF, CST has been shown to induce slow-wave sleep, reduce locomotor activity, and activate cation selective currents not responsive to somatostatin. Different lines of evidence also indicate that CST, like SRIF, is involved in learning and memory processes. CST-14 may also function as an endogenous anti-convulsant. In addition to its role in cortical synchronization, CST-14 has emerged as an important mediator of immunity and inflammation. This review will cover some of the basic properties of CST in the brain, and will discuss new data on the role of CST in cortical activity.

  View details for DOI 10.1016/j.mce.2007.12.014

  View details for Web of Science ID 000257024900013

  View details for PubMedID 18374474

• Somatostatin, cortistatin and their receptors in health and disease. Foreword. Molecular and cellular endocrinology Castaño, J. P., Ghigo, E., Kineman, R. D., de Lecea, L., Malagón, M. M., Vaudry, H. 2008; 286 (1-2): 1-2

  View details for DOI 10.1016/j.mce.2008.03.010

  View details for PubMedID 18456395

• Physiological arousal: a role for hypothalamic systems CELLULAR AND MOLECULAR LIFE SCIENCES Adamantidis, A., de Lecea, L. 2008; 65 (10): 1475-1488

  Abstract

  The lateral hypothalamus (LH) has long been known as a homeostasis center of the brain that modulates feeding behavior, arousal and reward. The hypocretins (Hcrts, also called orexins) and melanin-concentrating hormone (MCH) are neuropeptides produced in two intermingled populations of a few thousand neurons in the LH. The Hcrts have a prominent role in regulating the stability of arousal, since Hcrt system deficiency leads to narcolepsy. MCH is an important modulator of energy balance, as MCH system deficiency in mice leads to leanness and increased metabolism. Recently, MCH has been proposed to modulate rapid eye movement sleep in rodents. In this review, we propose a working model of the cross-talk between Hcrt and MCH circuits that may provide an arousal balance system to regulate complex goal-oriented behaviors.

  View details for DOI 10.1007/s00018-008-7521-8

  View details for Web of Science ID 000256252500003

  View details for PubMedID 18351292

• Neuropeptide interactions and REM sleep: A role for Urotensin II? 1st Meeting of the Japan Branch of the International-Neuropeptide-Society de Lecea, L., Bourgin, P. ELSEVIER SCIENCE INC. 2008: 845–51

  Abstract

  Urotensin II (UII) is a peptide with structural similarity to the somatostatin family with potent vasoconstrictor activity. UII receptor is expressed broadly in the periphery, and most notably in the heart and microvessels. In the brain, the UII receptor can be detected in the spinal cord and in cholinergic nuclei in the brainstem known to be involved in REM sleep regulation. Recent data suggest that, in addition to their vasoactive properties, UII receptor ligands may have excitatory activity on a selective group of neurons that modulate REM sleep. This review focuses on the implications of these findings for the neurobiology of REM sleep regulation and discusses the possible impact of UII and other neuropeptides on the balance of the alternation between sleep states.

  View details for DOI 10.1016/j.peptides.2008.02.009

  View details for Web of Science ID 000256007400025

  View details for PubMedID 18406008

• Somatostatin receptor subtype 4 couples to the m-current to regulate seizures JOURNAL OF NEUROSCIENCE Qiu, C., Zeyda, T., Johnson, B., Hochgeschwender, U., de Lecea, L., Tallent, M. K. 2008; 28 (14): 3567-3576

  Abstract

  The K(+) M-current (I(M), Kv7) is an important regulator of cortical excitability, and mutations in these channels cause a seizure disorder in humans. The neuropeptide somatostatin (SST), which has antiepileptic properties, augments I(M) in hippocampal CA1 pyramidal neurons. We used SST receptor knock-out mice and subtype-selective ligands to investigate the receptor subtype that couples to I(M) and mediates the antiepileptic effects of SST. Using pentylenetetrazole as a chemoconvulsant, SST(2), SST(3), and SST(4) receptor knock-out mice all had shorter latencies to different seizure stages and increased seizure severity when compared with wild-type mice. However, the most robust differences were observed in the SST(4) knock-outs. When seizures were induced by systemic injection of kainate, only SST(4) knock-outs showed an increase in seizure sensitivity. We next examined the action of SST and subtype-selective SST agonists on electrophysiological parameters in hippocampal slices of wild-type and receptor knock-out mice. SST(2) and SST(4) appear to mediate the majority of SST inhibition of epileptiform activity in CA1. SST lacked presynaptic effects in mouse CA1, in contrast to our previous findings in rat. SST increased I(M) in CA1 pyramidal neurons of wild-type and SST(2) knock-out mice, but not SST(4) knock-out mice. Using M-channel blockers, we found that SST(4) coupling to M-channels is critical to its inhibition of epileptiform activity. This is the first demonstration of an endogenous enhancer of I(M) that is important in controlling seizure activity. SST(4) receptors could therefore be an important novel target for developing new antiepileptic and antiepileptogenic drugs.

  View details for DOI 10.1523/JNEUROSCI.4679-07.2008

  View details for Web of Science ID 000254623500006

  View details for PubMedID 18385315

• Addiction and arousal: The hypocretin connection PHYSIOLOGY & BEHAVIOR Boutrel, B., de Lecea, L. 2008; 93 (4-5): 947-951

  Abstract

  The hypocretins, also known as orexins, are two neuropeptides now commonly described as critical components to maintain and regulate the stability of arousal. Several lines of evidence have raised the hypothesis that hypocretin-producing neurons are part of the circuitries that mediate the hypothalamic response to acute stress. Intracerebral administration of hypocretin leads to a dose-related reinstatement of drug and food seeking behaviors. Furthermore, stress-induced reinstatement can be blocked with hypocretin receptor 1 antagonism. These results, together with recent data showing that hypocretin is critically involved in cocaine sensitization through the recruitment of NMDA receptors in the ventral tegmental area, strongly suggest that activation of hypocretin neurons play a critical role in the development of the addiction process. The activity of hypocretin neurons may affect addictive behavior by contributing to brain sensitization or by modulating the brain reward system. Hypocretinergic cells, in coordination with brain stress systems may lead to a vulnerable state that facilitates the resumption of drug seeking behavior. Hence, the hypocretinergic system is a new drug target that may be used to prevent relapse of drug seeking.

  View details for DOI 10.1016/j.physbeh.2007.11.022

  View details for Web of Science ID 000255311100035

  View details for PubMedID 18262574

• Neural substrates of awakening probed with optogenetic control of hypocretin neurons 22nd Annual Meeting of the Associated-Professional-Sleep-Societies Adamantidis, A. R., Zhang, F., Aravanis, A., Deisseroth, K., De Lecea, L. AMER ACAD SLEEP MEDICINE. 2008: A364–A364

  View details for Web of Science ID 000255419001533

• Circuit-breakers: optical technologies for probing neural signals and systems NATURE REVIEWS NEUROSCIENCE Zhang, F., Aravanis, A. M., Adamantidis, A., de Lecea, L., Deisseroth, K. 2007; 8 (8): 577-581

  Abstract

  Neuropsychiatric disorders, which arise from a combination of genetic, epigenetic and environmental influences, epitomize the challenges faced in understanding the mammalian brain. Elucidation and treatment of these diseases will benefit from understanding how specific brain cell types are interconnected and signal in neural circuits. Newly developed neuroengineering tools based on two microbial opsins, channelrhodopsin-2 (ChR2) and halorhodopsin (NpHR), enable the investigation of neural circuit function with cell-type-specific, temporally accurate and reversible neuromodulation. These tools could lead to the development of precise neuromodulation technologies for animal models of disease and clinical neuropsychiatry.

  View details for DOI 10.1038/nrn2192

  View details for Web of Science ID 000248211800012

  View details for PubMedID 17643087

• Cortistatin promotes and negatively correlates with slow-wave sleep EUROPEAN JOURNAL OF NEUROSCIENCE Bourgin, P., Fabre, V., Huitron-Resendiz, S., Henriksen, S. J., Prospero-Garcia, O., Criado, J. R., de Lecea, L. 2007; 26 (3): 729-738

  Abstract

  Sleep need is characterized by the level of slow-wave activity (SWA) and increases with time spent awake. The molecular nature of this sleep homeostatic process is practically unknown. Here, we show that intracerebroventricular administration of the neuropeptide, cortistatin (CST-14), enhances EEG synchronization by selectively promoting deep slow-wave sleep (SWS) during both the light and dark period in rats. CST-14 also increases the level of slow-wave activity (SWA) within deep SWS during the first two hours following CST-14 administration. Steady-state levels of preprocortistatin mRNA oscillate during the light:dark cycle and are four-fold higher upon total 24-h sleep deprivation, returning progressively to normal levels after eight hours of sleep recovery. Preprocortistatin mRNA is expressed upon sleep deprivation in a particular subset of cortical interneurons that colocalize with c-fos. In contrast, the number of CST-positive cells coexpressing pERK1/2 decreases under sleep deprivation. The capacity of CST-14 to increase SWA, together with preprocortistatin's inverse correlation with time spent in SWS, suggests a potential role in sleep homeostatic processes.

  View details for DOI 10.1111/j.1460-9568.2007.05696.x

  View details for Web of Science ID 000248598100020

  View details for PubMedID 17686045

• Cortistatin as a therapeutic target in inflammation EXPERT OPINION ON THERAPEUTIC TARGETS Rubio, A., Avila, J., de Lecea, L. 2007; 11 (1): 1-9

  Abstract

  Cortistatin (CST) is a recently discovered neuropeptide from the somatostatin gene family, named after its predominantly cortical expression and ability to depress cortical activity. CST shows many remarkable structural and functional similarities to its related neuropeptide somatostatin, or somatotropin release-inhibiting factor. However, the many physiological differences between CST and somatostatin are just as remarkable as the similarities. CST-29 has recently been shown to prevent inflammation in rodent models for human diseases, raising novel therapeutic properties to this neuropeptide. In this review, the authors address a new possible role for CST in the immune system and evaluate the possible therapeutic use of CST to treat disorders associated with inflammation.

  View details for DOI 10.1517/14728222.11.1.1

  View details for Web of Science ID 000243300800002

  View details for PubMedID 17150030

• Identification of novel transcripts expressed in hypocretin-containing neurons 21st Annual Meeting of the American-Professional-Sleep-Societies Bourgin, P., Lin, L., Priest, M., Renier, C., Sakurai, T., De Lecea, L., Mignot, E. AMER ACAD SLEEP MEDICINE. 2007: A223–A223

  View details for Web of Science ID 000246224900659

• Transgenic mice with a reduced core body temperature have an increased life span SCIENCE Conti, B., Sanchez-Alavez, M., Winsky-Sommerer, R., Morale, M. C., Lucero, J., Brownell, S., Fabre, V., Huitron-Resendiz, S., Henriksen, S., Zorrilla, E. P., de Lecea, L., Bartfai, T. 2006; 314 (5800): 825-828

  Abstract

  Reduction of core body temperature has been proposed to contribute to the increased life span and the antiaging effects conferred by calorie restriction (CR). Validation of this hypothesis has been difficult in homeotherms, primarily due to a lack of experimental models. We report that transgenic mice engineered to overexpress the uncoupling protein 2 in hypocretin neurons (Hcrt-UCP2) have elevated hypothalamic temperature. The effects of local temperature elevation on the central thermostat resulted in a 0.3 degrees to 0.5 degrees C reduction of the core body temperature. Fed ad libitum, Hcrt-UCP2 transgenic mice had the same caloric intake as their wild-type littermates but had increased energy efficiency and a greater median life span (12% increase in males; 20% increase in females). Thus, modest, sustained reduction of core body temperature prolonged life span independent of altered diet or CR.

  View details for DOI 10.1126/science.1132191

  View details for Web of Science ID 000241729800049

  View details for PubMedID 17082459

• Addiction and arousal: Alternative roles of hypothalamic peptides JOURNAL OF NEUROSCIENCE de Lecea, L., Jones, B. E., Boutrel, B., Borgland, S. L., Nishino, S., Bubser, M., DiLeone, R. 2006; 26 (41): 10372-10375

  Abstract

  The importance of the lateral hypothalamus in the regulation of reward and motivation has long been recognized. However, the neuronal network involved in such a hypothalamic regulation of reward remains essentially unknown. Recently, hypocretin-containing neurons, a group of hypothalamic neurons known to be associated with the stability of arousal, have emerged as important structures in the control of brain reward function. This review summarizes a Mini-Symposium presented at the 2006 Annual Meeting of the Society for Neuroscience.

  View details for DOI 10.1523/JNEUROSCI.3118-06.2006

  View details for Web of Science ID 000241192800008

  View details for PubMedID 17035520

• Cortistatin: not just another somatostatin analog NATURE CLINICAL PRACTICE ENDOCRINOLOGY & METABOLISM de Lecea, L., Castano, J. P. 2006; 2 (7): 356-357

  View details for DOI 10.1038/ncpendmet0219

  View details for Web of Science ID 000239085200002

  View details for PubMedID 16932314

• New genes involved in cortical development DYSLEXIC BRAIN Burgaya, F., Garcia-Frigolo, C., Andres, R., Vitureira, N., Lopez-Domenech, G., De Lecea, L., Soriano, E. 2006: 143-?

  View details for Web of Science ID 000236464900008

• Role for hypocretin in mediating stress-induced reinstatement of cocaine-seeking behavior PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Boutrel, B., Kenny, P. J., Specio, S. E., Martin-Fardon, R., Markou, A., Koob, G. F., de Lecea, L. 2005; 102 (52): 19168-19173

  Abstract

  Hypocretin-1 and -2 (Hcrt-1 and Hcrt-2), also referred to as orexin-A and -B, are neuropeptides synthesized by a few thousand neurons in the lateral hypothalamus. Hypocretin-containing neurons project throughout the brain, with a prominent input to basal forebrain structures involved in motivation, reward, and stress. However, the role of hypocretins in addiction-related behaviors remains largely unexplored. Here we show that intracerebroventricular infusions of Hcrt-1 lead to a dose-related reinstatement of cocaine seeking without altering cocaine intake in rats. Hcrt-1 also dramatically elevates intracranial self-stimulation thresholds, indicating that, unlike treatments with reinforcing properties such as cocaine, Hcrt-1 negatively regulates the activity of brain reward circuitries. Hypocretin-induced reinstatement of cocaine seeking was prevented by blockade of noradrenergic and corticotropin-releasing factor systems, suggesting that Hcrt-1 reinstated drug seeking through induction of a stress-like state. Consistent with this interpretation, the selective Hcrt-1 receptor antagonist SB-334867 blocked footshock-induced reinstatement of previously extinguished cocaine-seeking behavior. These findings reveal a previously unidentified role for hypocretins in driving drug seeking through activation of stress pathways in the brain.

