Paul Buckmaster, DVM, PhD

All Publications

• Ventral hippocampal formation is the primary epileptogenic zone in a rat model of temporal lobe epilepsy. The Journal of neuroscience : the official journal of the Society for Neuroscience Buckmaster, P. S., Reyes, B., Kahn, T., Wyeth, M. 2022

  Abstract

  Temporal lobe epilepsy is common, but mechanisms of seizure initiation are unclear. We evaluated seizure initiation in female and male rats that had been systemically treated with pilocarpine, a widely used model of temporal lobe epilepsy. Local field potential recordings from many brain regions revealed variable sites of earliest recorded seizure activity, but mostly the ventral hippocampal formation. To test whether inactivation of the ventral hippocampal formation would reduce seizures, mini-osmotic pumps were used to continually and focally deliver tetrodotoxin. High doses of tetrodotoxin infused unilaterally into the ventral hippocampal formation blocked seizures reversibly but also reduced local field potential amplitudes in remote brain regions, indicating distant effects. A lower dose did not reduce local field potential amplitudes in remote brain regions but did not reduce seizures when infused unilaterally. Instead, seizures tended to initiate in the contralateral ventral hippocampal formation. Bilateral infusion of the lower dose into the ventral hippocampal formation reduced seizure frequency 85%. Similar bilateral treatment in the amygdala was not effective. Bilateral infusion of the dorsal hippocampus reduced seizure frequency, but only 17%. Together, these findings reveal that the ventral hippocampal formation is a primary bilaterally independent epileptogenic zone, and the dorsal hippocampus is a secondary epileptogenic zone in pilocarpine-treated rats. This is consistent with many human patients, and the results further validate the local field potential method for identifying seizure onset zones. Finally, the findings are more consistent with a focal mechanism of ictogenesis rather than one involving a network of interdependent nodes.SIGNIFICANCE STATEMENT:To better understand how seizures start, investigators need to know where seizures start in the animal models they study. In the widely used pilocarpine-treated rat model of temporal lobe epilepsy, earliest seizure activity was most frequently recorded in the ventral hippocampal formation. Confirming the primary role of the ventral hippocampal formation, seizure frequency was reduced most effectively when it was inactivated focally, bilaterally, and continually with infused tetrodotoxin. These findings suggest the ventral hippocampal formation is the primary site of seizure initiation in this animal model of temporal lobe epilepsy, consistent with findings in many human patients.

  View details for DOI 10.1523/JNEUROSCI.0429-22.2022

  View details for PubMedID 35995562

• Lack of hyper-inhibition of oriens-lacunosum moleculare cells by vasoactive intestinal peptide-expressing cells in a model of temporal lobe epilepsy. eNeuro Wyeth, M., Buckmaster, P. S. 2021

  Abstract

  Temporal lobe epilepsy remains a common disorder with no cure and inadequate treatments, potentially due to an incomplete understanding of how seizures start. CA1 pyramidal cells and many inhibitory interneurons increase their firing rate in the seconds-minutes before a spontaneous seizure in epileptic rats. However, some interneurons fail to do so, including those identified as putative interneurons with somata in oriens and axons targeting lacunosum-moleculare (OLM cells). Somatostatin-containing cells, including OLM cells, are the primary target of inhibitory vasoactive intestinal polypeptide and calretinin-expressing (VIP/CR) bipolar interneuron-selective interneurons, type 3 (ISI-3). The objective of this study was to test the hypothesis that in epilepsy inhibition of OLM cells by ISI-3 is abnormally increased, potentially explaining the failure of OLM recruitment when needed most during the ramp up of activity preceding a seizure. Stereological quantification of VIP/CR cells in a model of temporal lobe epilepsy demonstrated that they survive in epileptic mice, despite a reduction in their somatostatin-expressing cell targets. Paired recordings of unitary inhibitory postsynaptic currents from ISI-3 to OLM cells did not show increased connection probability or increased connection strength, and failure rate was unchanged. When miniature postsynaptic currents in ISI-3 were compared, only mIPSC frequency was increased in epileptic hippocampi. Nevertheless, spontaneous and miniature postsynaptic potentials were unchanged in OLM cells of epileptic mice. These results are not consistent with the hypothesis of hyper-inhibition from VIP/CR bipolar cells impeding recruitment of OLM cells in advance of a seizure.SignificanceInadequate recruitment of inhibitory cells in general, and OLM cells in particular, may be a mechanism of seizure initiation, making it important to determine why OLM cells do not fire faster and provide preictal feedback inhibition when presynaptic CA1 pyramidal activity is ramping up. This study excludes aberrantly increased inhibition of OLM cells by VIP bipolar cells as the cause, pointing to other possibilities for investigation.

  View details for DOI 10.1523/ENEURO.0299-21.2021

  View details for PubMedID 34819310

• Ictal onset sites and γ-aminobutyric acidergic neuron loss in epileptic pilocarpine-treated rats. Epilepsia Wyeth, M. n., Nagendran, M. n., Buckmaster, P. S. 2020

  Abstract

  The present study tested whether ictal onset sites are regions of more severe interneuron loss in epileptic pilocarpine-treated rats, a model of human temporal lobe epilepsy.Local field potential recordings were evaluated to identify ictal onset sites. Electrode sites were visualized in Nissl-stained sections. Adjacent sections were processed with proximity ligation in situ hybridization for glutamic acid decarboxylase 2 (Gad2). Gad2 neuron profile numbers at ictal onset sites were compared to contralateral regions. Other sections were processed with immunocytochemistry for reelin or nitric oxide synthase (NOS), which labeled major subtypes of granule cell layer-associated interneurons. Stereology was used to estimate numbers of reelin and NOS granule cell layer-associated interneurons per hippocampus.Ictal onset sites varied between and within rats but were mostly in the ventral hippocampus and were frequently bilateral. There was no conclusive evidence of more severe Gad2 neuron profile loss at sites of earliest seizure activity compared to contralateral regions. Numbers of granule cell layer-associated NOS neurons were reduced in the ventral hippocampus.In epileptic pilocarpine-treated rats, ictal onset sites were mostly in the ventral hippocampus, where there was loss of granule cell layer-associated NOS interneurons. These findings suggest the hypothesis that loss of granule cell layer-associated NOS interneurons in the ventral hippocampus is a mechanism of temporal lobe epilepsy.

  View details for DOI 10.1111/epi.16490

  View details for PubMedID 32242932

• Proportional loss of parvalbumin-immunoreactive synaptic boutons and granule cells from the hippocampus of sea lions with temporal lobe epilepsy JOURNAL OF COMPARATIVE NEUROLOGY Cameron, S., Lopez, A., Glabman, R., Abrams, E., Johnson, S., Field, C., Gulland, F. D., Buckmast, P. S. 2019; 527 (14): 2341–55

  View details for DOI 10.1002/cne.24680

  View details for Web of Science ID 000476915800010

• Seizure frequency correlates with loss of dentate gyrus GABAergic neurons in a mouse model of temporal lobe epilepsy. journal of comparative neurology Buckmaster, P. S., Abrams, E., Wen, X. 2017; 525 (11): 2592-2610

  Abstract

  Epilepsy occurs in one of 26 people. Temporal lobe epilepsy is common and can be difficult to treat effectively. It can develop after brain injuries that damage the hippocampus. Multiple pathophysiological mechanisms involving the hippocampal dentate gyrus have been proposed. This study evaluated a mouse model of temporal lobe epilepsy to test which pathological changes in the dentate gyrus correlate with seizure frequency and help prioritize potential mechanisms for further study. FVB mice (n = 127) that had experienced status epilepticus after systemic treatment with pilocarpine 31-61 days earlier were video-monitored for spontaneous, convulsive seizures 9 hr/day every day for 24-36 days. Over 4,060 seizures were observed. Seizure frequency ranged from an average of one every 3.6 days to one every 2.1 hr. Hippocampal sections were processed for Nissl stain, Prox1-immunocytochemistry, GluR2-immunocytochemistry, Timm stain, glial fibrillary acidic protein-immunocytochemistry, glutamic acid decarboxylase in situ hybridization, and parvalbumin-immunocytochemistry. Stereological methods were used to measure hilar ectopic granule cells, mossy cells, mossy fiber sprouting, astrogliosis, and GABAergic interneurons. Seizure frequency was not significantly correlated with the generation of hilar ectopic granule cells, the number of mossy cells, the extent of mossy fiber sprouting, the extent of astrogliosis, or the number of GABAergic interneurons in the molecular layer or hilus. Seizure frequency significantly correlated with the loss of GABAergic interneurons in or adjacent to the granule cell layer, but not with the loss of parvalbumin-positive interneurons. These findings prioritize the loss of granule cell layer interneurons for further testing as a potential cause of temporal lobe epilepsy.

  View details for DOI 10.1002/cne.24226

  View details for PubMedID 28425097

• More Docked Vesicles and Larger Active Zones at Basket Cell-to-Granule Cell Synapses in a Rat Model of Temporal Lobe Epilepsy. journal of neuroscience Buckmaster, P. S., Yamawaki, R., Thind, K. 2016; 36 (11): 3295-3308

  Abstract

  Temporal lobe epilepsy is a common and challenging clinical problem, and its pathophysiological mechanisms remain unclear. One possibility is insufficient inhibition in the hippocampal formation where seizures tend to initiate. Normally, hippocampal basket cells provide strong and reliable synaptic inhibition at principal cell somata. In a rat model of temporal lobe epilepsy, basket cell-to-granule cell (BC→GC) synaptic transmission is more likely to fail, but the underlying cause is unknown. At some synapses, probability of release correlates with bouton size, active zone area, and number of docked vesicles. The present study tested the hypothesis that impaired GABAergic transmission at BC→GC synapses is attributable to ultrastructural changes. Boutons making axosomatic symmetric synapses in the granule cell layer were reconstructed from serial electron micrographs. BC→GC boutons were predicted to be smaller in volume, have fewer and smaller active zones, and contain fewer vesicles, including fewer docked vesicles. Results revealed the opposite. Compared with controls, epileptic pilocarpine-treated rats displayed boutons with over twice the average volume, active zone area, total vesicles, and docked vesicles and with more vesicles closer to active zones. Larger active zones in epileptic rats are consistent with previous reports of larger amplitude miniature IPSCs and larger BC→GC quantal size. Results of this study indicate that transmission failures at BC→GC synapses in epileptic pilocarpine-treated rats are not attributable to smaller boutons or fewer docked vesicles. Instead, processes following vesicle docking, including priming, Ca(2+) entry, or Ca(2+) coupling with exocytosis, might be responsible.One in 26 people develops epilepsy, and temporal lobe epilepsy is a common form. Up to one-third of patients are resistant to currently available treatments. This study tested a potential underlying mechanism for previously reported impaired inhibition in epileptic animals at basket cell-to-granule cell (BC→GC) synapses, which normally are reliable and strong. Electron microscopy was used to evaluate 3D ultrastructure of BC→GC synapses in a rat model of temporal lobe epilepsy. The hypothesis was that impaired synaptic transmission is attributable to smaller boutons, smaller synapses, and abnormally low numbers of synaptic vesicles. Results revealed the opposite. These findings suggest that impaired transmission at BC→GC synapses in epileptic rats is attributable to later steps in exocytosis following vesicle docking.