  View details for DOI 10.1073/pnas.0507480102

  View details for Web of Science ID 000234350000069

  View details for PubMedID 16357203

• Stress and arousal - The corticotrophin-releasing factor/hypocretin circuitry MOLECULAR NEUROBIOLOGY Winsky-Sommerer, R., Boutrel, B., de Lecea, L. 2005; 32 (3): 285-294

  Abstract

  The hypocretins (also know as orexins) are two neuropeptides now commonly described as critical components for maintaining and regulating the stability of arousal. Several lines of evidence have raised the hypothesis that hypocretin-producing neurons are part of the circuitries that mediate the hypothalamic response to acute stress. New data indicate that the corticotrophin-releasing factor (CRF) peptidergic system directly innervates hypocretin-expressing neurons. CRF depolarizes hypocretin neurons, and this effect is blocked by a CRF-R1 antagonist. Furthermore, activation of hypocretinergic neurons by stress is impaired in CRF-R1 knockout mice. These data suggest that CRF-R1 receptor mediates the stress-induced activation of the hypocretinergic system. A significant amount of evidence also indicates that hypocretin cells connect reciprocally to the CRF system. We propose that upon stressor stimuli, CRF activates the hypocretin system, which relays these signals to brain stem nuclei involved in the modulation of arousal as well as to the extended amygdala, a structure involved in the negative motivational state that drives addiction.

  View details for Web of Science ID 000233294300006

  View details for PubMedID 16385142

• Cortistatin overexpression in transgenic mice produces deficits in synaptic plasticity and learning MOLECULAR AND CELLULAR NEUROSCIENCE Tallent, M. K., Veronique, F., Qiu, C., Calbet, M., Lamp, T., Baratta, M. V., Suzuki, C., Levy, C. L., Siggins, G. R., Henriksen, S. J., Criado, J. R., Roberts, A., de Lecea, L. 2005; 30 (3): 465-475

  Abstract

  Cortistatin-14 (CST) is a neuropeptide expressed in cortical and hippocampal interneurons that shares 11 of 14 residues with somatostatin. In contrast to somatostatin, infusion of CST decreases locomotor activity and selectively enhances slow wave sleep. Here, we show that transgenic mice that overexpress cortistatin under the control of neuron-specific enolase promoter do not express long-term potentiation in the dentate gyrus. This blockade of dentate LTP correlates with profound impairment of hippocampal-dependent spatial learning. Exogenously applied CST to slices of wild-type mice also blocked induction of LTP in the dentate gyrus. Our findings implicate cortistatin in the modulation of synaptic plasticity and cognitive function. Thus, increases in hippocampal cortistatin expression during aging could have an impact on age-related cognitive deficits.

  View details for DOI 10.1016/j.mcn.2005.08.010

  View details for Web of Science ID 000232746600016

  View details for PubMedID 16182561

• The hypocretins and sleep FEBS JOURNAL de Lecea, L., Sutcliffe, J. G. 2005; 272 (22): 5675-5688

  Abstract

  The hypocretins (also called the orexins) are two neuropeptides derived from the same precursor whose expression is restricted to a few thousand neurons of the lateral hypothalamus. Two G-protein coupled receptors for the hypocretins have been identified, and these show different distributions within the central nervous system and differential affinities for the two hypocretins. Hypocretin fibers project throughout the brain, including several areas implicated in regulation of the sleep/wakefulness cycle. Central administration of synthetic hypocretin-1 affects blood pressure, hormone secretion and locomotor activity, and increases wakefulness while suppressing rapid eye movement sleep. Most human patients with narcolepsy have greatly reduced levels of hypocretin peptides in their cerebral spinal fluid and no or barely detectable hypocretin-containing neurons in their hypothalamus. Multiple lines of evidence suggest that the hypocretinergic system integrates homeostatic, metabolic and limbic information and provides a coherent output that results in stability of the states of vigilance.

  View details for DOI 10.1111/j.1742-4658.2005.04981.x

  View details for Web of Science ID 000233143600002

  View details for PubMedID 16279933

• Expression, synaptic localization, and developmental regulation of Ack1/Pyk1, a cytoplasmic tyrosine kinase highly expressed in the developing and adult brain JOURNAL OF COMPARATIVE NEUROLOGY Urena, J. M., La Torre, A., Martinez, A., Lowenstein, E., Franco, N., Winsky-Sommerer, R., Fontana, X., Casaroli-Marano, R., Ibanez-Sabio, M. A., Pascual, M., del Rio, J. A., De Lecea, L., Soriano, E. 2005; 490 (2): 119-132

  Abstract

  Cytosolic tyrosine kinases play a critical role both in neural development and in adult brain function and plasticity. Here we isolated a cDNA with high homology to human Ack1 and mouse Tnk2. This cDNA directs the expression of a 125-kD protein that can be autophosphorylated in tyrosines. Initially, this clone was named Pyk1 for proline-rich tyrosine kinase (Lev et al., 1995); however, since it corresponds to the mouse homolog of Ack1, here we called it Ack1/Pyk1. In this study we show that Ack1/Pyk1 mRNA and protein is highly expressed in the developing and adult brain. The highest levels of Ack1/Pyk1 expression were detected in the hippocampus, neocortex, and cerebellum. Electron microscopy studies showed that Ack1/Pyk1 protein is expressed in these regions both at dendritic spines and presynaptic axon terminals, indicating a role in synaptic function. Furthermore, we demonstrate that Ack1/Pyk1 mRNA levels are strongly upregulated by increased neural activity, produced by intraperitoneal kainate injections. During development, Ack1/Pyk1 was also expressed in the proliferative ventricular zones and in postmitotic maturing neurons. In neuronal cultures, Ack1/Pyk1 was detected in developing dendrites and axons, including dendritic tips and growth cones. Moreover, Ack1/Pyk1 colocalized with Cdc42 GTPase in neuronal cultures and coimmunoprecipitated with Cdc42 in HEK 293T cells. Altogether, our findings indicate that Ack1/Pyk1 tyrosine kinase may be involved both in adult synaptic function and plasticity and in brain development.

  View details for DOI 10.1002/cne.20656

  View details for Web of Science ID 000231156000002

  View details for PubMedID 16052498

• Urotensin II modulates rapid eye movement sleep through activation of brainstem cholinergic neurons JOURNAL OF NEUROSCIENCE Huitron-Resendiz, S., Kristensen, M. P., Sanchez-Alavez, M., Clark, S. D., Grupke, S. L., Tyler, C., Suzuki, C., Nothacker, H. P., Civelli, O., Criado, J. R., Henriksen, S. J., Leonard, C. S., de Lecea, L. 2005; 25 (23): 5465-5474

  Abstract

  Urotensin II (UII) is a cyclic neuropeptide with strong vasoconstrictive activity in the peripheral vasculature. UII receptor mRNA is also expressed in the CNS, in particular in cholinergic neurons located in the mesopontine tegmental area, including the pedunculopontine tegmental (PPT) and lateral dorsal tegmental nuclei. This distribution suggests that the UII system is involved in functions regulated by acetylcholine, such as the sleep-wake cycle. Here, we tested the hypothesis that UII influences cholinergic PPT neuron activity and alters rapid eye movement (REM) sleep patterns in rats. Local administration of UII into the PPT nucleus increases REM sleep without inducing changes in the cortical blood flow. Intracerebroventricular injection of UII enhances both REM sleep and wakefulness and reduces slow-wave sleep 2. Intracerebroventricular, but not local, administration of UII increases cortical blood flow. Moreover, whole-cell recordings from rat-brain slices show that UII selectively excites cholinergic PPT neurons via an inward current and membrane depolarization that were accompanied by membrane conductance decreases. This effect does not depend on action potential generation or fast synaptic transmission because it persisted in the presence of TTX and antagonists of ionotropic glutamate, GABA, and glycine receptors. Collectively, these results suggest that UII plays a role in the regulation of REM sleep independently of its cerebrovascular actions by directly activating cholinergic brainstem neurons.

  View details for DOI 10.1523/JNEUROSCI.4501-04.2005

  View details for Web of Science ID 000229643000003

  View details for PubMedID 15944374

• Injection of neuropeptide W into paraventricular nucleus of hypothalamus increases food intake AMERICAN JOURNAL OF PHYSIOLOGY-REGULATORY INTEGRATIVE AND COMPARATIVE PHYSIOLOGY Levine, A. S., Winsky-Sommerer, R., Huitron-Resendiz, S., Grace, M. K., De Lecea, L. 2005; 288 (6): R1727-R1732

  Abstract

  Neuropeptide W (NPW) is an endogenous ligand for G protein-coupled receptor 7 (GPR7). There are two forms of the peptide, designated as neuropeptide W-23 (NPW23) and neuropeptide W-30 (NPW30). In the current study we found that intracerebroventricular administration of NPW23 increased c-Fos immunoreactivity (IR) in a variety of brain sites, many of which are involved in the regulation of feeding. In particular, we noted that c-Fos IR levels were increased in hypocretin-expressing neurons in the perifornical region of the lateral hypothalamus (LH). We then studied whether injection of NPW23 into the paraventricular nucleus of the hypothalamus (PVN) and the LH increased food intake over a 24-h time period. Intra-PVN injection of NPW23 at doses ranging from 0.1 to 3 nmol increased feeding for up to 4 h, and doses ranging from 0.3 to 3 nmol increased feeding for up to 24 h. In contrast, only the 3-nmol dose of NPW23 increased feeding after administration into the LH. Together, these data suggest a modulatory role for NPW in the control of food intake.

  View details for DOI 10.1152/ajpregu.00638.2003

  View details for Web of Science ID 000229013400036

  View details for PubMedID 15886360

• The corticotropin-releasing factor-hypocretin connection: Implications in stress response and addiction DRUG NEWS & PERSPECTIVES Paneda, C., Winsky-Sommerer, R., Boutrel, B., de Lecea, L. 2005; 18 (4): 250-255

  Abstract

  The hypothalamic neuropeptides hypocretins (orexins) play a crucial role in the stability of arousal and alertness. Recent data have raised the hypothesis that hypocretin neurons are also part of the circuitries that mediate the hypothalamic stress response. In particular, we have recently demonstrated that corticotrophin-releasing factor (CRF)-immunoreactive terminals make direct synaptic contacts with hypocretin-expressing neurons and that numerous hypocretinergic neurons express the CRF-R1/2 receptors. Furthermore, CRF excites hypocretinergic cells ex vivo through CRF-R1 receptors. Activation of hypocretinergic neurons in response to acute stress is severely impaired in CRF-R1 knockout mice. Moreover, the stress response is impaired in hypocretin-deficient mice. We propose that upon stressor stimuli, CRF stimulates the release of hypocretins, and this circuit contributes to activation and maintenance of arousal associated with the stress response and addiction.

  View details for DOI 10.1358/dnp.2005.18.4.908659

  View details for Web of Science ID 000230604600004

  View details for PubMedID 16034481

• Cortistatin radioligand binding in wild-type and somatostatin receptor-deficient mouse brain REGULATORY PEPTIDES Spier, A. D., Fabre, V., de Lecea, L. 2005; 124 (1-3): 179-186

  Abstract

  Cortistatin-14 (CST-14) is a recently discovered member of the somatostatin family of neuropeptides. It shares 11 of its 14 amino acids with somatostatin-14 (SRIF-14). In the present study, binding sites for cortistatin-14 in the mouse brain were examined and compared to those for somatostatin using iodinated cortistatin-14 and iodinated somatostatin-14. By in vitro receptor autoradiography, high densities of cortistatin-14 and somatostatin-14 specific binding sites were detected in the cortex, hippocampal formation, basolateral amygdala and medial habenula. Unlabeled 100 nM cortistatin-14 inhibited iodinated somatostatin-14 binding in the hippocampus, but not in the cortex or amygdaloid nuclei. In somatostatin receptor subtype-2 knock-out (KO) mice, autoradiographic iodinated somatostatin-14 binding was observed in the hippocampus and habenula but was removed in the cortex and amygdaloid nuclei, specific iodinated cortistatin-14 binding sites were found in the hippocampus, habenula and throughout the cortex. We conclude that the somatostatin receptor subtype-2 is responsible for somatostatin binding in cortical and amygdaloid regions and that cortistatin predominantly interacts with the same receptors as somatostatin.

  View details for DOI 10.1016/j.regpep.2004.07.015

  View details for Web of Science ID 000225638300023

  View details for PubMedID 15544857

• Cortistatin: a natural somatostatin analog. Journal of endocrinological investigation de Lecea, L. 2005; 28 (11): 10-14

  Abstract

  Cortistatin (CST) is a recently discovered neuropeptide from the somatostatin gene family named after its predominantly cortical expression and ability to depress cortical activity. CST shows many remarkable structural and functional similarities to its related neuropeptide somatostatin. However, the many physiological differences between CST and somatostatin are just as remarkable as the similarities. CST-14 shares 11 of its 14 amino acids with somatostatin-14, including the FWKT tetramer thought to be responsible for somatostatin's receptor interactions and the pair of cysteine residues that likely render the peptides cyclic. Yet the nucleotide sequences and chromosomal localizations of these genes clearly indicate they are products of separate genes and CST's activity in the brain is widely distinct from that of somatostatin. Now cloned from human, mouse and rat sources, in vitro assays show that CST is able to bind all five cloned somatostatin receptors and shares many pharmacological and functional properties with somatostatin, including the depression of neuronal activity and inhibition of GH release. However, distinct from somatostatin, CST has been shown to induce slow-wave sleep, reduce locomotor activity, and activate cation selective currents not responsive to, or antagonized by, somatostatin. Here we address the discovery and characterization of this novel somatostatin-like neuropeptide, including its cloning, expression and pharmacology. We also examine the evidence pointing towards a specific receptor for this novel neuropeptide member of the somatostatin gene family.

  View details for PubMedID 16625839

• Interaction between the corticotropin-releasing factor system and hypocretins (Orexins): A novel circuit mediating stress response JOURNAL OF NEUROSCIENCE Winsky-Sommerer, R., Yamanaka, A., Diano, S., Borok, E., Roberts, A. J., Sakurai, T., Kilduff, T. S., Horvath, T. L., de Lecea, L. 2004; 24 (50): 11439-11448

  Abstract

  The hypothalamic neuropeptides hypocretins (orexins) play a crucial role in the stability of arousal and alertness. We tested whether the hypocretinergic system is a critical component of the stress response activated by the corticotropin-releasing factor (CRF). Our results show that CRF-immunoreactive terminals make direct contact with hypocretin-expressing neurons in the lateral hypothalamus and that numerous hypocretinergic neurons express the CRF-R1/2 receptors. We also demonstrate that application of CRF to hypothalamic slices containing identified hypocretin neurons depolarizes membrane potential and increases firing rate in a subpopulation of hypocretinergic cells. CRF-induced depolarization was tetrodotoxin insensitive and was blocked by the peptidergic CRF-R1 antagonist astressin. Moreover, activation of hypocretinergic neurons in response to acute stress was severely impaired in CRF-R1 knock-out mice. Together, our data provide evidence of a direct neuroanatomical and physiological input from CRF peptidergic system onto hypocretin neurons. We propose that, after stressor stimuli, CRF stimulates the release of hypocretins and that this circuit contributes to activation and maintenance of arousal associated with the stress response.

  View details for DOI 10.1523/JNEUROSCI.3459-04.2004

  View details for Web of Science ID 000225766200028

  View details for PubMedID 15601950

• Overexpression of the human beta-amyloid precursor protein downregulates cortistatin mRNA in PDAPP mice BRAIN RESEARCH Winsky-Sommerer, R., Spier, A. D., Fabre, V., De Lecea, L., Criado, J. R. 2004; 1023 (1): 157-162

  Abstract

  We measured preprocortistatin mRNA expression in young and aged transgenic (Tg) mice overexpressing the human beta-amyloid precursor protein (hbetaAPP) under the platelet-derived growth factor-beta promoter. Our findings suggest that the significant increase in hippocampal cortistatin mRNA expression during normal aging is significantly attenuated in Tg mice at an age known to exhibit beta-amyloid protein (Abeta) deposition. These deficits in cortistatin expression may play a role in the deficits in hippocampal-dependent spatial learning and sleep/wake states previously demonstrated in aged Tg mice.

  View details for DOI 10.1016/j.brainres.2004.04.082

  View details for Web of Science ID 000224166600020

  View details for PubMedID 15364032

• Chronic morphine treatment alters N-methyl-D-aspartate receptors in freshly isolated neurons from nucleus accumbens JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Martin, G., Guadano-Ferraz, A., Morte, B., Ahmed, S., Koob, G. F., De Lecea, L., Siggins, G. R. 2004; 311 (1): 265-273

  Abstract

  Although there is now evidence of a role for N-methyl-D-aspartate (NMDA) receptors in nucleus accumbens (NAcc) neurons in the effects of chronic opiate treatment, the cellular and molecular mechanisms underlying this phenomenon are still unclear. Therefore, we studied the effects of chronic morphine on the pharmacological and biophysical properties of NMDA receptors in freshly isolated medium spiny neurons from NAcc. We found that chronic morphine treatment did not alter the affinity for NMDA receptor agonists such as glutamate, homoquinolinic acid, and NMDA, but decreased the affinity of glycine, the allosteric NMDA receptor coagonist, from 2.24 +/- 0.15 microM to 5.1 +/- 1.45 microM. Chronic morphine treatment also altered the affinity of two noncompetitive NMDA receptor antagonists, 7-chloro-kynurenic acid and ifenprodil. However, morphine had no effect on a third antagonist, D-(-)-2-amino-5-phosphonopentanoic acid. Single-exponential fits of desensitized NMDA current tails gave tau values ranging from 0.5 to 4 s in neurons from both control and morphine-treated rats. However, a shift to the left of the distribution of tau values after morphine treatment revealed that NMDA current desensitization rate was accelerated in a majority of NAcc neurons. Taken together with our recent molecular studies, our data are consistent with a shift away from NMDA receptor subunit (NR) NR2B and 2C function toward increased NR2A subunit expression or function after chronic morphine, a process that could alter excitability and integrative properties and may represent a neuroadaptation of NAcc medium spiny neurons underlying morphine dependence.

  View details for DOI 10.1124/jpet.104.067504

  View details for Web of Science ID 000223896100031

  View details for PubMedID 15263066

• Neuropeptide S: A neuropeptide promoting arousal and anxiolytic-like effects NEURON Xu, Y. L., Reinscheid, R. K., Huitron-Resendiz, S., Clark, S. D., Wang, Z. W., Lin, S. H., Brucher, F. A., Zeng, J. A., Ly, N. K., Henriksen, S. J., Lecea, L. C., Civelli, O. 2004; 43 (4): 487-497

  Abstract

  Arousal and anxiety are behavioral responses that involve complex neurocircuitries and multiple neurochemical components. Here, we report that a neuropeptide, neuropeptide S (NPS), potently modulates wakefulness and could also regulate anxiety. NPS acts by activating its cognate receptor (NPSR) and inducing mobilization of intracellular Ca2+. The NPSR mRNA is widely distributed in the brain, including the amygdala and the midline thalamic nuclei. Central administration of NPS increases locomotor activity in mice and decreases paradoxical (REM) sleep and slow wave sleep in rats. NPS was further shown to produce anxiolytic-like effects in mice exposed to four different stressful paradigms. Interestingly, NPS is expressed in a previously undefined cluster of cells located between the locus coeruleus (LC) and Barrington's nucleus. These results indicate that NPS could be a new modulator of arousal and anxiety. They also show that the LC region encompasses distinct nuclei expressing different arousal-promoting neurotransmitters.

  View details for Web of Science ID 000223436400008

  View details for PubMedID 15312648

• Distribution of CNT2 and ENT1 transcripts in rat brain: selective decrease of CNT2 mRNA in the cerebral cortex of sleep-deprived rats JOURNAL OF NEUROCHEMISTRY Guillen-Gomez, E., Calbet, M., Casado, J., De Lecea, L., Soriano, E., Pastor-Anglada, M., Burgaya, F. 2004; 90 (4): 883-893

  Abstract

  Nucleoside transport processes regulate the levels of adenosine available to modulate neurotransmission, vascular tone and other physiological events. However, although equilibrative transporter transcripts or proteins have been mapped in the central nervous system of rats and humans, little is known about the presence and distribution of the complete family of nucleoside transporters in brain. In this study, we analysed the distribution of the transcript encoding the high affinity adenosine-preferring concentrative transporter CNT2 in the rat central nervous system and compared it with that of the equilibrative transporter ENT1. Furthermore, we evaluated the changes in expression of these two transporters in a situation of increased extracellular levels of adenosine, such as sleep deprivation. CNT2 mRNA was widespread in rat brain, although most prevalent in the amygdala, the hippocampus, specific neocortical regions and the cerebellum. The distribution of CNT2 mRNA only partially overlapped that of ENT1. Most of the cells labelled were neurones. Total sleep deprivation dramatically diminished the amounts of CNT2 mRNA, whereas ENT1 mRNA remained unchanged. This specific decrease in CNT2 transcript suggests a new physiological role for the transporter in the modulation of extracellular adenosine levels and the sleep/wakefulness cycle.

  View details for DOI 10.1111/j.1471-4159.2004.02545.x

  View details for Web of Science ID 000223034200014

  View details for PubMedID 15287894

• Not asleep, not quite awake NATURE MEDICINE Sutcliffe, J. G., de Lecea, L. 2004; 10 (7): 673-674

  View details for DOI 10.1038/nm0704-673

  View details for Web of Science ID 000222460400014

  View details for PubMedID 15229508

• A collection of cDNAs enriched in upper cortical layers of the embryonic mouse brain MOLECULAR BRAIN RESEARCH Garcia-Frigola, C., Burgaya, F., Calbet, M., Lopez-Guillenno, G., De Lecea, L., Soriano, E. 2004; 122 (2): 133-150

  Abstract

  In an attempt to elucidate the molecular basis of neuronal migration and corticogenesis, we performed subtractive hybridization of mRNAs from the upper cortical layers (layer I and upper cortical plate) against mRNAs from the remaining cerebral cortex at E15-E16. We obtained a collection of subtracted cDNA clones and analyzed their 3' UTR sequences, 47% of which correspond to EST sequences, and may represent novel products. Among the cloned sequences, we identified gene products that have not been reported in brain or in the cerebral cortex before. We examined the expression pattern of 39 subtracted clones, which was enriched in the upper layers of the cerebral cortex at embryonic stages. The expression of most clones is developmentally regulated, and especially high in embryonic and early postnatal stages. Four of the unknown clones were studied in more detail and identified as a new member of the tetraspanin superfamily, a putative RNA binding protein, a specific product of the adult dentate gyrus and a protein containing a beta-catenin repeat. We thus cloned a collection of subtracted cDNAs coding for protein products that may be involved in the development of the cerebral cortex.

  View details for DOI 10.1016/j.molbrainres.2003.12.014

  View details for Web of Science ID 000220334500005

  View details for PubMedID 15010206

• The role of the hypcretinergic system in the integration of networks that dictate the states of arousal DRUG NEWS & PERSPECTIVES Winsky-Sommerer, R., Boutrel, B., De Lecea, L. 2003; 16 (8): 504-512

  Abstract

  Recent studies have led to the discovery of a neuropeptide system that regulates arousal states. The hypocretins (hcrt1 and hcrt2, also called the orexins) are neuropeptides of related sequence derived from the same precursor whose expression is restricted to a few thousand neurons of the lateral hypothalamus. Two G-protein-coupled receptors for the hypocretins have been identified, and these have different distributions within the central nervous system and differential affinities for the two hypocretins. Hypocretin fibers project throughout the brain, including several areas implicated in cardiovascular function and regulation of the sleep-wake cycle. Central administration of synthetic hypocretin-1 affects blood pressure, hormone secretion and locomotor activity, and increases wakefulness while suppressing rapid eye movement sleep. Most human patients with narcolepsy have greatly reduced levels of hypocretin peptides in their cerebral spinal fluid and no or barely detectable hypocretin neurons in their hypothalami, suggestive of autoimmune attack. Development of nonpeptidergic hypocretin antagonists may prove useful in sleep disorders, whereas hypocretin agonists may be used to treat narcolepsy and excessive daytime sleepiness. The hypocretins are also an excellent target for the pharmacological treatment of the deregulated arousal state that characterizes depression or addictive behavior.

  View details for Web of Science ID 000187028700006

  View details for PubMedID 14668948

• Glutamatergic transmission in opiate and alcohol dependence Conference on Glutamate and Disorders of Cognition and Motivation Siggins, G. R., Martin, G., Roberto, M., Nie, Z. G., Madamba, S., De Lecea, L. NEW YORK ACAD SCIENCES. 2003: 196–211

  Abstract

  Both the nucleus accumbens (NAcc) and central amygdala (CeA) are thought to play roles in tolerance to, and dependence on, abused drugs. Although our past studies in rat brain slices suggested a role for NMDA receptors (NMDARs) in NAcc neurons in the effects of acute and chronic opiate treatment, the cellular and molecular mechanisms remained unclear. Therefore, we examined the effects of morphine dependence on electrophysiological properties of NMDARs in freshly isolated NAcc neurons and on expression of mRNA coding for NR2A-C subunits using single-cell RT-PCR. Chronic morphine did not alter the affinity for NMDAR agonists glutamate, homoquinolinate, or NMDA, but decreased the affinity of the coagonist glycine. Chronic morphine altered the NMDAR inhibition by two NMDAR antagonists, 7-Cl-kynurenate and ifenprodil, but not that by d-APV or Mg2+. Chronic morphine accelerated the NMDA current desensitization rate in NAcc neurons. In single-cell RT-PCR, chronic morphine predominantly reduced the number of neurons expressing multiple NR2 subunits. Ethanol also alters NMDARs. We found that low ethanol concentrations (IC50 = 13 mM) inhibited NMDA currents and NMDA-EPSPs in most NAcc neurons in a slice preparation. NAcc neurons from ethanol-dependent rats showed enhanced NMDA sensitivity. In CeA neurons, acute ethanol decreased (by 10-25%) non-NMDA- and NMDA-EPSPs in most neurons. In CeA neurons from ethanol-dependent rats, acute ethanol decreased the non-NMDA-EPSPs to the same extent as in naïve rats, but inhibited (by 30-40%) NMDA-EPSPs significantly more than in controls, suggesting sensitization to ethanol. Preliminary studies with microdialysis and real-time PCR analysis support this idea: local ethanol administration in vivo had no effect on glutamate release, but chronic ethanol nearly tripled the expression of NR2B subunits (the most ethanol sensitive) in CeA. These combined findings suggest that changes in glutamatergic transmission in NAcc and CeA may underlie the neuroadaptions that lead to opiate and ethanol dependence.

  View details for DOI 10.1196/annals.1300.012

  View details for Web of Science ID 000188893500017

  View details for PubMedID 14684447

• Targeted disruption of RC3 reveals a calmodulin-based mechanism for regulating metaplasticity in the hippocampus JOURNAL OF NEUROSCIENCE Krucker, T., Siggins, G. R., McNamara, R. K., LINDSLEY, K. A., Dao, A., Allison, D. W., De Lecea, L., Lovenberg, T. W., Sutcliffe, J. G., Gerendasy, D. D. 2002; 22 (13): 5525-5535

  Abstract

  We used homologous recombination in the mouse to knock-out RC3, a postsynaptic, calmodulin-binding PKC substrate. Mutant brains exhibited lower immunoreactivity to phospho-Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) but had the same synaptic density as wild type and did not exhibit a gross neuroanatomical phenotype. Basal excitatory synaptic transmission in CA1 was depressed, long-term potentiation (LTP) was enhanced, and the depressant effects of the metabotropic glutamate receptor (mGluR) agonist (RS)-3,5-dihydroxyphenylglycine was occluded compared with littermate controls. The frequency-response curve was displaced to the left, and long-term depression (LTD) could not be induced unless low-frequency stimuli were preceded by high-frequency tetani. Depotentiation was much more robust in the mutant, and only one stimulus was required to saturate LTD in primed mutant hippocampi, whereas multiple low-frequency stimuli were required in wild-type slices. Thus, ablation of RC3 appears to render the postsynaptic neuron hypersensitive to Ca(2+), decreasing its LTD and LTP thresholds and accentuating the effects of priming stimuli. We propose an mGluR-dependent CaM-based sliding threshold mechanism for metaplasticity that is governed by the phosphorylation states of RC3 and CaMKII.