  View details for DOI 10.1523/JNEUROSCI.4049-15.2016

  View details for PubMedID 26985038

  View details for PubMedCentralID PMC4792940

• Unit Activity of Hippocampal Interneurons before Spontaneous Seizures in an Animal Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Toyoda, I., Fujita, S., Thamattoor, A. K., Buckmaster, P. S. 2015; 35 (16): 6600-6618

  Abstract

  Mechanisms of seizure initiation are unclear. To evaluate the possible roles of inhibitory neurons, unit recordings were obtained in the dentate gyrus, CA3, CA1, and subiculum of epileptic pilocarpine-treated rats as they experienced spontaneous seizures. Most interneurons in the dentate gyrus, CA1, and subiculum increased their firing rate before seizures, and did so with significant consistency from seizure to seizure. Identification of CA1 interneuron subtypes based on firing characteristics during theta and sharp waves suggested that a parvalbumin-positive basket cell and putative bistratified cells, but not oriens lacunosum moleculare cells, were activated preictally. Preictal changes occurred much earlier than those described by most previous in vitro studies. Preictal activation of interneurons began earliest (>4 min before seizure onset), increased most, was most prevalent in the subiculum, and was minimal in CA3. Preictal inactivation of interneurons was most common in CA1 (27% of interneurons) and included a putative ivy cell and parvalbumin-positive basket cell. Increased or decreased preictal activity correlated with whether interneurons fired faster or slower, respectively, during theta activity. Theta waves were more likely to occur before seizure onset, and increased preictal firing of subicular interneurons correlated with theta activity. Preictal changes by other hippocampal interneurons were largely independent of theta waves. Within seconds of seizure onset, many interneurons displayed a brief pause in firing and a later, longer drop that was associated with reduced action potential amplitude. These findings suggest that many interneurons inactivate during seizures, most increase their activity preictally, but some fail to do so at the critical time before seizure onset.

  View details for DOI 10.1523/JNEUROSCI.4786-14.2015

  View details for Web of Science ID 000353399800031

  View details for PubMedID 25904809

  View details for PubMedCentralID PMC4405565

• Rat strain differences in seizure frequency and hilar neuron loss after systemic treatment with pilocarpine. Epilepsy research Junghans, K., Wyeth, M., Buckmaster, P. S. 2024; 204: 107384

  Abstract

  At least 3 months after systemic treatment with pilocarpine to induce status epilepticus, Long-Evans and Sprague-Dawley rats were video-EEG monitored for seizures continuously for 1 month. Rats were then perfused, hippocampi were processed for Nissl staining, and hilar neurons were quantified. Seizure frequency in Long-Evans rats was 1/10th of that in Sprague-Dawley rats, and more variable. Hilar neuron loss was also less severe in Long-Evans rats. However, there was no correlation between hilar neuron loss and seizure frequency in either strain. The low and variable seizure frequency suggests limited usefulness of pilocarpine-treated Long-Evans rats for some epilepsy experiments.

  View details for DOI 10.1016/j.eplepsyres.2024.107384

  View details for PubMedID 38879905

• Pinniped electroencephalography: Methodology and findings in California sea lions (Zalophus californianus). Frontiers in veterinary science Williams, D. C., Haulena, M., Dennison, S., Waugh, L., Goldstein, T., Nutter, F., Bonn, B. V., Hoard, V., Laxer, K. D., Buckmaster, P. S., Gulland, F. M., Tharp, B. 2023; 10: 1040125

  Abstract

  This study was designed to identify abnormalities in the electroencephalograms (EEGs) recorded from stranded California sea lions (Zalophus californianus) with suspected domoic acid (DA) toxicosis. Recordings from animals presenting for non-neurological issues were also obtained to better understand the normal EEG (background activity and transient events) in this species, as, to date, studies have focused on examining natural sleep in pinnipeds. Most animals were sedated for electrode placement and EEG acquisition with some receiving antiepileptic medications or isoflurane during the procedure. A total of 103 recordings were read and scored from 0 (normal) to 3 (severely abnormal). Epileptiform discharges, consisting of spikes, sharp waves, slow waves, and/or spike waves, were present in all EEGs with scores of 1, 2, or 3. The distribution of these events over the scalp varied. While often generalized, others were lateralized over one hemisphere, bifrontal, bioccipital, and/or bitemporal, while some discharges were multifocal. Findings were different between sea lions and occasionally changed within the EEG on a given sea lion. No clinical seizures were observed during the recording but a few sea lions had findings consistent with electroencephalographic seizures. When available, supporting diagnostic results obtained from magnetic resonance imaging (MRI) and/or necropsy/histopathology were described, as well as the status of those sea lions that recovered and were released with satellite tags.

  View details for DOI 10.3389/fvets.2023.1040125

  View details for PubMedID 37065231

  View details for PubMedCentralID PMC10102506

• Cannabinoid receptor 1-labeled boutons in the sclerotic dentate gyrus of epileptic sea lions. Epilepsy research Seelman, A., Vu, K., Buckmaster, P., Mackie, K., Field, C., Johnson, S., Wyeth, M. 2022; 184: 106965

  Abstract

  Pathology in the dentate gyrus, including sclerosis, is a hallmark of temporal lobe epilepsy, and reduced inhibition to dentate granule cells may contribute to epileptogenesis. The perisomatic-targeting axonal boutons of parvalbumin-expressing interneurons decrease in proportion with granule cells in temporal lobe epilepsy. In contrast, dendrite-targeting axonal boutons of somatostatin-expressing interneurons sprout exuberantly in temporal lobe epilepsy. A third major class of GABAergic interneurons expresses cannabinoid receptor type 1 (CB1) on their terminal boutons, but there is conflicting evidence as to whether these boutons are increased or decreased in temporal lobe epilepsy. Naturally occurring temporal lobe epilepsy in California sea lions, with unilateral or bilateral sclerosis, offers the benefit of neuroanatomy and neuropathology akin to humans, but with the advantage that the entirety of both hippocampi from control and epileptic brains can be studied. Stereological quantification in the dentate gyrus revealed that sclerotic hippocampi from epileptic sea lions had fewer CB1-labeled boutons than controls. However, the reduction in the number of granule cells was greater, resulting in increased CB1-labeled boutons per granule cell in sclerotic hippocampi at temporal levels. This suggests that although CB1-expressing boutons are decreased in sclerotic dentate gyri, surviving cells have enhanced innervation from these boutons in epileptic sea lions.

  View details for DOI 10.1016/j.eplepsyres.2022.106965

  View details for PubMedID 35724601

• Non-invasive, neurotoxic surgery reduces seizures in a rat model of temporal lobe epilepsy. Experimental neurology Zhang, Y., Buckmaster, P. S., Qiu, L., Wang, J., Keunen, O., Ghobadi, S. N., Huang, A., Hou, Q., Li, N., Narang, S., Habte, F. G., Bertram, E. H., Lee, K. S., Wintermark, M. 2021: 113761

  Abstract

  Surgery can be highly effective for treating certain cases of drug resistant epilepsy. The current study tested a novel, non-invasive, surgical strategy for treating seizures in a rat model of temporal lobe epilepsy. The surgical approach uses magnetic resonance-guided, low-intensity focused ultrasound (MRgFUS) in combination with intravenous microbubbles to open the blood-brain barrier (BBB) in a transient and focal manner. During the period of BBB opening, a systemically administered neurotoxin (Quinolinic Acid: QA) that is normally impermeable to the BBB gains access to a targeted area in the brain, destroying neurons where the BBB has been opened. This strategy is termed Precise Intracerebral Non-invasive Guided Surgery (PING). Spontaneous recurrent seizures induced by pilocarpine were monitored behaviorally prior to and after PING or under control conditions. Seizure frequency in untreated animals or animals treated with MRgFUS without QA exhibited expected seizure rate fluctuations frequencies between the monitoring periods. In contrast, animals treated with PING targeting the intermediate-temporal aspect of the hippocampus exhibited substantial reductions in seizure frequency, with convulsive seizures being eliminated entirely in two animals. These findings suggest that PING could provide a useful alternative to invasive surgical interventions for treating drug resistant epilepsy, and perhaps for treating other neurological disorders in which aberrant neural circuitries play a role.

  View details for DOI 10.1016/j.expneurol.2021.113761

  View details for PubMedID 33991523

• Testing Different Combinations of Acoustic Pressure and Doses of Quinolinic Acid for Induction of Focal Neuron Loss in Mice Using Transcranial Low-Intensity Focused Ultrasound. Ultrasound in medicine & biology Zhang, Y., Liao, C., Qu, H., Huang, S., Jiang, H., Zhou, H., Abrams, E., Habte, F. G., Yuan, L., Bertram, E. H., Lee, K. S., Pauly, K. B., Buckmaster, P. S., Wintermark, M. 2018

  Abstract

  The goal of this study was to test different combinations of acoustic pressure and doses of quinolinic acid (QA) for producing a focal neuronal lesion in the murine hippocampus without causing unwanted damage to adjacent brain structures. Sixty male CD-1 mice were divided into 12 groups that underwent magnetic resonance-guided focused ultrasound at high (0.67 MPa), medium (0.5 MPa) and low (0.33 MPa) acoustic peak negative pressures and received QA at high (0.012 mmol), medium (0.006 mmol) and low (0.003 mmol) dosages. Neuronal loss occurred only when magnetic resonance-guided focused ultrasound with adequate acoustic power (0.67 or 0.5 MPa) was combined with QA. The animals subjected to the highest acoustic power had larger lesions than those treated with medium acoustic power, but two mice had evidence of bleeding. When the intermediate acoustic power was used, medium andhigh dosages of QA produced lesions larger than those produced by the low dosage.

  View details for PubMedID 30309748

• A single subconvulsant dose of domoic acid at mid-gestation does not cause temporal lobe epilepsy in mice NEUROTOXICOLOGY Demars, F., Clark, K., Wyeth, M. S., Abrams, E., Buckmaster, P. S. 2018; 66: 128–37

  Abstract

  Harmful blooms of domoic acid (DA)-producing algae are a problem in oceans worldwide. DA is a potent glutamate receptor agonist that can cause status epilepticus and in survivors, temporal lobe epilepsy. In mice, one-time low-dose in utero exposure to DA was reported to cause hippocampal damage and epileptiform activity, leading to the hypothesis that unrecognized exposure to DA from contaminated seafood in pregnant women can damage the fetal hippocampus and initiate temporal lobe epileptogenesis. However, development of epilepsy (i.e., spontaneous recurrent seizures) has not been tested. In the present study, long-term seizure monitoring and histology was used to test for temporal lobe epilepsy following prenatal exposure to DA. In Experiment One, the previous study's in utero DA treatment protocol was replicated, including use of the CD-1 mouse strain. Afterward, mice were video-monitored for convulsive seizures from 2 to 6 months old. None of the CD-1 mice treated in utero with vehicle or DA was observed to experience spontaneous convulsive seizures. After seizure monitoring, mice were evaluated for pathological evidence of temporal lobe epilepsy. None of the mice treated in utero with DA displayed the hilar neuron loss that occurs in patients with temporal lobe epilepsy and in the mouse pilocarpine model of temporal lobe epilepsy. In Experiment Two, a higher dose of DA was administered to pregnant FVB mice. FVB mice were tested as a potentially more sensitive strain, because they have a lower seizure threshold, and some females spontaneously develop epilepsy. Female offspring were monitored with continuous video and telemetric bilateral hippocampal local field potential recording at 1-11 months old. A similar proportion of vehicle- and DA-treated female FVB mice spontaneously developed epilepsy, beginning in the fourth month of life. Average seizure frequency and duration were similar in both groups. Seizure frequency was lower than that of positive-control pilocarpine-treated mice, but seizure duration was similar. None of the mice treated in utero with vehicle or DA displayed hilar neuron loss or intense mossy fiber sprouting, a form of aberrant synaptic reorganization that develops in patients with temporal lobe epilepsy and in pilocarpine-treated mice. FVB mice that developed epilepsy (vehicle- and DA-treated) displayed mild mossy fiber sprouting. Results of this study suggest that a single subconvulsive dose of DA at mid-gestation does not cause temporal lobe epilepsy in mice.

  View details for PubMedID 29625197

  View details for PubMedCentralID PMC5940543

• Hilar somatostatin interneuron loss reduces dentate gyrus inhibition in a mouse model of temporal lobe epilepsy EPILEPSIA Hofmann, G., Balgooyen, L., Mattis, J., Deisseroth, K., Buckmaster, P. S. 2016; 57 (6): 977-983

  Abstract

  In patients with temporal lobe epilepsy, seizures usually start in the hippocampus, and dentate granule cells are hyperexcitable. Somatostatin interneurons are a major subpopulation of inhibitory neurons in the dentate gyrus, and many are lost in patients and animal models. However, surviving somatostatin interneurons sprout axon collaterals and form new synapses, so the net effect on granule cell inhibition remains unclear.The present study uses optogenetics to activate hilar somatostatin interneurons and measure the inhibitory effect on dentate gyrus perforant path-evoked local field potential responses in a mouse model of temporal lobe epilepsy.In controls, light activation of hilar somatostatin interneurons inhibited evoked responses up to 40%. Epileptic pilocarpine-treated mice exhibited loss of hilar somatostatin interneurons and less light-induced inhibition of evoked responses.These findings suggest that severe epilepsy-related loss of hilar somatostatin interneurons can overwhelm the surviving interneurons' capacity to compensate by sprouting axon collaterals.

  View details for DOI 10.1111/epi.13376

  View details for Web of Science ID 000380151100017

  View details for PubMedID 27030321

  View details for PubMedCentralID PMC4903908

• Surviving Mossy Cells Enlarge and Receive More Excitatory Synaptic Input in a Mouse Model of Temporal Lobe Epilepsy HIPPOCAMPUS Zhang, W., Thamattoor, A. K., LeRoy, C., Buckmaster, P. S. 2015; 25 (5): 594-604

  Abstract

  Numerous hypotheses of temporal lobe epileptogenesis have been proposed, and several involve hippocampal mossy cells. Building on previous hypotheses we sought to test the possibility that after epileptogenic injuries surviving mossy cells develop into super-connected seizure-generating hub cells. If so, they might require more cellular machinery and consequently have larger somata, elongate their dendrites to receive more synaptic input, and display higher frequencies of miniature excitatory synaptic currents (mEPSCs). To test these possibilities pilocarpine-treated mice were evaluated using GluR2-immunocytochemistry, whole-cell recording, and biocytin-labeling. Epileptic pilocarpine-treated mice displayed substantial loss of GluR2-positive hilar neurons. Somata of surviving neurons were 1.4-times larger than in controls. Biocytin-labeled mossy cells also were larger in epileptic mice, but dendritic length per cell was not significantly different. The average frequency of mEPSCs of mossy cells recorded in the presence of tetrodotoxin and bicuculline was 3.2-times higher in epileptic pilocarpine-treated mice as compared to controls. Other parameters of mEPSCs were similar in both groups. Average input resistance of mossy cells in epileptic mice was reduced to 63% of controls, which is consistent with larger somata and would tend to make surviving mossy cells less excitable. Other intrinsic physiological characteristics examined were similar in both groups. Increased excitatory synaptic input is consistent with the hypothesis that surviving mossy cells develop into aberrantly super-connected seizure-generating hub cells, and soma hypertrophy is indirectly consistent with the possibility of axon sprouting. However, no obvious evidence of hyperexcitable intrinsic physiology was found. Furthermore, similar hypertrophy and hyper-connectivity has been reported for other neuron types in the dentate gyrus, suggesting mossy cells are not unique in this regard. Thus, findings of the present study reveal epilepsy-related changes in mossy cell anatomy and synaptic input but do not strongly support the hypothesis that mossy cells develop into seizure-generating hub cells. © 2014 Wiley Periodicals, Inc.

  View details for DOI 10.1002/hipo.22396

  View details for Web of Science ID 000353975200005

  View details for PubMedID 25488607

• Blockade of Excitatory Synaptogenesis With Proximal Dendrites of Dentate Granule Cells Following Rapamycin Treatment in a Mouse Model of Temporal Lobe Epilepsy JOURNAL OF COMPARATIVE NEUROLOGY Yamawaki, R., Thind, K., Buckmaster, P. S. 2015; 523 (2): 281-297

  Abstract

  Inhibiting the mammalian target of rapamycin (mTOR) signaling pathway with rapamycin blocks granule cell axon (mossy fiber) sprouting after epileptogenic injuries, including pilocarpine-induced status epilepticus. However, it remains unclear whether axons from other types of neurons sprout into the inner molecular layer and synapse with granule cell dendrites despite rapamycin treatment. If so, other aberrant positive-feedback networks might develop. To test this possibility stereological electron microscopy was used to estimate the numbers of excitatory synapses in the inner molecular layer per hippocampus in pilocarpine-treated control mice, in mice 5 days after pilocarpine-induced status epilepticus, and after status epilepticus and daily treatment beginning 24 hours later with rapamycin or vehicle for 2 months. The optical fractionator method was used to estimate numbers of granule cells in Nissl-stained sections so that numbers of excitatory synapses in the inner molecular layer per granule cell could be calculated. Control mice had an average of 2,280 asymmetric synapses in the inner molecular layer per granule cell, which was reduced to 63% of controls 5 days after status epilepticus, recovered to 93% of controls in vehicle-treated mice 2 months after status epilepticus, but remained at only 63% of controls in rapamycin-treated mice. These findings reveal that rapamycin prevented excitatory axons from synapsing with proximal dendrites of granule cells and raise questions about the recurrent excitation hypothesis of temporal lobe epilepsy.

  View details for DOI 10.1002/cne.23681

  View details for Web of Science ID 000345772600007

  View details for PubMedID 25234294

  View details for PubMedCentralID PMC4252507

• Preictal Activity of Subicular, CA1, and Dentate Gyrus Principal Neurons in the Dorsal Hippocampus before Spontaneous Seizures in a Rat Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Fujita, S., Toyoda, I., Thamattoor, A. K., Buckmaster, P. S. 2014; 34 (50): 16671-16687

  Abstract

  Previous studies suggest that spontaneous seizures in patients with temporal lobe epilepsy might be preceded by increased action potential firing of hippocampal neurons. Preictal activity is potentially important because it might provide new opportunities for predicting when a seizure is about to occur and insight into how spontaneous seizures are generated. We evaluated local field potentials and unit activity of single, putative excitatory neurons in the subiculum, CA1, CA3, and dentate gyrus of the dorsal hippocampus in epileptic pilocarpine-treated rats as they experienced spontaneous seizures. Average action potential firing rates of neurons in the subiculum, CA1, and dentate gyrus, but not CA3, increased significantly and progressively beginning 2-4 min before locally recorded spontaneous seizures. In the subiculum, CA1, and dentate gyrus, but not CA3, 41-57% of neurons displayed increased preictal activity with significant consistency across multiple seizures. Much of the increased preictal firing of neurons in the subiculum and CA1 correlated with preictal theta activity, whereas preictal firing of neurons in the dentate gyrus was independent of theta. In addition, some CA1 and dentate gyrus neurons displayed reduced firing rates preictally. These results reveal that different hippocampal subregions exhibit differences in the extent and potential underlying mechanisms of preictal activity. The finding of robust and significantly consistent preictal activity of subicular, CA1, and dentate neurons in the dorsal hippocampus, despite the likelihood that many seizures initiated in other brain regions, suggests the existence of a broader neuronal network whose activity changes minutes before spontaneous seizures initiate.

  View details for DOI 10.1523/JNEUROSCI.0584-14.2014

  View details for Web of Science ID 000346191500014

  View details for PubMedID 25505320

  View details for PubMedCentralID PMC4261094

• Hippocampal Neuropathology of Domoic Acid-Induced Epilepsy in California Sea Lions (Zalophus californianus) JOURNAL OF COMPARATIVE NEUROLOGY Buckmaster, P. S., Wen, X., Toyoda, I., Gulland, F. M., Van Bonn, W. 2014; 522 (7): 1691-1706

  Abstract

  California sea lions (Zalophus californianus) are abundant human-sized carnivores with large gyrencephalic brains. They develop epilepsy after experiencing status epilepticus when naturally exposed to domoic acid. We tested whether sea lions previously exposed to DA (chronic DA sea lions) display hippocampal neuropathology similar to that of human patients with temporal lobe epilepsy. Hippocampi were obtained from control and chronic DA sea lions. Stereology was used to estimate numbers of Nissl-stained neurons per hippocampus in the granule cell layer, hilus, and pyramidal cell layer of CA3, CA2, and CA1 subfields. Adjacent sections were processed for somatostatin immunoreactivity or Timm-stained, and the extent of mossy fiber sprouting was measured stereologically. Chronic DA sea lions displayed hippocampal neuron loss in patterns and extents similar but not identical to those reported previously for human patients with temporal lobe epilepsy. Similar to human patients, hippocampal sclerosis in sea lions was unilateral in 79% of cases, mossy fiber sprouting was a common neuropathological abnormality, and somatostatin-immunoreactive axons were exuberant in the dentate gyrus despite loss of immunopositive hilar neurons. Thus, hippocampal neuropathology of chronic DA sea lions is similar to that of human patients with temporal lobe epilepsy.

  View details for DOI 10.1002/cne.23509

  View details for Web of Science ID 000332916300011

  View details for PubMedID 24638960

• Does Mossy Fiber Sprouting Give Rise to the Epileptic State? Workshop on Issues in Clinical Epileptology - A View from the Bench held in honor of Phil Buckmaster, P. S. SPRINGER. 2014: 161–168

  Abstract

  Many patients with temporal lobe epilepsy display structural changes in the seizure initiating zone, which includes the hippocampus. Structural changes in the hippocampus include granule cell axon (mossy fiber) sprouting. The role of mossy fiber sprouting in epileptogenesis is controversial. A popular view of temporal lobe epileptogenesis contends that precipitating brain insults trigger transient cascades of molecular and cellular events that permanently enhance excitability of neuronal networks through mechanisms including mossy fiber sprouting. However, recent evidence suggests there is no critical period for mossy fiber sprouting after an epileptogenic brain injury. Instead, findings from stereological electron microscopy and rapamycin-delayed mossy fiber sprouting in rodent models of temporal lobe epilepsy suggest a persistent, homeostatic mechanism exists to maintain a set level of excitatory synaptic input to granule cells. If so, a target level of mossy fiber sprouting might be determined shortly after a brain injury and then remain constant. Despite the static appearance of synaptic reorganization after its development, work by other investigators suggests there might be continual turnover of sprouted mossy fibers in epileptic patients and animal models. If so, there may be opportunities to reverse established mossy fiber sprouting. However, reversal of mossy fiber sprouting is unlikely to be antiepileptogenic, because blocking its development does not reduce seizure frequency in pilocarpine-treated mice. The challenge remains to identify which, if any, of the many other structural changes in the hippocampus are epileptogenic.