  View details for Web of Science ID 000176599100032

  View details for PubMedID 12097504

• The hypocretins: Setting the arousal threshold NATURE REVIEWS NEUROSCIENCE Sutcliffe, J. G., De Lecea, L. 2002; 3 (5): 339-349

  Abstract

  Over a short period in the late 1990s, three groups converged on the discovery of a neuropeptide system, centred in the dorsolateral hypothalamus, that regulates arousal states, influences feeding and is implicated in the sleep disorder narcolepsy. Subsequent studies have illuminated many aspects of the circuitry of the hypocretin (also called orexin) system, which also influences hormone secretion and autonomic homeostasis, and have led to the hypothesis that most human narcolepsies result from an autoimmune attack against the hypocretin-producing neurons. The biochemical, physiological and anatomical components that regulate the switch between waking and sleeping are becoming clear. The rapidity with which the hypocretin story has emerged is a testament to both the conceptual and the technical evolution of genomic science in the past two decades.

  View details for DOI 10.1038/nrn808

  View details for Web of Science ID 000175350300015

  View details for PubMedID 11988773

• Hypocretins/orexins as integrators of physiological information: lessons from mutant animals NEUROPEPTIDES de Lecea, L., Sutcliffe, J. G., Fabre, V. 2002; 36 (2-3): 85-95

  Abstract

  The hypocretins/orexins (hcrts) are two recently described neuropeptides derived from the same precursor and expressed in a few thousand neurons in the perifornical area of the lateral hypothalamus, which project throughout the brain. The hypocretins bind to two G-protein coupled receptors with different selective affinities. Positional cloning of the gene responsible for a canine model of narcolepsy revealed that this disease is caused by mutations in hypocretin receptor type 2. Parallel studies with hypocretin/orexin knockout mice showed behavioral arrests reminiscent of narcolepsy-like attacks. Narcoleptic patients have decreased hypocretin-containing neurons suggesting that narcolepsy in humans is caused by selective neurodegeneration of hypocretinergic neurons. Additional functions for the hypocretins on regulation of energy balance neuroendocrine release and sympathetic outflow have been described. Here we review studies in humans and mutant animals that have provided clues about the functions of the hypocretinergic system, which appear to involve the coherent regulation of networks that dictate the states of arousal.

  View details for DOI 10.1054/npep.2002.0892

  View details for Web of Science ID 000178200500005

  View details for PubMedID 12359500

• Interaction of the hypocretins with neurotransmitters in the nucleus accumbens REGULATORY PEPTIDES Martin, G., Fabre, W., Siggins, G. R., de Lecea, L. 2002; 104 (1-3): 111-117

  Abstract

  The hypocretins (hcrt1 and hcrt2), also known as orexins, are two neuropeptides derived from the same precursor, expressed in a few thousand cells in the lateral hypothalamus. Hypocretin-containing cells project throughout the brain, including ascending projections to the olfactory bulb and cerebral cortex, through the medial septum and the nucleus accumbens. Here, we have studied the interactions of the hypocretins with different neurotransmitters by patch clamp recording of acutely dissociated cells from the nucleus accumbens. Application of hcrt1 or hcrt2 decreased postsynaptic NMDA currents, enhanced GABA currents but did not affect glycine-activated conductances. Our results strongly suggest that the hypocretin peptides may be inhibitory peptides, probably via binding hcrt receptor 2.

  View details for Web of Science ID 000174076900016

  View details for PubMedID 11830285

• Activation of hypocretin neurons and sleep 9th International Catecholamine Symposium Fabre, V., Bourgin, P., Sutcliffe, J. G., De Lecea, L. KLUWER ACADEMIC/PLENUM PUBL. 2002: 385–388

  View details for Web of Science ID 000180512100092

• Pattern of expression of the tetraspanin Tspan-5 during brain development in the mouse MECHANISMS OF DEVELOPMENT Garcia-Frigola, C., Burgaya, F., De Lecea, L., Soriano, E. 2001; 106 (1-2): 207-212

  Abstract

  Here, we report the pattern of expression of the tetraspanin mTspan-5 at various developmental stages. Expression was first seen at E10 and remained until adulthood, with increased levels between P0 and P5. mTspan-5 showed high expression in the cortical areas and Purkinje cells throughout development. Moreover, transcripts were also detected in the developing heart, forelimbs, and hindlimbs.

  View details for Web of Science ID 000170370900028

  View details for PubMedID 11472858

• Hypocretin (orexin) in the rat pineal gland: a central transmitter with effects on noradrenaline-induced release of melatonin EUROPEAN JOURNAL OF NEUROSCIENCE Mikkelsen, J. D., Hauser, F., deLecea, L., Sutcliffe, J. G., Kilduff, T. S., Calgari, C., Pevet, P., Simonneaux, V. 2001; 14 (3): 419-425

  Abstract

  Hypocretin-1 (HCRT-1) and hypocretin 2 (HCRT-2), also known as orexin-A and orexin-B, are two neuropeptides derived from the same precursor. Hypocretinergic neurons have been found exclusively in the hypothalamic dorsolateral area. These neurons are implicated in sleep and feeding through activation of specific G-protein-coupled orexin-1 and orexin-2 receptor (OR-R1 and OR-R2). The purpose of this study was to determine the existence of the HCRT peptides in the central input of the rat pineal gland. Further, OR-R1 and OR-R2 expression was determined in the pineal gland and the effect of HCRT-2 on melatonin synthesis and secretion was analysed in dissociated rat pinealocytes. A large contingent of HCRT-positive nerve fibres and terminals were observed in the epithalamus, many of which entered into the pineal parenchyma. A significant number of nerve fibres endowed with positive boutons were identified in the pineal stalk, though the number of positive fibres decreased along the extension of the stalk. So far, no positive fibres have been found in the superficial pineal gland. RT-PCR analysis revealed the expression of OR-R2 mRNA, whereas OR-R1-receptor mRNA was not detected. When tested alone, HCRT-2 had no effect on secretion of melatonin from cultured rat pinealocytes. However, HCRT-2 partially inhibited (by a maximum of 30%) the beta-adrenergic-induced melatonin secretion. The same effect was seen on activation of N-acetyltransferase activity. The distribution and the large number of HCRT-positive fibres together with the effect on noradrenaline-mediated melatonin release through specific receptors suggests that these peptides may be significant central transmitters in pineal function, probably mediating homeostatic signals to the pineal gland.

  View details for Web of Science ID 000171184400002

  View details for PubMedID 11553292

• Mapping of the mRNAs for the hypocretin/orexin and melanin-concentrating hormone receptors: Networks of overlapping peptide systems JOURNAL OF COMPARATIVE NEUROLOGY Kilduff, T. S., De Lecea, L. 2001; 435 (1): 1-5

  View details for Web of Science ID 000168841100001

  View details for PubMedID 11370007

• Immunohistochemical localization and biochemical characterization of hypocretin/orexin-related peptides in the central nervous system of the frog Rana ridibunda JOURNAL OF COMPARATIVE NEUROLOGY Galas, L., Vaudry, H., Braun, B., Van den Pol, A. N., De Lecea, L., Sutcliffe, J. G., Chartrel, N. 2001; 429 (2): 242-252

  Abstract

  In the present study, we have investigated the distribution and biochemical characteristics of hypocretin (hcrt) -like immunoreactivity in the central nervous system (CNS) of the frog Rana ridibunda by using an antiserum directed against rat hcrt2. Immunoreactive cell bodies were only detected in four diencephalic nuclei, including the anterior preoptic area and the suprachiasmatic, magnocellular, and ventral hypothalamic nuclei. In contrast, hcrt2-immunoreactive fibers were widely distributed throughout the frog CNS. In particular, a high density of hcrt-positive fibers was detected in several areas of the telencephalon, including the olfactory bulb, the nucleus of the diagonal band of Broca, and the amygdala. A dense network of hcrt-containing fibers was observed in all thalamic and hypothalamic nuclei. A low to moderate density of immunoreactive fibers was also found in the mesencephalon, rhombencephalon, and spinal cord. Reversed-phase high performance liquid chromatography analysis of frog brain extracts revealed that hcrt2-immunoreactive material eluted as two peaks, the major one exhibiting the same retention time as synthetic rat hcrt2. The present data provide the first detailed mapping of the hcrt neuronal system in the CNS of a nonmammalian vertebrate. The occurrence of hcrt-containing cell bodies in the hypothalamus and the widespread distribution of hcrt-immunoreactive fibers throughout the brain and spinal cord suggest that, in amphibians, hcrts may exert neuroendocrine, neurotransmitter, and/or neuromodulator activities.

  View details for Web of Science ID 000165642500005

  View details for PubMedID 11116217

• The hypocretins: Excitatory neuromodulatory peptides for multiple homeostatic systems, including sleep and feeding JOURNAL OF NEUROSCIENCE RESEARCH Sutcliffe, J. G., De Lecea, L. 2000; 62 (2): 161-168

  Abstract

  The hypocretins are two neuropeptides of related sequence that are produced from a common precursor whose expression is restricted to 1, 100 large neurons of the rat dorsal-lateral hypothalamus. The hypocretins have been detected immunohistochemically in secretory vesicles at synapses of fibers that project to areas within the posterior hypothalamus that are implicated in feeding behaviors and hormone secretion and diverse targets in other brain regions and in the spinal cord, including several areas implicated in cardiovascular function and sleep-wake regulation. The hypocretin-producing cells have receptors for leptin and receive input from arcuate neuropeptide Y neurons. The peptides are excitatory when applied to cultured hypothalamic, cortical, or spinal cord neurons. Two G protein-coupled receptors for the hypocretins have been identified, and these have different distributions within the CNS and differential affinities for the two hypocretins. Administration of the hypocretins stimulates food intake; affects blood pressure, hormone secretion, and locomotor activity; and increases wakefulness while suppressing REM sleep. The hypocretin mRNA accumulates during food deprivation. An inactivating insertion into the hypocretin receptor 2 gene in dogs results in narcolepsy. Mice whose hypocretin gene has been inactivated exhibit a narcolepsy-like phenotype. Human patients with narcolepsy have greatly reduced levels of hypocretin peptides in their cerebral spinal fluid. One aspect of hypocretin activity is the direct excitation of noradrenergic neurons in the locus coeruleus to prevent entry into REM sleep. These peptides appear to be part of a complex circuit that integrates aspects of energy metabolism, cardiovascular function, hormone homeostasis, and sleep-wake behaviors.

  View details for Web of Science ID 000089702300001

  View details for PubMedID 11020209

• Hypocretin-1 modulates rapid eye movement sleep through activation of locus coeruleus neurons JOURNAL OF NEUROSCIENCE Bourgin, P., Huitron-Resendiz, S., Spier, A. D., Fabre, V., Morte, B., Criado, J. R., Sutcliffe, J. G., Henriksen, S. J., de Lecea, L. 2000; 20 (20): 7760-7765

  Abstract

  The hypocretins (hcrts), also known as orexins, are two recently identified excitatory neuropeptides that in rat are produced by approximately 1200 neurons whose cell bodies are located in the lateral hypothalamus. The hypocretins/orexins have been implicated in the regulation of rapid eye movement (REM) sleep and the pathophysiology of narcolepsy. In the present study, we investigated whether the locus coeruleus (LC), a structure receiving dense hcrtergic innervation, which is quiescent during REM sleep, might be a target for hcrt to regulate REM sleep. Local administration of hcrt1 but not hcrt2 in the LC suppressed REM sleep in a dose-dependent manner and increased wakefulness at the expense of deep, slow-wave sleep. These effects were blocked with an antibody that neutralizes hcrt binding to hcrt receptor 1. In situ hybridization and immunocytochemistry showed the presence of hcrt receptor 1 but not the presence of hcrt receptor 2 in the LC. Iontophoretic application of hcrt1 enhanced the firing rate of LC neurons in vivo, and local injection of hcrt1 into the LC induced the expression of c-fos in the LC area. We propose that hcrt receptor 1 in the LC is a key target for REM sleep regulation and might be involved in the pathophysiological mechanisms of narcolepsy.

  View details for Web of Science ID 000089753300036

  View details for PubMedID 11027239

• Mouse Tspan-5, a member of the tetraspanin superfamily, is highly expressed in brain cortical structures NEUROREPORT Garcia-Frigola, C., Burgaya, F., Calbet, M., De Lecea, L., Soriano, E. 2000; 11 (14): 3181-3185

  Abstract

  Using a subtractive hybridization method for the identification of genes related to the development of the murine cerebral cortex, we cloned a mouse homologue of a human tetraspanin family member, Tspan-5. We have isolated a 3.1 Kb cDNA fragment containing the entire coding region. Analysis of the cDNA nucleotide sequence revealed that mouse Tspan-5 shares 98% amino acid sequence identity with its human homologue. The predicted length of the mouse protein is 268 amino acids, with four putative hydrophobic domains with N- and C-intracellular tails, and two extracellular domains. Northern blot analysis of adult mouse tissues showed a single transcript, which is preferentially expressed in the brain. In situ hybridization showed prominent expression of Tspan-5 in the neocortex, the hippocampus, amygdala and in Purkinje cells in the cerebellum. The pattern of expression of Tspan-5 in the mouse brain suggests a role for the tetraspanins in the maintenance of adult brain function.

  View details for Web of Science ID 000089833500028

  View details for PubMedID 11043545

• Cortistatin: a member of the somatostatin neuropeptide family with distinct physiological functions BRAIN RESEARCH REVIEWS Spier, A. D., de Lecea, L. 2000; 33 (2-3): 228-241

  Abstract

  Cortistatin is a recently discovered neuropeptide relative of somatostatin named after its predominantly cortical expression and ability to depress cortical activity. Cortistatin-14 shares 11 of the 14 amino acids of somatostatin-14 yet their nucleotide sequences and chromosomal localization clearly indicate they are products of separate genes. Now cloned from human, mouse and rat sources, cortistatin is known to bind all five cloned somatostatin receptors and share many pharmacological and functional properties with somatostatin including the depression of neuronal activity. However, cortistatin also has many properties distinct from somatostatin including induction of slow-wave sleep, apparently by antagonism of the excitatory effects of acetylcholine on the cortex, reduction of locomotor activity, and activation of cation selective currents not responsive to somatostatin. Expression of mRNA encoding cortistatin follows a circadian rhythm and is upregulated on deprivation of sleep, suggesting cortistatin is a sleep modulatory factor. This review summarizes recent advances in our understanding of the neurobiology of cortistatin, examines the similarities and differences between cortistatin and somatostatin, and asks the question: does cortistatin bind to a cortistatin-specific receptor?