  View details for Web of Science ID 000346021700015

  View details for PubMedID 25012375

• High-dose rapamycin blocks mossy fiber sprouting but not seizures in a mouse model of temporal lobe epilepsy. Epilepsia Heng, K., Haney, M. M., Buckmaster, P. S. 2013; 54 (9): 1535-1541

  Abstract

  The role of granule cell axon (mossy fiber) sprouting in temporal lobe epileptogenesis is unclear and controversial. Rapamycin suppresses mossy fiber sprouting, but its reported effects on seizure frequency are mixed. The present study used high-dose rapamycin to more completely block mossy fiber sprouting and to measure the effect on seizure frequency.Mice were treated with pilocarpine to induce status epilepticus. Beginning 24 h later and continuing for 2 months, vehicle or rapamycin (10 mg/kg/day) was administered. Starting 1 month after status epilepticus, mice were monitored by video 9 h per day, every day, for 1 month to measure the frequency of spontaneous motor seizures. At the end of seizure monitoring, a subset of mice was prepared for anatomic analysis. Mossy fiber sprouting was measured as the proportion of the granule cell layer and molecular layer that displayed black labeling in Timm-stained sections.Extensive mossy fiber sprouting developed in mice that experienced status epilepticus and were treated with vehicle. In rapamycin-treated mice, mossy fiber sprouting was blocked almost to the level of naive controls. Seizure frequency was similar in vehicle-treated and rapamycin-treated mice.These findings suggest that mossy fiber sprouting is not necessary for epileptogenesis in the mouse pilocarpine model. They also reveal that rapamycin does not have antiseizure or antiepileptogenic effects in this model.

  View details for DOI 10.1111/epi.12246

  View details for PubMedID 23848506

• Early Activation of Ventral Hippocampus and Subiculum during Spontaneous Seizures in a Rat Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Toyoda, I., Bower, M. R., Leyva, F., Buckmaster, P. S. 2013; 33 (27): 11100-11115

  Abstract

  Temporal lobe epilepsy is the most common form of epilepsy in adults. The pilocarpine-treated rat model is used frequently to investigate temporal lobe epilepsy. The validity of the pilocarpine model has been challenged based largely on concerns that seizures might initiate in different brain regions in rats than in patients. The present study used 32 recording electrodes per rat to evaluate spontaneous seizures in various brain regions including the septum, dorsomedial thalamus, amygdala, olfactory cortex, dorsal and ventral hippocampus, substantia nigra, entorhinal cortex, and ventral subiculum. Compared with published results from patients, seizures in rats tended to be shorter, spread faster and more extensively, generate behavioral manifestations more quickly, and produce generalized convulsions more frequently. Similarities to patients included electrographic waveform patterns at seizure onset, variability in sites of earliest seizure activity within individuals, and variability in patterns of seizure spread. Like patients, the earliest seizure activity in rats was recorded most frequently within the hippocampal formation. The ventral hippocampus and ventral subiculum displayed the earliest seizure activity. Amygdala, olfactory cortex, and septum occasionally displayed early seizure latencies, but not above chance levels. Substantia nigra and dorsomedial thalamus demonstrated consistently late seizure onsets, suggesting their unlikely involvement in seizure initiation. The results of the present study reveal similarities in onset sites of spontaneous seizures in patients with temporal lobe epilepsy and pilocarpine-treated rats that support the model's validity.

  View details for DOI 10.1523/JNEUROSCI.0472-13.2013

  View details for Web of Science ID 000321258000017

  View details for PubMedID 23825415

  View details for PubMedCentralID PMC3718374

• Short-term treatment with the GABAA receptor antagonist pentylenetetrazole produces a sustained pro-cognitive benefit in a mouse model of Down's syndrome. British journal of pharmacology Colas, D., Chuluun, B., Warrier, D., Blank, M., Wetmore, D. Z., Buckmaster, P., Garner, C. C., Heller, H. C. 2013; 169 (5): 963-973

  Abstract

  BACKGROUND AND PURPOSE: Down's syndrome (DS) is a common genetic cause of intellectual disability yet there are no drug therapies. Mechanistic studies in a model of DS (Ts65Dn mice) demonstrated that impaired cognitive function is due to excessive neuronal inhibitory tone. These deficits can be normalized by low doses of GABAA receptor (GABAA R) antagonists in adult animals. In this study, we explore the therapeutic potential of pentylenetetrazole (PTZ), a GABAA R antagonist which has a history of safe use in man. EXPERIMENTAL APPROACH: Long-term memory was assessed by the Novel Object Recognition (NOR) test in different cohorts of Ts65Dn mice after a delay following a short-term chronic treatment with PTZ. Seizure susceptibility, as an index of treatment safety, was studied by means of EEG, behavior and hippocampus morphology. EEG spectral analysis was used as a biometric of the treatment. RESULTS: PTZ has a broad therapeutic window (0.03-3mg.kg(-1) ) that is >10-1000 fold below the seizure threshold for this drug, and chronic PTZ treatment does not lower the seizure threshold. Remarkably, short-term, low, chronic dose regimens of PTZ elicit long-lasting (>1week) normalization of cognitive function in young and aged mice. PTZ effectiveness is time of day dependent: cognitive performance improves when PTZ is delivered during the light (inactive) phase, but not during the dark (active) phase. Chronic PTZ treatment results in EEG power normalization. CONCLUSIONS: PTZ at very low dosage can be administered safely, produces long lasting cognitive improvements and has the potential of fulfilling an unmet therapeutic need in DS.

  View details for DOI 10.1111/bph.12169

  View details for PubMedID 23489250

• Factors affecting outcomes of pilocarpine treatment in a mouse model of temporal lobe epilepsy EPILEPSY RESEARCH Buckmaster, P. S., Haney, M. M. 2012; 102 (3): 153-159

  Abstract

  Pilocarpine-treated mice are an increasingly used model of temporal lobe epilepsy. However, outcomes of treatment can be disappointing, because many mice die or fail to develop status epilepticus. To improve animal welfare and outcomes of future experiments we analyzed results of previous pilocarpine treatments to identify factors that correlate with development of status epilepticus and survival. All treatments were performed by one investigator with mice of the FVB background strain. Results from 2413 mice were evaluated for effects of sex, age, body weight, and latency between administration of atropine methyl bromide and pilocarpine. All parameters correlated with effects on outcomes. Best results were obtained from male mice, 6-7 weeks old, and 21-25 g, with pilocarpine administered 18-30 min after atropine methyl bromide. In that group only 23% failed to develop status epilepticus, and 64% developed status epilepticus and survived. Those results are substantially better than that of the total sample in which 31% failed to develop status epilepticus and only 34% developed status epilepticus and survived.

  View details for DOI 10.1016/j.eplepsyres.2012.05.012

  View details for Web of Science ID 000312519400002

  View details for PubMedID 22721955

  View details for PubMedCentralID PMC3458129

• Mossy cell dendritic structure quantified and compared with other hippocampal neurons labeled in rats in vivo EPILEPSIA Buckmaster, P. S. 2012; 53: 9-17

  Abstract

  Mossy cells are likely to contribute to normal hippocampal function and to the pathogenesis of neurologic disorders that involve the hippocampus, including epilepsy. Mossy cells are the least well-characterized excitatory neurons in the hippocampus. Their somatic and dendritic morphology has been described qualitatively but not quantitatively. In the present study rat mossy cells were labeled intracellularly with biocytin in vivo. Somatic and dendritic structure was reconstructed three-dimensionally. For comparison, granule cells, CA3 pyramidal cells, and CA1 pyramidal cells were labeled and analyzed using the same approach. Among the four types of hippocampal neurons, granule cells had the smallest somata, fewest primary dendrites and dendritic branches, and shortest total dendritic length. CA1 pyramidal cells had the most dendritic branches and longest total dendritic length. Mossy cells and CA3 pyramidal cells both had large somata and similar total dendritic lengths. However, mossy cell dendrites branched less than CA3 pyramidal cells, especially close to the soma. These findings suggest that mossy cells have dendritic features that are not identical to any other type of hippocampal neuron. Therefore, electrotonic properties that depend on soma-dendritic structure are likely to be distinct in mossy cells compared to other neurons.

  View details for DOI 10.1111/j.1528-1167.2012.03470.x

  View details for Web of Science ID 000304257700003

  View details for PubMedID 22612804

  View details for PubMedCentralID PMC3536534

• Distinct Neuronal Coding Schemes in Memory Revealed by Selective Erasure of Fast Synchronous Synaptic Transmission NEURON Xu, W., Morishita, W., Buckmaster, P. S., Pang, Z. P., Malenka, R. C., Suedhof, T. C. 2012; 73 (5): 990-1001

  Abstract

  Neurons encode information by firing spikes in isolation or bursts and propagate information by spike-triggered neurotransmitter release that initiates synaptic transmission. Isolated spikes trigger neurotransmitter release unreliably but with high temporal precision. In contrast, bursts of spikes trigger neurotransmission reliably (i.e., boost transmission fidelity), but the resulting synaptic responses are temporally imprecise. However, the relative physiological importance of different spike-firing modes remains unclear. Here, we show that knockdown of synaptotagmin-1, the major Ca(2+) sensor for neurotransmitter release, abrogated neurotransmission evoked by isolated spikes but only delayed, without abolishing, neurotransmission evoked by bursts of spikes. Nevertheless, knockdown of synaptotagmin-1 in the hippocampal CA1 region did not impede acquisition of recent contextual fear memories, although it did impair the precision of such memories. In contrast, knockdown of synaptotagmin-1 in the prefrontal cortex impaired all remote fear memories. These results indicate that different brain circuits and types of memory employ distinct spike-coding schemes to encode and transmit information.

  View details for DOI 10.1016/j.neuron.2011.12.036

  View details for PubMedID 22405208

• Identification of new epilepsy treatments: Issues in preclinical methodology EPILEPSIA Galanopoulou, A. S., Buckmaster, P. S., Staley, K. J., Moshe, S. L., Perucca, E., Engel, J., Loescher, W., Noebels, J. L., Pitkanen, A., Stables, J., White, H. S., O'Brien, T. J., Simonato, M. 2012; 53 (3): 571-582

  Abstract

  Preclinical research has facilitated the discovery of valuable drugs for the symptomatic treatment of epilepsy. Yet, despite these therapies, seizures are not adequately controlled in a third of all affected individuals, and comorbidities still impose a major burden on quality of life. The introduction of multiple new therapies into clinical use over the past two decades has done little to change this. There is an urgent demand to address the unmet clinical needs for: (1) new symptomatic antiseizure treatments for drug-resistant seizures with improved efficacy/tolerability profiles, (2) disease-modifying treatments that prevent or ameliorate the process of epileptogenesis, and (3) treatments for the common comorbidities that contribute to disability in people with epilepsy. New therapies also need to address the special needs of certain subpopulations, that is, age- or gender-specific treatments. Preclinical development in these treatment areas is complex due to heterogeneity in presentation and etiology, and may need to be formulated with a specific seizure, epilepsy syndrome, or comorbidity in mind. The aim of this report is to provide a framework that will help define future guidelines that improve and standardize the design, reporting, and validation of data across preclinical antiepilepsy therapy development studies targeting drug-resistant seizures, epileptogenesis, and comorbidities.

  View details for DOI 10.1111/j.1528-1167.2011.03391.x

  View details for Web of Science ID 000300838400023

  View details for PubMedID 22292566

• Increased Excitatory Synaptic Input to Granule Cells from Hilar and CA3 Regions in a Rat Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Zhang, W., Huguenard, J. R., Buckmaster, P. S. 2012; 32 (4): 1183-1196

  Abstract

  One potential mechanism of temporal lobe epilepsy is recurrent excitation of dentate granule cells through aberrant sprouting of their axons (mossy fibers), which is found in many patients and animal models. However, correlations between the extent of mossy fiber sprouting and seizure frequency are weak. Additional potential sources of granule cell recurrent excitation that would not have been detected by markers of mossy fiber sprouting in previous studies include surviving mossy cells and proximal CA3 pyramidal cells. To test those possibilities in hippocampal slices from epileptic pilocarpine-treated rats, laser-scanning glutamate uncaging was used to randomly and focally activate neurons in the granule cell layer, hilus, and proximal CA3 pyramidal cell layer while measuring evoked EPSCs in normotopic granule cells. Consistent with mossy fiber sprouting, a higher proportion of glutamate-uncaging spots in the granule cell layer evoked EPSCs in epileptic rats compared with controls. In addition, stimulation spots in the hilus and proximal CA3 pyramidal cell layer were more likely to evoke EPSCs in epileptic rats, despite significant neuron loss in those regions. Furthermore, synaptic strength of recurrent excitatory inputs to granule cells from CA3 pyramidal cells and other granule cells was increased in epileptic rats. These findings reveal substantial levels of excessive, recurrent, excitatory synaptic input to granule cells from neurons in the hilus and proximal CA3 field. The aberrant development of these additional positive-feedback circuits might contribute to epileptogenesis in temporal lobe epilepsy.

  View details for DOI 10.1523/JNEUROSCI.5342-11.2012

  View details for Web of Science ID 000299801100005

  View details for PubMedID 22279204

• Is there a critical period for mossy fiber sprouting in a mouse model of temporal lobe epilepsy? EPILEPSIA Lew, F. H., Buckmaster, P. S. 2011; 52 (12): 2326-2332

  Abstract

  Dentate granule cell axon (mossy fiber) sprouting creates an aberrant positive-feedback circuit that might be epileptogenic. Presumably, mossy fiber sprouting is initiated by molecular signals, but it is unclear whether they are expressed transiently or persistently. If transient, there might be a critical period when short preventative treatments could permanently block mossy fiber sprouting. Alternatively, if signals persist, continuous treatment would be necessary. The present study tested whether temporary treatment with rapamycin has long-term effects on mossy fiber sprouting.Mice were treated daily with 1.5 mg/kg rapamycin or vehicle (i.p.) beginning 24 h after pilocarpine-induced status epilepticus. Mice were perfused for anatomic evaluation immediately after 2 months of treatment ("0 delay") or after an additional 6 months without treatment ("6-month delay"). One series of sections was Timm-stained, and an adjacent series was Nissl-stained. Stereologic methods were used to measure the volume of the granule cell layer plus molecular layer and the Timm-positive fraction. Numbers of Nissl-stained hilar neurons were estimated using the optical fractionator method.At 0 delay, rapamycin-treated mice had significantly less black Timm staining in the granule cell layer plus molecular layer than vehicle-treated animals. However, by 6-month delay, Timm staining had increased significantly in mice that had been treated with rapamycin. Percentages of the granule cell layer plus molecular layer that were Timm-positive were high and similar in 0 delay vehicle-treated, 6-month delay vehicle-treated, and 6-month delay rapamycin-treated mice. Extent of hilar neuron loss was similar among all groups that experienced status epilepticus and, therefore, was not a confounding factor. Compared to naive controls, average volume of the granule cell layer plus molecular layer was larger in 0 delay vehicle-treated mice. The hypertrophy was partially suppressed in 0 delay rapamycin-treated mice. However, 6-month delay vehicle- and 6-month delay rapamycin-treated animals had similar average volumes of the granule cell layer plus molecular layer that were significantly larger than those of all other groups.Status epilepticus-induced mossy fiber sprouting and dentate gyrus hypertrophy were suppressed by systemic treatment with rapamycin but resumed after treatment ceased. These findings suggest that molecular signals that drive mossy fiber sprouting and dentate gyrus hypertrophy might persist for >2 months after status epilepticus in mice. Therefore, prolonged or continuous treatment might be required to permanently suppress mossy fiber sprouting.

  View details for DOI 10.1111/j.1528-1167.2011.03315.x

  View details for Web of Science ID 000297695100025

  View details for PubMedID 22092282

  View details for PubMedCentralID PMC3233231

• Rapamycin suppresses axon sprouting by somatostatin interneurons in a mouse model of temporal lobe epilepsy EPILEPSIA Buckmaster, P. S., Wen, X. 2011; 52 (11): 2057-2064

  Abstract

  In temporal lobe epilepsy many somatostatin interneurons in the dentate gyrus die. However, some survive and sprout axon collaterals that form new synapses with granule cells. The functional consequences of γ-aminobutyric acid (GABA)ergic synaptic reorganization are unclear. Development of new methods to suppress epilepsy-related interneuron axon sprouting might be useful experimentally.Status epilepticus was induced by systemic pilocarpine treatment in green fluorescent protein (GFP)-expressing inhibitory nerurons (GIN) mice in which a subset of somatostatin interneurons expresses GFP. Beginning 24 h later, mice were treated with vehicle or rapamycin (3 mg/kg intraperitoneally) every day for 2 months. Stereologic methods were then used to estimate numbers of GFP-positive hilar neurons per dentate gyrus and total length of GFP-positive axon in the molecular layer plus granule cell layer. GFP-positive axon density was calculated. The number of GFP-positive axon crossings of the granule cell layer was measured. Regression analyses were performed to test for correlations between GFP-positive axon length versus number of granule cells and dentate gyrus volume.After pilocarpine-induced status epilepticus, rapamycin- and vehicle-treated mice had approximately half as many GFP-positive hilar neurons as did control animals. Despite neuron loss, vehicle-treated mice had over twice the GFP-positive axon length per dentate gyrus as controls, consistent with GABAergic axon sprouting. In contrast, total GFP-positive axon length was similar in rapamycin-treated mice and controls. GFP-positive axon length correlated most closely with dentate gyrus volume.These findings suggest that rapamycin suppressed axon sprouting by surviving somatostatin/GFP-positive interneurons after pilocarpine-induced status epilepticus in GIN mice. It is unclear whether the effect of rapamycin on axon length was on interneurons directly or secondary, for example, by suppressing growth of granule cell dendrites, which are synaptic targets of interneuron axons. The mammalian target of rapamycin (mTOR) signaling pathway might be a useful drug target for influencing GABAergic synaptic reorganization after epileptogenic treatments, but additional side effects of rapamycin treatment must be considered carefully.

  View details for DOI 10.1111/j.1528-1167.2011.03253.x

  View details for Web of Science ID 000297049700022

  View details for PubMedID 21883182

  View details for PubMedCentralID PMC3204172

• Rapamycin Suppresses Mossy Fiber Sprouting But Not Seizure Frequency in a Mouse Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Buckmaster, P. S., Lew, F. H. 2011; 31 (6): 2337-2347

  Abstract

  Temporal lobe epilepsy is prevalent and can be difficult to treat effectively. Granule cell axon (mossy fiber) sprouting is a common neuropathological finding in patients with mesial temporal lobe epilepsy, but its role in epileptogenesis is unclear and controversial. Focally infused or systemic rapamycin inhibits the mammalian target of rapamycin (mTOR) signaling pathway and suppresses mossy fiber sprouting in rats. We tested whether long-term systemic treatment with rapamycin, beginning 1 d after pilocarpine-induced status epilepticus in mice, would suppress mossy fiber sprouting and affect the development of spontaneous seizures. Mice that had experienced status epilepticus and were treated for 2 months with rapamycin displayed significantly less mossy fiber sprouting (42% of vehicle-treated animals), and the effect was dose dependent. However, behavioral and video/EEG monitoring revealed that rapamycin- and vehicle-treated mice displayed spontaneous seizures at similar frequencies. These findings suggest mossy fiber sprouting is neither pro- nor anti-convulsant; however, there are caveats. Rapamycin treatment also reduced epilepsy-related hypertrophy of the dentate gyrus but did not significantly affect granule cell proliferation, hilar neuron loss, or generation of ectopic granule cells. These findings are consistent with the hypotheses that hilar neuron loss and ectopic granule cells might contribute to temporal lobe epileptogenesis.

  View details for DOI 10.1523/JNEUROSCI.4852-10.2011

  View details for Web of Science ID 000287389400040

  View details for PubMedID 21307269

  View details for PubMedCentralID PMC3073836

• Seizure-induced basal dendrites on granule cells EPILEPSIA Ribak, C. E., Shapiro, L. A., Yan, X., Dashtipour, K., Nadler, J. V., Obenaus, A., Spigelman, I., Buckmaster, P. S. 2010; 51: 43-43

  View details for DOI 10.1111/j.1528-1167.2010.02829.x

  View details for Web of Science ID 000285388600038

• Mossy fiber sprouting in the dentate gyrus EPILEPSIA Buckmaster, P. S. 2010; 51: 39-39

  View details for DOI 10.1111/j.1528-1167.2010.02825.x

  View details for Web of Science ID 000285388600034

• Excitatory Input Onto Hilar Somatostatin Interneurons Is Increased in a Chronic Model of Epilepsy JOURNAL OF NEUROPHYSIOLOGY Halabisky, B., Parada, I., Buckmaster, P. S., Prince, D. A. 2010; 104 (4): 2214-2223

  Abstract

  The density of somatostatin (SOM)-containing GABAergic interneurons in the hilus of the dentate gyrus is significantly decreased in both human and experimental temporal lobe epilepsy. We used the pilocarpine model of status epilepticus and temporal lobe epilepsy in mice to study anatomical and electrophysiological properties of surviving somatostatin interneurons and determine whether compensatory functional changes occur that might offset loss of other inhibitory neurons. Using standard patch-clamp techniques and pipettes containing biocytin, whole cell recordings were obtained in hippocampal slices maintained in vitro. Hilar SOM cells containing enhanced green fluorescent protein (EGFP) were identified with fluorescent and infrared differential interference contrast video microscopy in epileptic and control GIN (EGFP-expressing Inhibitory Neurons) mice. Results showed that SOM cells from epileptic mice had 1) significant increases in somatic area and dendritic length; 2) changes in membrane properties, including a small but significant decrease in resting membrane potential, and increases in time constant and whole cell capacitance; 3) increased frequency of slowly rising spontaneous excitatory postsynaptic currents (sEPSCs) due primarily to increased mEPSC frequency, without changes in the probability of release; 4) increased evoked EPSC amplitude; and 5) increased spontaneous action potential generation in cell-attached recordings. Results suggest an increase in excitatory innervation, perhaps on distal dendrites, considering the slower rising EPSCs and increased output of hilar SOM cells in this model of epilepsy. In sum, these changes would be expected to increase the inhibitory output of surviving SOM interneurons and in part compensate for interneuronal loss in the epileptogenic hippocampus.

  View details for DOI 10.1152/jn.00147.2010

  View details for Web of Science ID 000282649900037

  View details for PubMedID 20631216

• Stress coping stimulates hippocampal neurogenesis in adult monkeys PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Lyons, D. M., Buckmaster, P. S., Lee, A. G., Wu, C., Mitra, R., Duffey, L. M., Buckmaster, C. L., Her, S., Patel, P. D., Schatzberg, A. F. 2010; 107 (33): 14823-14827

  Abstract

  Coping with intermittent social stress is an essential aspect of living in complex social environments. Coping tends to counteract the deleterious effects of stress and is thought to induce neuroadaptations in corticolimbic brain systems. Here we test this hypothesis in adult squirrel monkey males exposed to intermittent social separations and new pair formations. These manipulations simulate conditions that typically occur in male social associations because of competition for limited access to residency in mixed-sex groups. As evidence of coping, we previously confirmed that cortisol levels initially increase and then are restored to prestress levels within several days of each separation and new pair formation. Follow-up studies with exogenous cortisol further established that feedback regulation of the hypothalamic-pituitary-adrenal axis is not impaired. Now we report that exposure to intermittent social separations and new pair formations increased hippocampal neurogenesis in squirrel monkey males. Hippocampal neurogenesis in rodents contributes to spatial learning performance, and in monkeys we found that spatial learning was enhanced in conditions that increased hippocampal neurogenesis. Corresponding changes were discerned in the expression of genes involved in survival and integration of adult-born granule cells into hippocampal neural circuits. These findings support recent indications that stress coping stimulates hippocampal neurogenesis in adult rodents. Psychotherapies designed to promote stress coping potentially have similar effects in humans with major depression.