  View details for Web of Science ID 000165360200006

  View details for PubMedID 11011067

• Developmental regulation of two isoforms of Ca2+/calmodulin-dependent protein kinase I beta in rat brain BRAIN RESEARCH Loseth, O. P., De Lecea, L., Calbet, M., Danielson, P. E., Gautvik, V., Hovring, P. I., Walaas, S. I., Gautvik, K. M. 2000; 869 (1-2): 137-145

  Abstract

  Subtractive hybridization analysis of region-specific gene expression in brain has demonstrated a mRNA species enriched in rat hypothalamus [K.M. Gautvik, L. de Lecea, V.T. Gautvik, P.E. Danielson, P. Tranque, A. Dopazo, F.E. Bloom, J.G. Sutcliffe, Proc. Natl. Acad. Sci. USA 93 (1996) 8733-8738.]. We here show that this mRNA encodes a Ca(2+)/calmodulin-dependent (CaM) kinase belonging in the CaM kinase I beta subgroup. cDNA analysis showed that this enzyme was differentially spliced into two isoforms (designated beta1 and beta2) with distinct C-termini. The C-terminal of the translated CaM kinase I beta2 protein (38.5 kDa molecular size), contained 25 amino acid residues not present in the beta1 isoform. The two isoforms were differentially developmentally regulated, with the beta1 isoform being present in rat embryos from day 18 and the beta2 isoform being present from day 5 postnatally. In situ hybridization analysis of adult rat CNS showed CaM kinase I beta2 mRNA being enriched in the hypothalamus and the hippocampal formation. Expression was also observed in a number of ventral limbic structures and in the thalamus. Northern blot analysis showed additional expression of multiple beta2 isoforms in heart and skeletal muscle. The human mRNA showed a similar distribution. Our data suggest that the two isoforms of CaM kinase I beta, created by a splicing process occurring within a week around birth, may have distinct pre- and postnatal functions in a distinct set of CNS neurons and excitable tissues.

  View details for Web of Science ID 000087990300017

  View details for PubMedID 10865068

• The hypocretins/orexins: novel hypothalamic neuropeptides involved in different physiological systems CELLULAR AND MOLECULAR LIFE SCIENCES de Lecea, L., Sutcliffe, J. G. 1999; 56 (5-6): 473-480

  Abstract

  The hypothalamus regulates diverse physiological functions, including the control of energy metabolism, circadian rhythms, stress and anxiety, sexual and reproductive behaviors. An overview of the most prevalent hypothalamus-enriched mRNAs revealed that this area of the brain specializes in producing intercellular signaling molecules. Two new secreted peptides derived from a single neuropeptide precursor, named hypocretins and orexins by two different groups, are synthesized in a small set of neurons in the perifornical area of the hypothalamus. Intracerebroventricular injection of the hypocretins/orexins increases food consumption in rats. Here we review recent progress in identifying the role of the hypocretins/orexins in the control of energy balance and in other physiological systems.

  View details for Web of Science ID 000083612100009

  View details for PubMedID 11212299

• Cortistatin and somatostatin mRNAs are differentially regulated in response to kainate MOLECULAR BRAIN RESEARCH Calbet, M., Guadano-Ferraz, A., Spier, A. D., Maj, M., Sutcliffe, J. G., Przewlocki, R., de Lecea, L. 1999; 72 (1): 55-64

  Abstract

  Cortistatin (CST) is a presumptive neuropeptide that shares 11 of its 14 amino acids with somatostatin (SST). CST and SST are expressed in partially overlapping but distinct populations of cortical interneurons. In the hippocampal formation, most CST-positive cells are also positive for SST. In contrast to SST, administration of CST into the rat brain ventricles reduces locomotor activity and specifically enhances slow wave sleep. Intracerebroventricular injection of CST or SST has been shown to protect against the neurotoxic effects of kainic acid. Here, we show that CST and SST mRNAs respond differently to kainate-induced seizures. Furthermore, comparison of the upstream sequences from the CST and SST precursor genes reveal that they contain binding motifs for different transcriptional regulatory factors. Our data demonstrate that CST and SST, which are often co-expressed in the same neurons, are regulated by different stimuli.

  View details for Web of Science ID 000082766900006

  View details for PubMedID 10521599

• Leptin receptor- and STAT3-immunoreactivities in hypocretin/orexin neurones of the lateral hypothalamus JOURNAL OF NEUROENDOCRINOLOGY Hakansson, M. L., De Lecea, L., Sutcliffe, J. G., Yanagisawa, M., Meister, B. 1999; 11 (8): 653-663

  Abstract

  Hypocretins/orexins are recently characterized peptides that are synthesized in neurones of the lateral hypohalamus and stimulate food intake in rats. To clarify whether leptin may interact with hypocretin/orexin to reduce ingestive behaviour, the presence of leptin receptor-immunoreactivity in hypocretin/orexin-containing neurones was examined. Many leptin receptor-and hypocretin/orexin-immunoreactive neurones were demonstrated in the lateral hypothalamic area and perifornical region. Both direct double-labelling and elution-restaining methods showed that leptin receptor-immunoreactivity was present in the vast majority of hypocretin/orexin-containing neurones. Immunoreactivity for STAT3, a transcription factor activated by leptin, was also demonstrated in hypocretin/orexin-containing neurones. Isolated hypocretin/orexin cell bodies in the dorsal part of the lateral hypothalamic area and the ventral perifornical region were shown to contain immunoreactivity for galanin, another peptide known to affect feeding. Galanin neurones were also seen to contain leptin receptor-and STAT3-immunoreactivity. Melanin-concentrating hormone (MCH)-containing neurones constituted a cell population within the lateral hypothalamus distinct from the one containing hypocretin/orexin-immunoreactivity, as shown by elution-restaining methodology. The presence of leptin receptor-and STAT3-immunoreactivities in hypocretin/orexin-containing neurones of the lateral hypothalamus suggests that leptin may directly regulate these hypothalamic neurones, most likely via an inhibitory action on hypocretin/orexin expression and/or secretion resulting in reduced food intake.

  View details for Web of Science ID 000081806200010

  View details for PubMedID 10447804

• OCD-like behaviors caused by a neuropotentiating transgene targeted to cortical and limbic D1+neurons JOURNAL OF NEUROSCIENCE Campbell, K. M., De Lecea, L., Severynse, D. M., Caron, M. G., McGrath, M. J., Sparber, S. B., Sun, L. Y., Burton, F. H. 1999; 19 (12): 5044-5053

  Abstract

  To study the behavioral role of neurons containing the D1 dopamine receptor (D1+), we have used a genetic neurostimulatory approach. We generated transgenic mice that express an intracellular form of cholera toxin (CT), a neuropotentiating enzyme that chronically activates stimulatory G-protein (Gs) signal transduction and cAMP synthesis, under the control of the D1 promoter. Because the D1 promoter, like other CNS-expressed promoters, confers transgene expression that is regionally restricted to different D1+ CNS subsets in different transgenic lines, we observed distinct but related psychomotor disorders in different D1CT-expressing founders. In a D1CT line in which transgene expression was restricted to the following D1+ CNS regions-the piriform cortex layer II, layers II-III of somatosensory cortical areas, and the intercalated nucleus of the amygdala-D1CT mice showed normal CNS and D1+ neural architecture but increased cAMP content in whole extracts of the piriform and somatosensory cortex. These mice also exhibited a constellation of compulsive behavioral abnormalities that strongly resembled human cortical-limbic-induced compulsive disorders such as obsessive-compulsive disorder (OCD). These compulsive behaviors included episodes of perseverance or repetition of any and all normal behaviors, repetitive nonaggressive biting of siblings during grooming, and repetitive leaping. These results suggest that chronic potentiation of cortical and limbic D1+ neurons thought to induce glutamatergic output to the striatum causes behaviors reminiscent of those in human cortical-limbic-induced compulsive disorders.

  View details for Web of Science ID 000080753800035

  View details for PubMedID 10366637

• Structural and compositional determinants of cortistatin activity JOURNAL OF NEUROSCIENCE RESEARCH Criado, J. R., Li, H. T., Jiang, X. H., Spina, M., Huitron-Resendiz, S., Liapakis, G., Calbet, M., Siehler, S., Henriksen, S. J., Koob, G., Hoyer, D., Sutcliffe, J. G., Goodman, M., De Lecea, L. 1999; 56 (6): 611-619

  Abstract

  Cortistatin-14 (CST-14) is a putative novel neuropeptide that shares 11 of its 14 residues with somatostatin-14 (SRIF-14), yet its effects on sleep physiology, locomotor behavior and hippocampal function are different from those of somatostatin. We studied the structural basis for cortistatin's distinct biological activities. As with SRIF-14, CST-14 does not show any preferred conformation in solution, as determined by circular dichroism and nuclear magnetic resonance. Synthetic cortistatin analogs were designed and synthesized based on the cyclic structure of octreotide. Biological assays were carried out to determine their binding affinities to five somatostatin receptors (sstl-5) and their ability to produce changes in locomotor activity and to modulate hippocampal physiology and sleep. The results show that the compound with N-terminal proline and C-terminal lysine amide exhibits cortistatin-like biological activities, including reduction of population spike amplitudes in the hippocampal CA1 region, decrease in locomotor activity and enhancement of slow-wave sleep 2. These findings suggest that both proline and lysine are necessary for cortistatin binding to its specific receptor.

  View details for Web of Science ID 000080526700007

  View details for PubMedID 10374816

• Cortistatin affects glutamate sensitivity in mouse hypothalamic neurons through activation of sst2 somatostatin receptor subtype NEUROSCIENCE Vasilaki, A., Lanneau, C., Dournaud, P., De Lecea, L., Gardette, R., Epelbaum, J. 1999; 88 (2): 359-364

  Abstract

  Cortistatin is a 14-residue putative neuropeptide with strong structural similarity to somatostatin. Even if it shares several biological properties with somatostatin, the effects of cortistatin on cortical electrical activity and sleep are opposite to those elicited by somatostatin. We recently demonstrated that somatostatin could modulate glutamate sensitivity, either positively through activation of the sstl receptor subtype, or negatively through activation of the sst2 receptor subtype in hypothalamic neurons in culture which express almost exclusively these two sst subtypes. Thus, in the present study we compared the effects of cortistatin and somatostatin in hypothalamic neurons in culture, in order to define the former peptide activity on both subtypes. We first determined that the affinities of cortistatin and somatostatin were similar on cloned rat sstl and sst2 receptor subtypes in transfected cells and hypothalamic neurons membranes. We then found that cortistatin, like somatostatin, depresses the glutamate response but, unlike somatostatin, never potentiates glutamate sensitivity in hypothalamic neurons. The observed effect of cortistatin is strongly suggestive of an activation of the somatostatin sst2 receptor subtype in hypothalamic neurons in culture.

  View details for Web of Science ID 000076506700004

  View details for PubMedID 10197759

• Novel neurotransmitters for sleep and energy homeostasis. Results and problems in cell differentiation Sutcliffe, J. G., De Lecea, L. 1999; 26: 239-255

  Abstract

  We have developed methodologies for identifying mRNAs with highly restricted expression within the brain. One postnatal-onset mRNA, restricted to sparse GABAergic interneurons of the cerebral cortex and hippocampus, encodes preprocortistatin, the precursor of a 14-residue peptide that shares 11 amino acids with somatostatin. Cortistatin binds to all five cloned somatostatin receptors when they are expressed in transfected cells and depresses neuronal activity, but, unlike somatostatin, it reduces locomotor activity and induces slow-wave sleep. Cortistatin, whose mRNA accumulates during sleep deprivation, apparently acts by antagonizing the effects of acetylcholine on cortical excitability, thereby causing synchronization brain slow waves. A single amino acid difference with somatostatin accounts for the dramatic differences in the effects of the two peptides on physiology and behavior. A second postnatal-onset mRNA, restricted to 1100 large neuronal cell bodies of the dorsal-lateral hypothalamus, encodes preprohypocretin, the precursor of two peptides that share homology with each other and with members of the secretin peptide family. The peptides are detected immunohistochemically in secretory vesicles at synapses of fibers that project to posterior hypothalamus and diverse targets in other brain regions. The peptides are excitatory when applied to cultured hypothalamic neurons. Recent studies by Sakurai and colleagues (1998) have identified the hypocretin peptides (called the orexins by those workers) as ligands for two orphan receptors at which they stimulate food-intake behavior. Sakurai and collaborators showed that the mRNA for these peptides accumulates during food deprivation. The hypocretin projections suggest additional homeostatic roles for the peptides. These studies suggest the common mechanism of regulation for necessary, but voluntary, behaviors (sleep and feeding) by transcription-based accumulation of peptide transmitters that create a pressure for the voluntary activities.

  View details for PubMedID 10453467

• Hypocretin/orexin- and melanin-concentrating hormone-expressing cells form distinct populations in the rodent lateral hypothalamus: Relationship to the neuropeptide Y and Agouti gene-related protein systems JOURNAL OF COMPARATIVE NEUROLOGY Broberger, C., De Lecea, L., Sutcliffe, J. G., Hokfelt, T. 1998; 402 (4): 460-474

  Abstract

  Cells in the lateral hypothalamus and in the arcuate nucleus play prominent roles in the central control of food intake; however, a neurochemical link connecting these potential components of a hypothalamic circuitry regulating energy metabolism remains to be established. In the present study, the topographical relationship between cells expressing mRNAs encoding melanin-concentrating hormone and the newly discovered neuropeptide family hypocretins/orexins was studied in the rat and mouse lateral hypothalamus by using double-labeling in situ hybridization. Cells expressing the two mRNAs formed completely distinct populations, with hypocretin/orexin cells located primarily perifornically and in the magnocellular lateral hypothalamic nucleus; melanin-concentrating hormone cells extended in a wider area both laterally and periventricularly and appeared to partly surround the hypocretin/orexin population. In the arcuate nucleus, cells expressing neuropeptide Y and agouti gene-related protein were studied by routine fluorescence and/or confocal microscopy immunohistochemistry. Double staining demonstrated that a large proportion of the neuropeptide Y-positive cell bodies in this nucleus also contained agouti gene-related protein-like immunoreactivity. Moreover, these two peptides also coexisted in nerve terminals surrounding and in close relationship to perikarya and processes of both hypocretin/orexin- and melanin-concentrating hormone-immunoreactive cells in the lateral hypothalamus, whereby the former appeared to receive a more dense innervation. These results thus provide evidence for an arcuate-lateral hypothalamic neuropeptide Y/agouti gene-related protein pathway. Furthermore, the results implicate hypocretin/orexin and melanin-concentrating hormone-expressing cells as downstream targets in neuropeptide Y-induced feeding.