  View details for DOI 10.1073/pnas.0914568107

  View details for Web of Science ID 000281287600055

  View details for PubMedID 20675584

  View details for PubMedCentralID PMC2930418

• Initial Loss but Later Excess of GABAergic Synapses with Dentate Granule Cells in a Rat Model of Temporal Lobe Epilepsy JOURNAL OF COMPARATIVE NEUROLOGY Thind, K. K., Yamawaki, R., Phanwar, I., Zhang, G., Wen, X., Buckmaster, P. S. 2010; 518 (5): 647-667

  Abstract

  Many patients with temporal lobe epilepsy display neuron loss in the dentate gyrus. One potential epileptogenic mechanism is loss of GABAergic interneurons and inhibitory synapses with granule cells. Stereological techniques were used to estimate numbers of gephyrin-positive punctae in the dentate gyrus, which were reduced short-term (5 days after pilocarpine-induced status epilepticus) but later rebounded beyond controls in epileptic rats. Stereological techniques were used to estimate numbers of synapses in electron micrographs of serial sections processed for postembedding GABA-immunoreactivity. Adjacent sections were used to estimate numbers of granule cells and glutamic acid decarboxylase-positive neurons per dentate gyrus. GABAergic neurons were reduced to 70% of control levels short-term, where they remained in epileptic rats. Integrating synapse and cell counts yielded average numbers of GABAergic synapses per granule cell, which decreased short-term and rebounded in epileptic animals beyond control levels. Axo-shaft and axo-spinous GABAergic synapse numbers in the outer molecular layer changed most. These findings suggest interneuron loss initially reduces numbers of GABAergic synapses with granule cells, but later, synaptogenesis by surviving interneurons overshoots control levels. In contrast, the average number of excitatory synapses per granule cell decreased short-term but recovered only toward control levels, although in epileptic rats excitatory synapses in the inner molecular layer were larger than in controls. These findings reveal a relative excess of GABAergic synapses and suggest that reports of reduced functional inhibitory synaptic input to granule cells in epilepsy might be attributable not to fewer but instead to abundant but dysfunctional GABAergic synapses.

  View details for DOI 10.1002/cne.22235

  View details for Web of Science ID 000273620800005

  View details for PubMedID 20034063

  View details for PubMedCentralID PMC3098130

• Surviving Hilar Somatostatin Interneurons Enlarge, Sprout Axons, and Form New Synapses with Granule Cells in a Mouse Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Zhang, W., Yamawaki, R., Wen, X., Uhl, J., Diaz, J., Prince, D. A., Buckmaster, P. S. 2009; 29 (45): 14247-14256

  Abstract

  In temporal lobe epilepsy, seizures initiate in or near the hippocampus, which frequently displays loss of neurons, including inhibitory interneurons. It is unclear whether surviving interneurons function normally, are impaired, or develop compensatory mechanisms. We evaluated GABAergic interneurons in the hilus of the dentate gyrus of epileptic pilocarpine-treated GIN mice, specifically a subpopulation of somatostatin interneurons that expresses enhanced green fluorescence protein (GFP). GFP-immunocytochemistry and stereological analyses revealed substantial loss of GFP-positive hilar neurons (GPHNs) but increased GFP-positive axon length per dentate gyrus in epileptic mice. Individual biocytin-labeled GPHNs in hippocampal slices from epileptic mice also had larger somata, more axon in the molecular layer, and longer dendrites than controls. Dual whole-cell patch recording was used to test for monosynaptic connections from hilar GPHNs to granule cells. Unitary IPSCs (uIPSCs) recorded in control and epileptic mice had similar average rise times, amplitudes, charge transfers, and decay times. However, the probability of finding monosynaptically connected pairs and evoking uIPSCs was 2.6 times higher in epileptic mice compared to controls. Together, these findings suggest that surviving hilar somatostatin interneurons enlarge, extend dendrites, sprout axon collaterals in the molecular layer, and form new synapses with granule cells. These epilepsy-related changes in cellular morphology and connectivity may be mechanisms for surviving hilar interneurons to inhibit more granule cells and compensate for the loss of vulnerable interneurons.

  View details for DOI 10.1523/JNEUROSCI.3842-09.2009

  View details for Web of Science ID 000271664000019

  View details for PubMedID 19906972

  View details for PubMedCentralID PMC2802278

• Inhibition of the Mammalian Target of Rapamycin Signaling Pathway Suppresses Dentate Granule Cell Axon Sprouting in a Rodent Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Buckmaster, P. S., Ingram, E. A., Wen, X. 2009; 29 (25): 8259-8269

  Abstract

  Dentate granule cell axon (mossy fiber) sprouting is a common abnormality in patients with temporal lobe epilepsy. Mossy fiber sprouting creates an aberrant positive-feedback network among granule cells that does not normally exist. Its role in epileptogenesis is unclear and controversial. If it were possible to block mossy fiber sprouting from developing after epileptogenic treatments, its potential role in the pathogenesis of epilepsy could be tested. Previous attempts to block mossy fiber sprouting have been unsuccessful. The present study targeted the mammalian target of rapamycin (mTOR) signaling pathway, which regulates cell growth and is blocked by rapamycin. Rapamycin was focally, continuously, and unilaterally infused into the dorsal hippocampus for prolonged periods beginning within hours after rats sustained pilocarpine-induced status epilepticus. Infusion for 1 month reduced aberrant Timm staining (a marker of mossy fibers) in the granule cell layer and molecular layer. Infusion for 2 months inhibited mossy fiber sprouting more. However, after rapamycin infusion ceased, aberrant Timm staining developed and approached untreated levels. When onset of infusion began after mossy fiber sprouting had developed for 2 months, rapamycin did not reverse aberrant Timm staining. These findings suggest that inhibition of the mTOR signaling pathway suppressed development of mossy fiber sprouting. However, suppression required continual treatment, and rapamycin treatment did not reverse already established axon reorganization.

  View details for DOI 10.1523/JNEUROSCI.4179-08.2009

  View details for Web of Science ID 000267339000032

  View details for PubMedID 19553465

  View details for PubMedCentralID PMC2819377

• Dysfunction of the Dentate Basket Cell Circuit in a Rat Model of Temporal Lobe Epilepsy JOURNAL OF NEUROSCIENCE Zhang, W., Buckmaster, P. S. 2009; 29 (24): 7846-7856

  Abstract

  Temporal lobe epilepsy is common and difficult to treat. Reduced inhibition of dentate granule cells may contribute. Basket cells are important inhibitors of granule cells. Excitatory synaptic input to basket cells and unitary IPSCs (uIPSCs) from basket cells to granule cells were evaluated in hippocampal slices from a rat model of temporal lobe epilepsy. Basket cells were identified by electrophysiological and morphological criteria. Excitatory synaptic drive to basket cells, measured by mean charge transfer and frequency of miniature EPSCs, was significantly reduced after pilocarpine-induced status epilepticus and remained low in epileptic rats, despite mossy fiber sprouting. Paired recordings revealed higher failure rates and a trend toward lower amplitude uIPSCs at basket cell-to-granule cell synapses in epileptic rats. Higher failure rates were not attributable to excessive presynaptic inhibition of GABA release by activation of muscarinic acetylcholine or GABA(B) receptors. High-frequency trains of action potentials in basket cells generated uIPSCs in granule cells to evaluate readily releasable pool (RRP) size and resupply rate of recycling vesicles. Recycling rate was similar in control and epileptic rats. However, quantal size at basket cell-to-granule cell synapses was larger and RRP size smaller in epileptic rats. Therefore, in epileptic animals, basket cells receive less excitatory synaptic drive, their pools of readily releasable vesicles are smaller, and transmission failure at basket cell-to-granule cell synapses is increased. These findings suggest dysfunction of the dentate basket cell circuit could contribute to hyperexcitability and seizures.

  View details for DOI 10.1523/JNEUROSCI.6199-08.2009

  View details for Web of Science ID 000267131000024

  View details for PubMedID 19535596

  View details for PubMedCentralID PMC2838908

• Prolonged infusion of inhibitors of calcineurin or L-type calcium channels does not block mossy fiber sprouting in a model of temporal lobe epilepsy EPILEPSIA Ingram, E. A., Toyoda, I., Wen, X., Buckmaster, P. S. 2009; 50 (1): 56-64

  Abstract

  It would be useful to selectively block granule cell axon (mossy fiber) sprouting to test its functional role in temporal lobe epileptogenesis. Targeting axonal growth cones may be an effective strategy to block mossy fiber sprouting. L-type calcium channels and calcineurin, a calcium-activated phosphatase, are critical for normal growth cone function. Previous studies have provided encouraging evidence that blocking L-type calcium channels or inhibiting calcineurin during epileptogenic treatments suppresses mossy fiber sprouting.Rats were treated systemically with pilocarpine to induce status epilepticus, which lasted at least 2 h. Then, osmotic pumps and cannulae were implanted to infuse calcineurin inhibitors (FK506 or cyclosporin A) or an L-type calcium channel blocker (nicardipine) into the dorsal dentate gyrus. After 28 days of continuous infusion, extent of mossy fiber sprouting was evaluated with Timm staining and stereological methods.Percentages of volumes of the granule cell layer plus molecular layer that were Timm-positive were similar in infused and noninfused hippocampi.These findings suggest inhibiting calcineurin or L-type calcium channels does not block mossy fiber sprouting in the pilocarpine-treated rat model of temporal lobe epilepsy.

  View details for DOI 10.1111/j.1528-1167.2008.01704.x

  View details for Web of Science ID 000262224500005

  View details for PubMedID 18616558

  View details for PubMedCentralID PMC3007596

• Synaptic input to dentate granule cell basal dendrites in a rat model of temporal lobe epilepsy JOURNAL OF COMPARATIVE NEUROLOGY Thind, K. K., Ribak, C. E., Buckmaster, P. S. 2008; 509 (2): 190-202

  Abstract

  In patients with temporal lobe epilepsy some dentate granule cells develop basal dendrites. The extent of excitatory synaptic input to basal dendrites is unclear, nor is it known whether basal dendrites receive inhibitory synapses. We used biocytin to intracellularly label individual granule cells with basal dendrites in epileptic pilocarpine-treated rats. An average basal dendrite had 3.9 branches, was 612 microm long, and accounted for 16% of a cell's total dendritic length. In vivo intracellular labeling and postembedding GABA-immunocytochemistry were used to evaluate synapses with basal dendrites reconstructed from serial electron micrographs. An average of 7% of 1,802 putative synapses were formed by GABA-positive axon terminals, indicating synaptogenesis by interneurons. Ninety-three percent of the identified synapses were GABA-negative. Most GABA-negative synapses were with spines, but at least 10% were with dendritic shafts. Multiplying basal dendrite length/cell and synapse density yielded an estimate of 180 inhibitory and 2,140 excitatory synapses per granule cell basal dendrite. Based on previous estimates of synaptic input to granule cells in control rats, these findings suggest an average basal dendrite receives approximately 14% of the total inhibitory and 19% of excitatory synapses of a cell. These findings reveal that basal dendrites are a novel source of inhibitory input, but they primarily receive excitatory synapses.

  View details for DOI 10.1002/cne.21745

  View details for Web of Science ID 000256553300006

  View details for PubMedID 18461605

  View details for PubMedCentralID PMC2667124

• Changes in granule cell firing rates precede locally recorded spontaneous seizures by minutes in an animal model of temporal lobe epilepsy JOURNAL OF NEUROPHYSIOLOGY Bower, M. R., Buckmaster, P. S. 2008; 99 (5): 2431-2442

  Abstract

  Although much is known about persistent molecular, cellular, and circuit changes associated with temporal lobe epilepsy, mechanisms of seizure onset remain unclear. The dentate gyrus displays many persistent epilepsy-related abnormalities and is in the mesial temporal lobe where seizures initiate in patients. However, little is known about seizure-related activity of individual neurons in the dentate gyrus. We used tetrodes to record action potentials of multiple, single granule cells before and during spontaneous seizures in epileptic pilocarpine-treated rats. Subsets of granule cells displayed four distinct activity patterns: increased firing before seizure onset, decreased firing before seizure onset, increased firing only after seizure onset, and unchanged firing rates despite electrographic seizure activity in the immediate vicinity. No cells decreased firing rate immediately after seizure onset. During baseline periods between seizures, action potential waveforms and firing rates were similar among the four subsets of granule cells in epileptic rats and in granule cells of control rats. The mean normalized firing rate of granule cells whose firing rates increased before seizure onset deviated from baseline earliest, beginning 4 min before dentate gyrus electrographic seizure onset, and increased progressively, more than doubling by seizure onset. It is generally assumed that neuronal firing rates increase abruptly and synchronously only when electrographic seizures begin. However, these findings show heterogeneous and gradually building changes in activity of individual granule cells minutes before spontaneous seizures.

  View details for DOI 10.1152/jn.01369.2007

  View details for Web of Science ID 000255811500033

  View details for PubMedID 18322007

• Neuron-specific nuclear antigen NeuN is not detectable in gerbil subtantia nigra pars reticulata BRAIN RESEARCH Kumar, S. S., Buckmaster, P. S. 2007; 1142: 54-60

  Abstract

  NeuN (Neuronal Nuclei), the neuron-specific marker of nuclear protein is used extensively in histological procedures to identify major cell-types in adult vertebrate nervous systems of a variety of species including rodents and humans. Some notable exceptions (i.e., NeuN-negative neurons) include Purkinje cells in cerebellum, mitral cells in olfactory bulb, and photoreceptors in retina. Here we report that neurons in gerbil (Meriones unguiculatus) substantia nigra pars reticulata (SNr), whose "neuronal" phenotype was confirmed via electrophysiology, biocytin-labeling, histology, and in situ hybridization, are also devoid of NeuN-immunoreactivity as assayed with the widely used monoclonal antibody A60. Immunohistochemistry of rat SNr using the same antibody yielded robust staining. These data suggest lack of NeuN-immonoreactivity observed in certain cell-types and brain regions can be species-specific.

  View details for DOI 10.1016/j.brainres.2007.01.027

  View details for Web of Science ID 000245785000007

  View details for PubMedID 17291468

  View details for PubMedCentralID PMC2691720

• Recurrent circuits in layer II of medial entorhinal cortex in a model of temporal lobe epilepsy JOURNAL OF NEUROSCIENCE Kumar, S. S., Jin, X., Buckmaster, P. S., Huguenard, J. R. 2007; 27 (6): 1239-1246

  Abstract

  Patients and laboratory animal models of temporal lobe epilepsy display loss of layer III pyramidal neurons in medial entorhinal cortex and hyperexcitability and hypersynchrony of less vulnerable layer II stellate cells. We sought to test the hypothesis that loss of layer III pyramidal neurons triggers synaptic reorganization and formation of recurrent, excitatory synapses among layer II stellate cells in epileptic pilocarpine-treated rats. Laser-scanning photo-uncaging of glutamate focally activated neurons in layer II while excitatory synaptic responses were recorded in stellate cells. Photostimulation revealed previously unidentified, functional, recurrent, excitatory synapses between layer II stellate cells in control animals. Contrary to the hypothesis, however, control and epileptic rats displayed similar levels of recurrent excitation. Recently, hyperexcitability of layer II stellate cells has been attributed, at least in part, to loss of GABAergic interneurons and inhibitory synaptic input. To evaluate recurrent inhibitory circuits in layer II, we focally photostimulated interneurons while recording inhibitory synaptic responses in stellate cells. IPSCs were evoked more than five times more frequently in slices from control versus epileptic animals. These findings suggest that in this model of temporal lobe epilepsy, reduced recurrent inhibition contributes to layer II stellate cell hyperexcitability and hypersynchrony, but increased recurrent excitation does not.

  View details for DOI 10.1523/JNEUROSCI.3182-06.2007

  View details for Web of Science ID 000244070000003

  View details for PubMedID 17287497

• Hyperexcitability, interneurons, and loss of GABAergic synapses in entorhinal cortex in a model of temporal lobe epilepsy JOURNAL OF NEUROSCIENCE Kumar, S. S., Buckmaster, P. S. 2006; 26 (17): 4613-4623

  Abstract

  Temporal lobe epilepsy is the most common type of epilepsy in adults, and its pathophysiology remains unclear. Layer II stellate cells of the entorhinal cortex, which are hyperexcitable in animal models of temporal lobe epilepsy, provide the predominant synaptic input to the hippocampal dentate gyrus. Previous studies have ascribed the hyperexcitability of layer II stellate cells to GABAergic interneurons becoming "dormant" after disconnection from their excitatory synaptic inputs, which has been reported to occur during preferential loss of layer III pyramidal cells. We used whole-cell recording from slices of entorhinal cortex in pilocarpine-treated epileptic rats to test the dormant interneuron hypothesis. Hyperexcitability appeared as multiple action potentials and prolonged depolarizations evoked in layer II stellate cells of epileptic rats but not controls. However, blockade of glutamatergic synaptic transmission caused similar percentage reductions in the frequency of spontaneous IPSCs in layer II stellate cells of control and epileptic rats, suggesting similar levels of excitatory synaptic input to GABAergic interneurons. Direct recordings and biocytin labeling revealed two major types of interneurons in layer III whose excitatory synaptic drive in epileptic animals was undiminished. Interneurons in layer III did not appear to be dormant; therefore, we tested whether loss of GABAergic synapses might underlie hyperexcitability of layer II stellate cells. Stereological evidence of fewer GABAergic interneurons, fewer gephyrin-immunoreactive punctae, and reduced frequency of spontaneous IPSCs and miniature IPSCs (recorded in tetrodotoxin) confirmed that layer II stellate cell hyperexcitability is attributable, at least in part, to reduced inhibitory synaptic input.

  View details for DOI 10.1523/JNEUROSCI.0064-06.2006

  View details for Web of Science ID 000237271400020

  View details for PubMedID 16641241

• GABA(A) receptor-mediated IPSCs and alpha 1 subunit expression are not reduced in the substantia nigra pars reticulata of gerbils with inherited epilepsy JOURNAL OF NEUROPHYSIOLOGY Kumar, S. S., Wen, X. L., Yang, Y. F., Buckmaster, P. S. 2006; 95 (4): 2446-2455

  Abstract

  Domestic Mongolian gerbils, a model of inherited epilepsy, begin having spontaneous seizures at approximately 1.5 mo of age, making it possible to evaluate them during epileptic and pre-epileptic stages. Previous studies have shown that GABA binding is reduced in the substantia nigra pars reticulata (SNr) of both epileptic and pre-epileptic gerbils compared with controls, suggesting that reduced expression of GABAA receptors in SNr might be epileptogenic in this model. To test this hypothesis, we measured the expression of the GABAA receptor alpha1 subunit, the dominant alpha subunit expressed in the SNr, and evaluated GABAA receptor-mediated postsynaptic currents in SNr neurons. GABA(A) alpha1 subunit mRNA levels in substantia nigra-rich tissue from pre-epileptic animals were similar to controls, and immunocytochemistry for the alpha1 subunit showed similar strong expression in the SNr in both groups. Western analysis confirmed that expression of the alpha1 subunit protein was similar in substantia nigra-rich tissue from pre-epileptic and control gerbils. The frequency and amplitude of spontaneous inhibitory postsynaptic currents (IPSCs) and the frequency of miniature (m)IPSCs in SNr neurons of pre-epileptic gerbil were similar to those of controls. The amplitude of mIPSCs in the pre-epileptics was significantly larger than controls. Zolpidem, an alpha1 subunit-specific modulator of the GABAA receptor, was equally efficacious in prolonging the decay time of mIPSCs in both groups. Hence, contrary to the predictions of the hypothesis, mRNA and protein expression levels of the major GABAA receptor alpha subunit were normal, and neurons of the SNr in pre-epileptic gerbils displayed normal or enhanced IPSC frequencies and amplitudes. Therefore reduced expression of GABAA receptors in SNr is not likely to be an epileptogenic mechanism in this model.

  View details for DOI 10.1152/jn.01173.2005

  View details for Web of Science ID 000236152100038

  View details for PubMedID 16407426

• Stereological analysis of forebrain regions in kainate-treated epileptic rats BRAIN RESEARCH Chen, S. Y., Buckmaster, P. S. 2005; 1057 (1-2): 141-152

  Abstract

  Patients and models of temporal lobe epilepsy display neuron loss in the hippocampal formation, but neuropathological changes also occur in other forebrain regions. We sought to evaluate the specificity and extent of volume loss of the major forebrain regions in epileptic rats months after kainate-induced status epilepticus. In systematic series of Nissl-stained sections, the areas of major forebrain regions were measured, and volumes were estimated using the Cavalieri principle. In some regions, the optical fractionator method was used to estimate neuron numbers. Most kainate-treated rats showed significant volume loss in the amygdala, olfactory cortex, and septal region, but others displayed different patterns, with significant loss only in the hippocampus or thalamus, for example. Average volume loss was most severe in the amygdala and olfactory cortex (82-83% of controls), especially the caudal parts of both regions. In the piriform cortex (including the endopiriform nucleus) of epileptic rats, an average of approximately one-third of Nissl-stained neurons and one-third of the GABAergic interneurons labeled by in situ hybridization for GAD67 mRNA were lost, and the extent of neuron loss was correlated with the extent of volume loss. Volumetric analysis of major forebrain regions was insensitive to specific neuron loss in subregions such as layer III of the entorhinal cortex and the hilus of the dentate gyrus. These findings provide quantitative evidence that kainate-treated rats tend to display extensive neuron and volume loss in the amygdala and olfactory cortex, although the patterns and extent of loss in forebrain regions vary considerably among individuals. In this status epilepticus-based model, extrahippocampal damage appears to be more extensive and hippocampal damage appears to be less extensive than that reported for patients with temporal lobe epilepsy.