  View details for Web of Science ID 000077534100003

  View details for PubMedID 9862321

• Neurons containing hypocretin (orexin) project to multiple neuronal systems JOURNAL OF NEUROSCIENCE Peyron, C., Tighe, D. K., Van den Pol, A. N., De Lecea, L., Heller, H. C., Sutcliffe, J. G., Kilduff, T. S. 1998; 18 (23): 9996-10015

  Abstract

  The novel neuropeptides called hypocretins (orexins) have recently been identified as being localized exclusively in cell bodies in a subregion of the tuberal part of the hypothalamus. The structure of the hypocretins, their accumulation in vesicles of axon terminals, and their excitatory effect on cultured hypothalamic neurons suggest that the hypocretins function in intercellular communication. To characterize these peptides further and to help understand what physiological functions they may serve, we undertook an immunohistochemical study to examine the distribution of preprohypocretin-immunoreactive neurons and fibers in the rat brain. Preprohypocretin-positive neurons were found in the perifornical nucleus and in the dorsal and lateral hypothalamic areas. These cells were distinct from those that express melanin-concentrating hormone. Although they represent a restricted group of cells, their projections were widely distributed in the brain. We observed labeled fibers throughout the hypothalamus. The densest extrahypothalamic projection was found in the locus coeruleus. Fibers were also seen in the septal nuclei, the bed nucleus of the stria terminalis, the paraventricular and reuniens nuclei of the thalamus, the zona incerta, the subthalamic nucleus, the central gray, the substantia nigra, the raphe nuclei, the parabrachial area, the medullary reticular formation, and the nucleus of the solitary tract. Less prominent projections were found in cortical regions, central and anterior amygdaloid nuclei, and the olfactory bulb. These results suggest that hypocretins are likely to have a role in physiological functions in addition to food intake such as regulation of blood pressure, the neuroendocrine system, body temperature, and the sleep-waking cycle.

  View details for Web of Science ID 000077169800038

  View details for PubMedID 9822755

• Regional and cellular patterns of reelin mRNA expression in the forebrain of the developing and adult mouse JOURNAL OF NEUROSCIENCE Alcantara, S., Ruiz, M., D'Arcangelo, G., Ezan, F., De Lecea, L., Curran, T., Sotelo, C., Soriano, E. 1998; 18 (19): 7779-7799

  Abstract

  The reelin gene encodes an extracellular protein that is crucial for neuronal migration in laminated brain regions. To gain insights into the functions of Reelin, we performed high-resolution in situ hybridization analyses to determine the pattern of reelin expression in the developing forebrain of the mouse. We also performed double-labeling studies with several markers, including calcium-binding proteins, GAD65/67, and neuropeptides, to characterize the neuronal subsets that express reelin transcripts. reelin expression was detected at embryonic day 10 and later in the forebrain, with a distribution that is consistent with the prosomeric model of forebrain regionalization. In the diencephalon, expression was restricted to transverse and longitudinal domains that delineated boundaries between neuromeres. During embryogenesis, reelin was detected in the cerebral cortex in Cajal-Retzius cells but not in the GABAergic neurons of layer I. At prenatal stages, reelin was also expressed in the olfactory bulb, and striatum and in restricted nuclei in the ventral telencephalon, hypothalamus, thalamus, and pretectum. At postnatal stages, reelin transcripts gradually disappeared from Cajal-Retzius cells, at the same time as they appeared in subsets of GABAergic neurons distributed throughout neocortical and hippocampal layers. In other telencephalic and diencephalic regions, reelin expression decreased steadily during the postnatal period. In the adult, there was prominent expression in the olfactory bulb and cerebral cortex, where it was restricted to subsets of GABAergic interneurons that co-expressed calbindin, calretinin, neuropeptide Y, and somatostatin. This complex pattern of cellular and regional expression is consistent with Reelin having multiple roles in brain development and adult brain function.

  View details for Web of Science ID 000076060100016

  View details for PubMedID 9742148

• Endogenous protein kinase A inhibitor (PKI alpha) modulates synaptic activity JOURNAL OF NEUROSCIENCE RESEARCH De Lecea, L., Criado, J. R., Rivera, S., Wen, W., Soriano, E., Henriksen, S. J., Taylor, S. S., Gall, C. M., Sutcliffe, J. G. 1998; 53 (3): 269-278

  Abstract

  Protein kinase A (PKA) has long been known to be involved in major regulatory mechanisms underlying synaptic plasticity and complex behaviors such as learning and memory. The endogenous PKA inhibitor, PKIalpha, has been extensively studied for its effects on PKA and PKA-mediated signal transduction. Clear functions for PKIalpha in vivo, however, remain to be established. Here we describe that several forms of synaptic stimulation in the rat hippocampus cause a dramatic decrease in the concentration of PKIalpha in dentate granule cells. Furthermore, chronic infusion of antisense oligonucleotides against PKIalpha into the rat brain results in a dramatic reduction of the excitability of these neurons and elimination of their ability to exhibit long-term potentiation (LTP) and long-term depression (LTD), suggesting a stimulus-dependent regulatory role for PKIalpha in PKA signal transduction.

  View details for Web of Science ID 000074995100001

  View details for PubMedID 9698155

• The hypocretins: Hypothalamus-specific peptides with neuroexcitatory activity PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA De Lecea, L., Kilduff, T. S., Peyron, C., Gao, X. B., Foye, P. E., Danielson, P. E., Fukuhara, C., Battenberg, E. L., Gautvik, V. T., Bartlett, F. S., Frankel, W. N., Van den Pol, A. N., Bloom, F. E., Gautvik, K. M., Sutcliffe, J. G. 1998; 95 (1): 322-327

  Abstract

  We describe a hypothalamus-specific mRNA that encodes preprohypocretin, the putative precursor of a pair of peptides that share substantial amino acid identities with the gut hormone secretin. The hypocretin (Hcrt) protein products are restricted to neuronal cell bodies of the dorsal and lateral hypothalamic areas. The fibers of these neurons are widespread throughout the posterior hypothalamus and project to multiple targets in other areas, including brainstem and thalamus. Hcrt immunoreactivity is associated with large granular vesicles at synapses. One of the Hcrt peptides was excitatory when applied to cultured, synaptically coupled hypothalamic neurons, but not hippocampal neurons. These observations suggest that the hypocretins function within the CNS as neurotransmitters.

  View details for Web of Science ID 000071429500062

  View details for PubMedID 9419374

• Cortistatin is expressed in a distinct subset of cortical interneurons JOURNAL OF NEUROSCIENCE deLecea, L., delRio, J. A., Criado, J. R., Alcantara, S., Morales, M., Danielson, P. E., Henriksen, S. J., Soriano, E., Sutcliffe, J. G. 1997; 17 (15): 5868-5880

  Abstract

  Cortistatin is a presumptive neuropeptide that shares 11 of its 14 amino acids with somatostatin. In contrast to somatostatin, administration of cortistatin into the rat brain ventricles specifically enhances slow wave sleep, apparently by antagonizing the effects of acetylcholine on cortical excitability. Here we show that preprocortistatin mRNA is expressed in a subset of GABAergic cells in the cortex and hippocampus that partially overlap with those containing somatostatin. A significant percentage of cortistatin-positive neurons is also positive for parvalbumin. In contrast, no colocalization was found between cortistatin and calretinin, cholecystokinin, or vasoactive intestinal peptide. During development there is a transient increase in cortistatin-expressing cells in the second postnatal week in all cortical areas and in the dentate gyrus. A transient expression of preprocortistatin mRNA in the hilar region at P16 is paralleled by electrophysiological changes in dentate granule cells. Together, these observations suggest mechanisms by which cortistatin may regulate cortical activity.

  View details for Web of Science ID A1997XU91800020

  View details for PubMedID 9221784

• Cloning, mRNA expression, and chromosomal mapping of mouse and human preprocortistatin GENOMICS deLecea, L., RUIZLOZANO, P., Danielson, P. E., PeelleKirley, J., Foye, P. E., Frankel, W. N., Sutcliffe, J. G. 1997; 42 (3): 499-506

  Abstract

  Cortistatin is a 14-residue putative neuropeptide with strong structural similarity to somatostatin and is expressed predominantly in cortical GABAergic interneurons of rats. Administration of cortistatin into the brain ventricles specifically enhances slow-wave sleep, presumably by antagonizing the effects of acetylcholine on cortical excitability. Here we report the identification of cDNAs corresponding to mouse and human preprocortistatin and the mRNA distribution and gene mapping of mouse cortistatin. Analysis of the nucleotide and predicted amino acid sequences from rat and mouse reveals that the 14 C-terminal residues of preprocortistatin, which make up the sequence that is most similar to somatostatin, are conserved between species. Lack of conservation of other dibasic amino acid residues whose cleavage by prohormone convertases would give rise to additional peptides suggests that cortistatin-14 is the only active peptide derived from the precursor. As in the rat, mouse preprocortistatin mRNA is present in GABAergic interneurons in the cerebral cortex and hippocampus. The preprocortistatin gene maps to mouse chromosome 4, in a region showing conserved synteny with human 1p36. The human putative cortistatin peptide has an arginine for lysine substitution, compared to the rat and mouse products, and is N-terminally extended by 3 amino acids.

  View details for Web of Science ID A1997XG51900018

  View details for PubMedID 9205124

• Peptides, sleep and cortistatin MOLECULAR PSYCHIATRY deLecea, L., Sutcliffe, J. G. 1996; 1 (5): 349-351

  View details for Web of Science ID A1996VV91100002

  View details for PubMedID 9154223

• The type 3 serotonin receptor is expressed in a subpopulation of GABAergic neurons in the rat neocortex and hippocampus BRAIN RESEARCH Morales, M., Battenberg, E., deLecea, L., Bloom, F. E. 1996; 731 (1-2): 199-202

  Abstract

  We used in situ hybridization and immunocytochemistry to investigate the presence of GABA in neurons that express the type 3 serotonin receptor (5-HT3R). Quantitative analysis indicated that more than 90% of 5-HT3R expressing cells are GABAergic in the neocortex and hippocampus. The co-existence of 5-HT3R and GABA in cortical and hippocampal neurons indicates that serotonin, via 5-HT3R, can affect GABA release and suggests the participation of 5-HT3R in the inhibitory regulation of forebrain neurons.

  View details for Web of Science ID A1996VH44800023

  View details for PubMedID 8883870

• Overview of the most prevalent hypothalamus-specific mRNAs, as identified by directional tag PCR subtraction PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Gautvik, K. M., deLecea, L., Gautvik, V. T., Danielson, P. E., Tranque, P., Dopazo, A., Bloom, F. E., Sutcliffe, J. G. 1996; 93 (16): 8733-8738

  Abstract

  We applied the directional tag PCR subtractive hybridization method to construct a rat hypothalamic cDNA library from which cerebellar and hippocampal sequences had been depleted, enriching 20-30-fold for sequences expressed selectively in the hypothalamus. We studied a sample of 94 clones selected for enrichment in the subtracted library. These clones corresponded to 43 distinct mRNA species, about half of which were novel. Thirty-eight of these 43 mRNAs (corresponding to 85 of the clones in the sample) exhibited enrichment in the hypothalamus; 23 were highly enriched. In situ hybridization studies revealed that one novel species was restricted to cells in a small bilaterally symmetric area of the paraventricular hypothalamus. Other novel mRNAs showed substantial enrichment in basal diencephalic structures, particularly the hypothalamus, without restriction to single hypothalamic nuclei. The data suggest that the hypothalamus utilizes at least two distinct strategies for employing its selectively expressed proteins. Secretory neuropeptides utilized for intercellular communication are produced by functionally discrete nuclei, while several other proteins are shared by structures that are unrelated in their physiological roles but may share biochemical systems.

  View details for Web of Science ID A1996VB32500105

  View details for PubMedID 8710940

• Transient colocalization of parvalbumin and calbindin D28k in the postnatal cerebral cortex: Evidence for a phenotypic shift in developing nonpyramidal neurons EUROPEAN JOURNAL OF NEUROSCIENCE Alcantara, S., deLecea, L., delRio, J. A., Ferrer, I., Soriano, E. 1996; 8 (7): 1329-1339

  Abstract

  In the adult rat cerebral cortex the calcium-binding proteins parvalbumin and calbindin D28k are present in essentially non-overlapping populations of GABAergic interneurons. These proteins follow different developmental patterns in the cortex: calbindin D28k-immunoreactive nonpyramidal neurons are abundant until the second postnatal week and decrease markedly thereafter; it is at this time that parvalbumin immunoreactivity develops in cortical nonpyramidal neurons. To determine whether parvalbumin-immunoreactive neurons derive from calbindin D38k positive cells we used double-immunofluorescence studies for both calcium-binding proteins, together with combined immunocytochemistry for calbindin D28k and in situ hybridization for parvalbumin mRNA during postnatal development. Double-labelled cells were found in all cortical layers between P9 and P21, coinciding with the onset of parvalbumin expression. The percentage of colocalization of the two calcium-binding proteins depended on the age and layer examined. Colocalization reached a peak (80-100%) during the second postnatal week. Double-labelled neurons were rare in layer V at all ages studied. The present results indicate a phenotypic shift during the development of some cortical interneurons that halts the expression of calbindin D28k while parvalbumin expression starts. These findings agree with lineage analyses reporting that different types of nonpyramidal neuron arise from a common progenitor.

  View details for Web of Science ID A1996VD72200003

  View details for PubMedID 8758940

• Expression of NGF and NT3 mRNAs in hippocampal interneurons innervated by the GABAergic septohippocampal pathway JOURNAL OF NEUROSCIENCE Rocamora, N., Pascual, M., Acsady, L., deLecea, L., Freund, T. F., Soriano, E. 1996; 16 (12): 3991-4004

  Abstract

  We used in situ hybridization for the detection of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin 3 (NT3) mRNAs combined with immunocytochemistry against the calcium-binding proteins parvalbumin (PARV), calbindin 28k (CALB), and calretinin (CALR) to determine the expression of neurotrophins in functionally distinct subsets of hippocampal interneurons. Most PARV-immunoreactive neurons in the hippocampus were NGF mRNA-positive (82%), which corresponds to 71% of NGF-positive neurons in the hippocampus proper and in the dentate gyrus (excluding granule cells). In contrast, only a subset of CALB- and CALR-immunoreactive interneurons (24% and 23%, respectively) displayed hybridization signals for NGF. Small subsets of PARV- and CALR-positive cells expressed NT3 mRNA, but we did not find hippocampal interneurons expressing BDNF mRNA. These results show that NGF and NT3 genes are differentially regulated in distinct subsets of GABAergic cells, and these interneurons are a major source of NGF production in the hippocampus. We also addressed whether hippocampal interneurons expressing neurotrophins were targets of the GABAergic septohippocampal pathway. We developed a triple-labeling method that combines anterograde tracing of this pathway by means of Phaseolus vulgaris leucoagglutinin injections, with in situ hybridization for the detection of neurotrophins, and immunocytochemistry for calcium-binding proteins. Virtually every PARV-positive neuron innervated by GABAergic septohippocampal baskets expressed NGF mRNA (86%), whereas 39-59% of CALR- and CALB-positive interneurons that were contacted by GABAergic septohippocampal axons showed NGF gene expression. A small subset of NT3 mRNA-expressing interneurons was also innervated by septohippocampal baskets. These findings show that the GABAergic septohippocampal pathway preferentially terminates on interneurons expressing NGF mRNA, suggesting that this neurotrophic factor might be involved in the specification of this connection and in its maintenance and normal function in the adult brain.

  View details for Web of Science ID A1996UP38300021

  View details for PubMedID 8656293

• A cortical neuropeptide with neuronal depressant and sleep-modulating properties NATURE deLecea, L., Criado, J. R., PROSPEROGARCIA, O., Gautvik, K. M., Schweitzer, P., Danielson, P. E., Dunlop, C. L., Siggins, G. R., Henriksen, S. J., Sutcliffe, J. G. 1996; 381 (6579): 242-245

  Abstract

  Acetylcholine (ACh) plays a key role in the transitions between the different phases of sleep: Slow-wave sleep requires low ACh concentrations in the brain, whereas rapid-eye-movement (REM) sleep is associated with high levels of ACh. Also, these phases of sleep are differentially sensitive to a number of endogenous neuropeptides and cytokines, including somatostatin, which has been shown to increase REM sleep without significantly affecting other phases. Here we report the cloning and initial characterization of cortistatin, a neuropeptide that exhibits strong structural similarity to somatostatin, although it is the product of a different gene. Administration of cortistatin depresses neuronal electrical activity but, unlike somatostatin, induces low-frequency waves in the cerebral cortex and antagonizes the effects of acetylcholine on hippocampal and cortical measures of excitability. This suggests a mechanism for cortical synchronization related to sleep.

  View details for Web of Science ID A1996UL24900056

  View details for PubMedID 8622767

• Cell-specific effects of thyroid hormone on RC3/neurogranin expression in rat brain ENDOCRINOLOGY Iniguez, M. A., deLecea, L., GUADANOFERRAZ, A., Morte, B., Gerendasy, D., Sutcliffe, J. G., Bernal, J. 1996; 137 (3): 1032-1041

  Abstract

  To identify thyroid hormone-sensitive neuronal populations in the forebrain, we studied the effects of thyroid hormone deficiency and replacement on the expression of RC3 messenger RNA (mRNA) in the rat brain by in situ hybridization. RC3/neurogranin is a brain-specific, calmodulin-binding, protein kinase C substrate that has been implicated in postsynaptic events involving calcium as a second messenger. We have previously shown that RC3 mRNA and protein concentrations are thyroid hormone dependent in developing and adult rats. In normal developing rats, RC3 expression occurs in two phases. Before postnatal day 10 (P10), RC3 mRNA was detected mainly in layers II/III and V of cerebral cortex and the CA fields of the hippocampus. From P10 to P15, it decreased in layer V and increased in layer VI, the retrosplenial cortex, the caudate-putamen nucleus, and the dentate gyrus. Expression in the caudate followed a lateral to medial gradient. Thyroid hormone deficiency interfered with the late phase of RC3 expression, such that developing hypothyroid rats showed lower RC3 expression in layer VI, the retrosplenial cortex, the dentate gyrus, and the caudate, and increased expression in layer V. These changes were reverted by T4 treatment. Adult- onset hyperthyroidism also reversibly decreased hybridization in the striatum. In contrast to other molecular targets of thyroid hormone in the brain, such as myelin genes, expression of RC3 was also affected by long term hypothyroidism in the absence of hormone replacement, indicating that thyroid hormone is a required factor for the cell-specific control of RC3 expression. In addition to identifying thyroid hormone-sensitive neurons, our results suggest that one action of thyroid hormone during brain development is the timely coordination of gene expression among phenotypically different, region-specific neuronal populations.

  View details for Web of Science ID A1996TW60600033

  View details for PubMedID 8603571

• Cellular and subcellular immunolocalization of the type 3 serotonin receptor in the rat central nervous system MOLECULAR BRAIN RESEARCH Morales, M., Battenberg, E., deLecea, L., Sanna, P. P., Bloom, F. E. 1996; 36 (2): 251-260

  Abstract

  We developed and characterized 14 polyclonal antibodies against peptides whose sequences were predicted from the type 3 serotonin receptor subunit A (5-HT3R-A) cDNA. One such antiserum, 0165, raised against a peptide corresponding to the large putative intracellular loop, immunoprecipitated in vitro translated 5-HT3R-A protein and recognized both recombinant and neuronal 5-HT3R-A protein by Western blot at a high titer. Furthermore, when antiserum 0165 was used to immunolabel brain sections previously hybridized with a riboprobe specific for 5-HT3R-A transcripts, neuronal co-localization of immunoproduct and transcript was widely found throughout the brain. The study of the distribution of 5-HT3R-A-immunoreactivity in the rat central nervous system with antiserum 0165 revealed intensely immunolabeled neurons in the forebrain (isocortex, olfactory regions, hippocampal formation and amygdala), brainstem (sensory and motor nuclei and nuclei of the reticular formation) and spinal cord (dorsal and ventral horn). At the subcellular level, the 5-HT3R-A was found in endomembranes involved in translation (nuclear envelope and endoplasmic reticulum) and in the dendritic plasma-membrane. The present report is the first description of the 5-HT3R-A immunolocalization in the CNS. The wide distribution of the 5-HT3R-A in the brain and spinal cord based on ligand binding, in situ hybridization and immunolocalization studies support its participation in a large array of central nervous system functions.

  View details for Web of Science ID A1996UD31500006

  View details for PubMedID 8965645

• DEVELOPMENTAL EXPRESSION OF PARVALBUMIN MESSENGER-RNA IN THE CEREBRAL-CORTEX AND HIPPOCAMPUS OF THE RAT MOLECULAR BRAIN RESEARCH deLecea, L., delRio, J. A., Soriano, E. 1995; 32 (1): 1-13

  Abstract

  Parvalbumin (PARV) belongs to the family of calcium-binding proteins bearing the EF hand domain. Immunocytochemical studies in the cerebral cortex have demonstrated that neurons containing PARV include two types of GABAergic interneurons, namely, basket and axo-axonic chandelier cells. The present study examines the onset and pattern of PARV mRNA expression during the development of rat neocortex and hippocampus by means of 'in situ' hybridization with an oligonucleotide probe corresponding to rat PARV cDNA. In animals aged P0-P6 no signal was detected above background in neocortex or hippocampus. At P8, a few cortical cells displayed a number of silver grains just above background levels. By P10 PARV mRNA-expressing cells in the neocortex were detected almost exclusively in layer V of somatosensory, frontal and cingulate cortices. At P12 PARV mRNA was mainly detected in layers IV, V and VIa. By P14 there was a marked overall increase in the entire neocortex, including layer II-III, both in the number of cells and in their intensity of labelling. Further maturation in the pattern of PARV mRNA concentration was observed between P16 and P21. In the hippocampus low hybridization was observed at P10-P12. In subsequent stages both the number of positive cells and the intensity of labelling increased steadily. No clear-cut radial gradients for the expression of PARV mRNA were observed in the hippocampal region. Our results show that the developmental radial gradient followed by PARV mRNA expression in the neocortex does not follow an 'inside-out' gradient, consistent with previous immunocytochemical findings. Taken together, these data indicate that the developmental sequence followed by the PARV protein directly reflects mRNA abundance and suggest that PARV mRNA expression correlates with the functional maturation of cortical interneurons.

  View details for Web of Science ID A1995RL35500001

  View details for PubMedID 7494447

• THE GENE ENCODING RAT PHOSPHOGLYCERATE MUTASE SUBUNIT-M - CLONING AND PROMOTER ANALYSIS IN SKELETAL-MUSCLE CELLS GENE RUIZLOZANO, P., deLecea, L., Buesa, C., DELAOSA, P. P., LePage, D., Gualberto, A., Walsh, K., Pons, G. 1994; 147 (2): 243-248

  Abstract

  The expression of the gene encoding the muscle-specific (M)-subunit of phosphoglycerate mutase (PGAM-M) is restricted to adult skeletal and cardiac muscle. In order to study its expression in muscle, the rat PGAM-M gene has been isolated and sequenced. Rat PGAM-M spans about 2.2 kb and is composed of three exons: 442, 181 and 186-bp long, and two introns of 97 bp and 1.3 bp. The analysis of the 5'-flanking region reveals a promoter which contains multiple DNA regulatory elements and constitutes an ideal model to study muscle gene transcriptional regulation. Thus, the elements responsible for rat PGAM-M muscle-specific expression have been identified by transient transfection in chicken embryo primary cultures, using chimeric constructs of the rat promoter linked to a cat reporter gene. Here, we report that in spite of the abundance of E-box motifs in the rat PGAM-M promoter known for their involvement in muscle gene expression, two DNA elements regulate the muscle-specific transcription of rat PGAM-M: an A/T motif, the putative MEF-2-binding site (myocyte-specific enhancer-binding factor 2), and a proximal 27-bp element which is conserved between the rat and human genes. These two elements define a small promoter (170 bp) sufficient to support potent and skeletal-muscle-specific expression. The conserved 27-bp region contains a transcriptional regulatory element able to confer muscle-specific expression when located upstream from a heterologous TATA box.

  View details for Web of Science ID A1994PK88000015

  View details for PubMedID 7926808

• THE DEVELOPMENT OF PARVALBUMIN-IMMUNOREACTIVITY IN THE NEOCORTEX OF THE MOUSE DEVELOPMENTAL BRAIN RESEARCH delRio, J. A., deLecea, L., Ferrer, I., Soriano, E. 1994; 81 (2): 247-259

  Abstract

  In the present study the postnatal development of parvalbumin-immunoreactivity was examined in the neocortex of the mouse. Postnatal mice were processed at different developmental stages using a well-characterized monoclonal antibody against parvalbumin, and immunocytochemistry. The first immunoreactive neurons appeared in the first parietal and retrosplenial cortices at postnatal day 10 (P10). From P11 to P12, immunoreactivity emerged in the second parietal, cingular, frontal, hindlimb-forelimb, first temporal, primary and secondary occipital and gustatory cortices, and at P14, parvalbumin-positive cells were present in the remaining regions. In general, parvalbumin-immunoreactivity appeared first in the primary sensory/motor areas, and then in second sensory/motor or associative areas. The maturation of parvalbumin-immunoreactivity, however, was a long-lasting process, which was not completed until adult stages. In all cortical regions, parvalbumin-immunoreactive cells were present first in layer V, from which immunoreactivity expanded to the upper and inner cortical layers at subsequent developmental stages. This pattern of maturation differed from the usual 'inside-out' gradient of neocortical neurogenesis and maturation. At the cellular level, parvalbumin-immunoreactivity appeared first in cell somata, and staining of dendrites and boutons was apparent two days later. From the second postnatal week onwards, an immunoreactive axonal system was observed in the neocortical white matter and the corpus callosum. We conclude that the emergence and maturation of parvalbumin-immunoreactivity in the mouse neocortex shows marked area-specific differences, but proceeds following a similar center-to-outside radial gradient. These features may reflect the acquisition of certain physiological properties by a subset of GABAergic inhibitory neurons.

  View details for Web of Science ID A1994PG25000011

  View details for PubMedID 7813046

• G-PROTEIN GAMMA-7 SUBUNIT IS SELECTIVELY EXPRESSED IN MEDIUM-SIZED NEURONS AND DENDRITES OF THE RAT NEOSTRIATUM JOURNAL OF NEUROSCIENCE RESEARCH Watson, J. B., COULTER, P. M., Margulies, J. E., deLecea, L., Danielson, P. E., Erlander, M. G., Sutcliffe, J. G. 1994; 39 (1): 108-116

  Abstract

  We used subtractive hybridization to isolate clones of gamma 7, a 68 residue G-protein gamma subunit. Northern blotting and in situ hybridization reveal that the gamma 7 subunit mRNA is expressed primarily in medium-sized neurons of the neostriatum and nucleus accumbens and neurons of the olfactory tubercle, and at low levels in the dentate gyrus of the hippocampal formation and laminae II-III, and V of the neocortex. The gamma 7 mRNA is translocated into dendrites of neurons in the neostriatum and the dentate gyrus of the hippocampus. gamma 7 is expressed at relatively very low concentrations in peripheral tissues. The selective pattern of gamma 7 expression within the brain is highly reminiscent of those of the striatum-enriched adenylyl cyclase ACST, dopamine receptors, and the alpha subunit of G(olf), suggesting that, in striatum, gamma 7 may be a subunit of a G(olf) alpha-containing G protein that couples dopamine receptors selectively to ACST.

  View details for Web of Science ID A1994PF02700012

  View details for PubMedID 7807587

• TRANSCRIPTS ENCODING A NEURAL MEMBRANE CD26 PEPTIDASE-LIKE PROTEIN ARE STIMULATED BY SYNAPTIC ACTIVITY MOLECULAR BRAIN RESEARCH deLecea, L., Soriano, E., Criado, J. R., Steffensen, S. C., Henriksen, S. J., Sutcliffe, J. G. 1994; 25 (3-4): 286-296

  Abstract

  We isolated a cDNA clone, named BSPL, that encodes a brain-specific dipeptidyl peptidase-like protein with 30% identity and 50% similarity to CD26, a lymphocyte membrane antigen involved in T-cell activation. BSPL lacks, however, the catalytic residue responsible for peptidase activity. The expression of BSPL is widespread throughout the CNS but restricted to neurons under normal conditions. Twenty-four hours after injection of kainic acid into the hippocampus, a dramatic increase in the concentration of BSPL mRNA was detected by in situ hybridization in the CA3 region of the injected hemisphere as compared with the contralateral hemisphere or sham-injected animals. An increase in the steady-state level of BSPL mRNA concentration was also found following tetanic stimulation of the perforant path to produce LTP in granule cells of the dentate gyrus. Hybridization signals could be detected in dendritic processes of pyramidal neurons and in some glial cells upon either type of stimulation. These data suggest that BSPL may be involved in synaptic plasticity.

  View details for Web of Science ID A1994PD78700013

  View details for PubMedID 7808228

• ISOLATION OF CLONES OF RAT STRIATUM-SPECIFIC MESSENGER-RNAS BY DIRECTIONAL TAG PCR SUBTRACTION JOURNAL OF NEUROSCIENCE Usui, H., Falk, J. D., Dopazo, A., deLecea, L., Erlander, M. G., Sutcliffe, J. G. 1994; 14 (8): 4915-4926

  Abstract

  We report an improved subtractive cDNA cloning procedure, named "directional tag PCR subtraction," for isolating clones of mRNAs enriched in a target tissue compared to a second tissue, the driver. In this method, the target and driver are prepared from directional cDNA libraries constructed in different vectors, and the target cDNA contains tag sequences at both its 5' and 3' ends for PCR amplification. This method avoids several limitations of previous subtraction procedures, and was demonstrated to be technically easy and efficient. Using the directional tag PCR subtraction and improved screening procedures, cDNA clones corresponding to mRNAs expressed in the striatum but not in the cerebellum of the rat brain were efficiently isolated, including mRNAs encoding calmodulin-dependent phosphodiesterase, a transcriptional regulatory protein, and several previously uncharacterized species. Our data suggest that approximately 1% of the striatal polyA+ RNA mass potentially encoding more than 300 distinct proteins corresponds to RNA species reduced in concentration or absent from the cerebellum, of which about one-third are expressed prominently only in the striatum. This unexpected finding suggests that the striatum has a unique biochemical character within the brain, and that characterization of these mRNAs will be important for understanding the biochemical basis of striatal function.

  View details for Web of Science ID A1994PA95300030

  View details for PubMedID 8046460

• Four structurally distinct neuron-specific olfactomedin-related glycoproteins produced by differential promoter utilization and alternative mRNA splicing from a single gene. Journal of neuroscience research Danielson, P. E., Forss-Petter, S., Battenberg, E. L., deLecea, L., Bloom, F. E., Sutcliffe, J. G. 1994; 38 (4): 468-478

  Abstract

  Four structurally related neuron-specific 1B426b mRNAs, designated AMY, BMY, AMZ, and BMZ, have been isolated from rat brain cDNA libraries. The four mRNAs are related to one another by their shared M region and by two pairs of alternative 5' (A, B) or 3' (Y, Z) regions. All four possible combinations were detected. The four transcripts are derived by differential promoter utilization (to generate A or B 5' ends) and alternative splicing (to generate Y or Z 3' ends) of the primary transcripts of the single D2Sutle gene. All four mRNAs were detected in most brain regions, but were enriched within the cortex and hippocampus. In the pituitary only the two A-type and in the adrenal glands only the two B-type mRNAs were detected. In situ hybridization shows a highly heterogeneous distribution across brain regions, paralleling the Northern blot results and additionally identifying the reactive cells as neurons. The cDNAs encode related glycoproteins of 125, 153, 457, and 485 amino acids, which have been detected immunochemically. The AMZ and BMZ proteins show significant sequence similarity with olfactomedin, an extracellular matrix protein of bullfrog olfactory epithelium, suggesting the possibility of a matrix-related function for these rat glycoproteins in neurons and neurosecretory cells.

  View details for PubMedID 7932877

• CHROMOSOMAL MAPPING OF MOUSE GENES EXPRESSED SELECTIVELY WITHIN THE CENTRAL-NERVOUS-SYSTEM GENOMICS Danielson, P. E., Watson, J. B., Gerendasy, D. D., Erlander, M. G., Lovenberg, T. W., deLecea, L., Sutcliffe, J. G., Frankel, W. N. 1994; 19 (3): 454-461

  Abstract

  We have used RFLP analysis on DNA from a panel of interspecific (C57BL/6J x Mus spretus) F1 x M. spretus backcross offspring to assign the genes encoding 10 neuron-specific mRNAs and 2 loci corresponding to cyclophilin 2-related sequences to the mouse chromosomal map. The Pss1 locus encoding the forebrain-enriched protein kinase C substrate RC3, a component of dendritic spines, mapped to proximal Chr 9. The Camkl locus encoding the calmodulin-binding protein kinase-like vesicle protein 1G5 mapped to distal Chr 9. The Gng7 locus encoding the gamma 7 G-protein subunit, highly enriched in the striatum and presumptively coupled to dopamine receptors, mapped to mid-Chr 10. The Htr1f, Htr5a, Htr5b, and Htr7loci, encoding four serotonin receptors, mapped to Chr 16, 5, 1, and 19, respectively. The Peplb locus, encoding a CD26 ectopeptidase-like neuronal membrane protein activated by kainate and long-term potentiation, mapped to Chr 5. The D2Sut1e and Cpu3 loci, encoding neural proteins of unknown functions, mapped to Chrs 2 and 9, respectively. Two cyclophilin 2-related loci, Cphn2-r1 and Cphn2-r2, mapped to different regions of Chr 9. Comparison of these 12 newly mapped loci with the existing mouse map and known regions of syntenic homology with the human map, along with the known features and expression profiles of the products of these genes, suggests a few candidates for mouse mutations and human neurological and immunological deficits, including the Tourette syndrome and Bloom syndrome genes.

  View details for Web of Science ID A1994NA60500007

  View details for PubMedID 7910582

• A NOVEL ADENYLYL CYCLASE-ACTIVATING SEROTONIN RECEPTOR (5-HT7) IMPLICATED IN THE REGULATION OF MAMMALIAN CIRCADIAN-RHYTHMS NEURON Lovenberg, T. W., Baron, B. M., deLecea, L., Miller, J. D., Prosser, R. A., Rea, M. A., Foye, P. E., Racke, M., SLONE, A. L., SIEGEL, B. W., Danielson, P. E., Sutcliffe, J. G., Erlander, M. G. 1993; 11 (3): 449-458

  Abstract

  We report the cloning and characterization of a novel serotonin receptor, designated as 5-HT7, which is coupled to the stimulation of adenylyl cyclase. 5-HT7 mRNA is expressed discretely throughout the CNS, predominantly in the thalamus and hypothalamus. 5-HT7 has a unique pharmacological profile that redefines agonist and antagonist classification of ligands previously thought to be "selective." The circadian phase of spontaneous neuronal activity of the rat suprachiasmatic nucleus of the hypothalamus advances in response to serotonin ligands with a pharmacological profile consistent exclusively with that of 5-HT7. These findings suggest a physiological role in the regulation of circadian rhythms for one subtype of serotonin receptor, 5-HT7, and provide a pharmacological test to evaluate its role in other neuronal systems.

  View details for Web of Science ID A1993LY49800005

  View details for PubMedID 8398139

• Two members of a distinct subfamily of 5-hydroxytryptamine receptors differentially expressed in rat brain. Proceedings of the National Academy of Sciences of the United States of America Erlander, M. G., Lovenberg, T. W., Baron, B. M., De Lecea, L., Danielson, P. E., Racke, M., SLONE, A. L., SIEGEL, B. W., Foye, P. E., Cannon, K. 1993; 90 (8): 3452-3456

  Abstract

  We report two serotonin (5-hydroxytryptamine, 5-HT) receptors, MR22 and REC17, that belong to the G-protein-associated receptor superfamily. MR22 and REC17 are 371 and 357 amino acids long, respectively, as deduced from nucleotide sequence and share 68% mutual amino acid identity and 30-35% identity with known catecholamine and 5-HT receptors. Saturable binding of 125I-labeled (+)-lysergic acid diethylamide to transiently expressed MR22 in COS-M6 cells was inhibited by ergotamine > methiothepin > 5-carboxamidotryptamine > 5-HT. For REC17, the rank of potency was ergotamine > 5-carboxamidotryptamine > methiothepin > methysergide > 5-HT. Both were insensitive to 5-HT1A, 5-HT1D or 5-HT2 serotonergic ligands [8-hydroxy-2-(di-n-propylamino)tetralin, sumatriptan, and 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane]. The mRNAs encoding MR22 were detected in the CA1 region of hippocampus, the medial habenula, and raphe nuclei. In contrast, mRNAs encoding REC17 were found throughout the rat central nervous system. We propose that REC17 and MR22, designated as 5-HT5 alpha and 5-HT5 beta, represent a distinct subfamily of 5-HT receptors.

  View details for PubMedID 7682702

• 2 MEMBERS OF A DISTINCT SUBFAMILY OF 5-HYDROXYTRYPTAMINE RECEPTORS DIFFERENTIALLY EXPRESSED IN RAT-BRAIN PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Erlander, M. G., Lovenberg, T. W., Baron, B. M., deLecea, L., Danielson, P. E., Racke, M., SLONE, A. L., SIEGEL, B. W., Foye, P. E., Cannon, K., Burns, J. E., Sutcliffe, J. G. 1993; 90 (8): 3452-3456

  View details for Web of Science ID A1993KX81600070

• MOLECULAR APPROACH TO HYPOTHALAMIC RHYTHMS - ISOLATION OF NOVEL INDOLEAMINE RECEPTOR GENES JOURNAL OF BIOLOGICAL RHYTHMS Erlander, M. G., Lovenberg, T. W., Baron, B. M., deLecea, L., Sutcliffe, J. G. 1993; 8: S25-S31

  Abstract

  We have utilized polymerase chain reaction with primers corresponding to conserved amino acid sequences within membrane-spanning regions of known serotonin receptors to identify clones of four putative new indoleamine receptors. We have determined complete amino acid sequences of these four receptors, which fall into three subfamilies; two of these subfamilies are novel. The sites of expression within the brain have been determined for each of the genes. Expression in mammalian cells demonstrates that each new protein is a receptor for serotonin and that each has a distinct pharmacology when compared to known receptors. Two of the new receptors are coupled to cyclic adenosine monophosphate, one negatively (Gi) and one positively (Gs). The latter is a candidate for the serotonin receptor that mediates phase advances in circadian rhythms of the suprachiasmatic nucleus.

  View details for Web of Science ID A1993MB81800005

  View details for PubMedID 8274759

• LATE APPEARANCE OF PARVALBUMIN-IMMUNOREACTIVE NEURONS IN THE RODENT CEREBRAL-CORTEX DOES NOT FOLLOW AN INSIDE-OUT SEQUENCE NEUROSCIENCE LETTERS Soriano, E., delRio, J. A., Ferrer, I., Auladell, C., deLecea, L., Alcantara, S. 1992; 142 (2): 147-150

  Abstract

  Parvalbumin (PARV), a Ca(2+)-binding protein believed to play a role in neuronal excitability, is contained in certain GABAergic inhibitory neurons of the cerebral cortex. Here we report that expression of PARV in the developing neocortex of rats and mice occurs with a sequence which does not follow the usual 'inside-out' gradient of cortical development. Thus, PARV-immunoreactive neurons appear first in layer V and only thereafter in the remaining cortical layers. An adult-like pattern of immunoreactivity is reached simultaneously in layers II-III and VIb. These observations indicate that the mechanisms regulating the functional maturation of PARV-containing inhibitory neurons are different from those that generally govern developmental processes in the cortex.

  View details for Web of Science ID A1992JK73700010

  View details for PubMedID 1454208

• CLONING AND SEQUENCING OF A CDNA-ENCODING 2,3-BISPHOSPHOGLYCERATE-INDEPENDENT PHOSPHOGLYCERATE MUTASE FROM MAIZE - POSSIBLE RELATIONSHIP TO THE ALKALINE-PHOSPHATASE FAMILY JOURNAL OF BIOLOGICAL CHEMISTRY Grana, X., deLecea, L., ELMAGHRABI, M. R., Urena, J. M., CAELLAS, C., Carreras, J., Puigdomenech, P., Pilkis, S. J., Climent, F. 1992; 267 (18): 12797-12803

  Abstract

  The primary sequence of maize 2,3-bisphosphoglycerate-independent phosphoglycerate mutase was deduced from cDNAs isolated from maize cDNA libraries by screening with specific antibodies to the cofactor-independent enzyme and from a maize genomic clone. The genomic clone provided the 5'-nucleotide sequence encoding the N-terminal amino acids which could not be obtained from the cDNA. Confirmation that the nucleotide sequence was for the cofactor-independent phosphoglycerate mutase was obtained by sequencing the peptides generated from cyanogen bromide cleavage of the purified protein. This is the first report of the amino acid sequence of a 2,3-bisphosphoglycerate cofactor-independent phosphoglycerate mutase, which consists of 559 amino acids and is twice the molecular size of the mammalian cofactor-dependent enzyme subunit. Analysis of the cofactor-independent phosphoglycerate mutase amino acid sequence revealed no identity with the cofactor-dependent mutase types. Northern blot analysis confirmed this difference since the maize cofactor-independent phosphoglycerate mutase cDNA did not hybridize with mRNA of the cofactor-dependent mutase. The lack of amino acid identity between cofactor-dependent and -independent enzymes is consistent with their different catalytic mechanisms and suggests that both enzymes are unrelated evolutionarily and arose from two independent ancestral genes. However, a constellation of residues which are involved in metal ion binding in various alkaline phosphatases is conserved in the maize cofactor-independent phosphoglycerate mutase, which suggests that the enzyme is a member of the alkaline phosphatase family of enzymes.

  View details for Web of Science ID A1992HZ48300063

  View details for PubMedID 1535626

• ISOLATION AND SEQUENCING OF A CDNA-ENCODING THE B-ISOZYME OF RAT PHOSPHOGLYCERATE MUTASE GENE Urena, J. M., Grana, X., deLecea, L., Ruiz, P., Castella, J., Carreras, J., Pons, G., Climent, F. 1992; 113 (2): 281-282

  Abstract

  Phosphoglycerate mutase consists of two kinds of different subunits, M and B. We previously sequenced a rat cDNA encoding the type-M subunit. Here, we report the sequence of the type-B subunit-encoding cDNA. This cDNA has 1754 bp and contains a long 3'-untranslated region of 897 bp.

  View details for Web of Science ID A1992HT14500020

  View details for PubMedID 1533381