  View details for DOI 10.1016/j.brainres.2005.07.058

  View details for Web of Science ID 000232550300017

  View details for PubMedID 16122711

• Prolonged infusion of cycloheximide does not block mossy fiber sprouting in a model of temporal lobe epilepsy EPILEPSIA Toyoda, I., Buckmaster, P. S. 2005; 46 (7): 1017-1020

  Abstract

  The role of protein synthesis in mossy fiber sprouting is unclear. Conflicting reports exist on whether a single dose of the protein synthesis-blocker cycloheximide administered around the time of an epileptogenic injury can block the eventual development of mossy fiber sprouting.In rats, osmotic minipumps and cannulae were implanted to deliver 8 mg/ml cycloheximide to one dentate gyrus and vehicle to the other. This method has been used to block protein synthesis in the infused region for up to 5 days with minimal neurotoxic effects (Taha and Stryker, Neuron 2002;34:425-36). After 2 days of infusion, rats were treated with pilocarpine to induce status epilepticus. Pumps were removed 3 days later. Thirty days after pilocarpine treatment, rats were perfused, and hippocampal sections were processed for Timm staining.Timm staining revealed aberrant mossy fiber sprouting in the inner molecular layer regardless of whether hippocampi were treated with cycloheximide or vehicle. Cycloheximide-treated hippocampi displayed more aberrant Timm staining and more tissue damage around the infusion site than did vehicle-treated hippocampi.Prolonged infusion of cycloheximide, spanning the period of pilocarpine treatment, did not block mossy fiber sprouting. This finding suggests that protein-dependent mechanisms around the time of an epileptogenic injury are not necessary for the eventual development of synaptic reorganization.

  View details for Web of Science ID 000230431800004

  View details for PubMedID 16026553

• Does a unique type of CA3 pyramidal cell in primates bypass the dentate gate? JOURNAL OF NEUROPHYSIOLOGY Buckmaster, P. S. 2005; 94 (1): 896-900

  Abstract

  The predominant excitatory synaptic input to the hippocampus arises from entorhinal cortical axons that synapse with dentate granule cells, which in turn synapse with CA3 pyramidal cells. Thus two highly excitable brain areas--the entorhinal cortex and the CA3 field--are separated by dentate granule cells, which have been proposed to function as a gate or filter. However, unlike rats, primates have "dentate" CA3 pyramidal cells with an apical dendrite that extends into the molecular layer of the dentate gyrus, where they could receive strong, monosynaptic, excitatory synaptic input from the entorhinal cortex. To test this possibility, the dentate gyrus molecular layer was stimulated while intracellular recordings were obtained from CA3 pyramidal cells in hippocampal slices from neurologically normal macaque monkeys. Stimulus intensity of the outer molecular layer of the dentate gyrus was standardized by the threshold intensity for evoking a dentate gyrus field potential population spike. Recorded proximal CA3 pyramidal cells were labeled with biocytin, processed with diaminobenzidine for visualization, and classified according to their dendritic morphology. In response to stimulation of the dentate gyrus molecular layer, action potential thresholds were similar in proximal CA3 pyramidal cells with different dendritic morphologies. These findings do not support the hypothesis that dentate CA3 pyramidal cells receive stronger synaptic input from the entorhinal cortex than do other proximal CA3 pyramidal cells.

  View details for DOI 10.1152/jn.01216.2004

  View details for Web of Science ID 000230135500081

  View details for PubMedID 15800071

• Laboratory animal models of temporal lobe epilepsy COMPARATIVE MEDICINE Buckmaster, P. S. 2004; 54 (5): 473-485

  Abstract

  Temporal lobe epilepsy is a common human disease that is difficult to treat. The pathogenesis of temporal lobe epilepsy, which holds many unresolved questions, and opportunities for creating more effective treatments and preventative strategies are reviewed herein. Laboratory animal models are essential to meet these challenges. How models are created, how they compare with each other and with the disease in human patients, and how they advance our understanding of temporal lobe epilepsy are described.

  View details for Web of Science ID 000225357200003

  View details for PubMedID 15575361

• Recurrent excitation of granule cells with basal dendrites and low interneuron density and inhibitory postsynaptic current frequency in the dentate gyrus of macaque monkeys JOURNAL OF COMPARATIVE NEUROLOGY Austin, J. E., Buckmaster, P. S. 2004; 476 (3): 205-218

  Abstract

  Temporal lobe epilepsy is often associated with pathological changes in the dentate gyrus, and such changes may be more common in humans than in some nonprimate species. To examine species-specific characteristics that might predispose the dentate gyrus to epileptogenic damage, we evaluated recurrent excitation of granule cells with and without basal dendrites in macaque monkeys, measured miniature inhibitory postsynaptic currents (mIPSCs) of granule cells in macaque monkeys and compared them to rats, and estimated the granule cell-to-interneuron ratio in macaque monkeys and rats. In hippocampal slices from monkeys, whole-cell patch recording revealed antidromically evoked excitatory PSCs that were four times larger and inhibitory PSCs that were over two times larger in granule cells with basal dendrites than without. These findings suggest that granule cells with basal dendrites receive more recurrent excitation and, to a lesser degree, more recurrent inhibition. Miniature IPSC amplitude was slightly larger in monkey granule cells with basal dendrites than in those without, but mIPSC frequency was similar and only 26% of that reported for rats. In situ hybridization for glutamic acid decarboxylase and immunocytochemistry for somatostatin, parvalbumin, and neuronal nuclei revealed interneuron proportions and distributions in monkeys that were similar to those reported for rats. However, the interneuron-to-granule cell ratio was lower in monkeys (1:28) than in rats (1:11). These findings suggest that in the primate dentate gyrus, recurrent excitation is enhanced and inhibition is reduced compared with rodents. These primate characteristics may contribute to the susceptibility of the human dentate gyrus to epileptogenic injuries.

  View details for DOI 10.1002/cne.20182

  View details for PubMedID 15269966

• Prolonged infusion of tetrodotoxin does not block mossy fiber sprouting in pilocarpine-treated rats EPILEPSIA Buckmaster, P. S. 2004; 45 (5): 452-458

  Abstract

  Mossy fiber sprouting is a common abnormality found in patients and models of temporal lobe epilepsy. The role of mossy fiber sprouting in epileptogenesis is unclear, and its blockade would be useful experimentally and perhaps therapeutically. Results from previous attempts to block mossy fiber sprouting have been disappointing or controversial. In some brain regions, prolonged application of the sodium channel blocker tetrodotoxin prevents axon sprouting and posttrauma epileptogenesis. The present study tested the hypothesis that prolonged, focal infusion of tetrodotoxin would block mossy fiber sprouting after an epileptogenic treatment.Adult rats were treated with pilocarpine to induce status epilepticus. Several hours to 3 days after pilocarpine treatment, a pump with a cannula directed toward the dentate gyrus was implanted to deliver 10 microM tetrodotoxin or vehicle alone at 0.25 microl/h. This method blocks local EEG activity in the hippocampus (Galvan et al. J Neurosci 2000; 20:2904-16). After 28 days of continuous infusion, rats were perfused with fixative, and their hippocampi analyzed anatomically with stereologic techniques.Tetrodotoxin infusion was verified immunocytochemically in tetrodotoxin-treated but not vehicle-treated hippocampi. Tetrodotoxin-infused and vehicle-infused hippocampi displayed similar levels of hilar neuron loss. The Timm stain revealed mossy fiber sprouting regardless of whether hippocampi were treated with tetrodotoxin infusion, vehicle infusion, or neither.Prolonged infusion of tetrodotoxin did not block mossy fiber sprouting. This finding suggests that sodium channel-mediated neuronal activity is not necessary for mossy fiber sprouting after an epileptogenic treatment.

  View details for Web of Science ID 000221213500006

  View details for PubMedID 15101826

• Dendritic morphology, local circuitry, and intrinsic electrophysiology of principal neurons in the entorhinal cortex of macaque monkeys JOURNAL OF COMPARATIVE NEUROLOGY Buckmaster, P. S., Alonso, A., Canfield, D. R., AMARAL, D. G. 2004; 470 (3): 317-329

  Abstract

  Little is known about the neuroanatomical or electrophysiological properties of individual neurons in the primate entorhinal cortex. We have used intracellular recording and biocytin-labeling techniques in the entorhinal slice preparation from macaque monkeys to investigate the morphology and intrinsic electrophysiology of principal neurons. These neurons have previously been studied most extensively in rats. In monkeys, layer II neurons are usually stellate cells, as in rats, but they occasionally have a pyramidal shape. They tend to discharge trains, not bursts, of action potentials, and some display subthreshold membrane potential oscillations. Layer III neurons are pyramidal, and they do not appear to display membrane potential oscillations. The distribution of dendrites and of axon collaterals suggests that neurons in layers II and III are interconnected by a network of associational fibers. Layer V and VI neurons are pyramidal and tend to discharge trains of action potentials. The distribution of dendrites and axon collaterals suggests that there is an associative network of principal neurons in layers V and VI, and they also project axon collaterals toward superficial layers. Importantly, entorhinal cortical neurons in monkeys appear to exhibit significant differences from those in rats. Morphologically, neurons in monkey entorhinal layers II and III have more primary dendrites, more dendritic branches, and greater total dendritic length than in rats. Electrophysiologically, layer II neurons in monkeys exhibit less sag, and subthreshold oscillations are less robust and slower. Some monkey layer III neurons discharge bursts of action potentials that are not found in rats. The interspecies differences revealed by this study may influence information processing and pathophysiological processes in the primate entorhinal cortex. J. Comp. Neurol. 470:317-329, 2004.

  View details for DOI 10.1002/cne.20014

  View details for Web of Science ID 000188782000007

  View details for PubMedID 14755519

• Reduced inhibition and increased output of layer II neurons in the medial entorhinal cortex in a model of temporal lobe epilepsy JOURNAL OF NEUROSCIENCE Kobayashi, M., Wen, X. L., Buckmaster, P. S. 2003; 23 (24): 8471-8479

  Abstract

  Temporal lobe epilepsy is the most common type of epilepsy in adults, and its underlying mechanisms are unclear. To investigate how the medial entorhinal cortex might contribute to temporal lobe epilepsy, we evaluated the histology and electrophysiology of slices from rats 3-7 d after an epileptogenic injury (pilocarpine-induced status epilepticus). Nissl staining, NeuN immunocytochemistry, and in situ hybridization for GAD65 mRNA were used to verify the preferential loss of glutamatergic neurons and the relative sparing of GABAergic interneurons in layer III. From slices adjacent to those that were used for anatomy, we obtained whole-cell patch recordings from layer II medial entorhinal cortical neurons. Recordings under current-clamp conditions revealed similar intrinsic electrophysiological properties (resting membrane potential, input resistance, single spike, and repetitive firing properties) to those of controls. Spontaneous IPSCs were less frequent (68% of controls), smaller in amplitude (57%), and transferred less charge (51%) than in controls. However, the frequency, amplitude, and rise time of miniature IPSCs were normal. These findings suggest that after epileptogenic injuries the layer II entorhinal cortical neurons receive less GABA(A) receptor-mediated synaptic input because presynaptic inhibitory interneurons become less active. To investigate the possible consequences of reduced spontaneous inhibitory input to layer II neurons, we recorded field potentials in the dentate gyrus, their major synaptic target. At 5 d after pilocarpine-induced status epilepticus the spontaneous field potentials recorded in vivo were over three times more frequent than in controls. These findings suggest that an epileptogenic injury reduces inhibition of layer II neurons and results in excessive synaptic input to the dentate gyrus.

  View details for Web of Science ID 000185452900006

  View details for PubMedID 13679415

• Reduced inhibition of dentate granule cells in a model of temporal lobe epilepsy JOURNAL OF NEUROSCIENCE Kobayashi, M., Buckmaster, P. S. 2003; 23 (6): 2440-2452

  Abstract

  Patients and models of temporal lobe epilepsy have fewer inhibitory interneurons in the dentate gyrus than controls, but it is unclear whether granule cell inhibition is reduced. We report the loss of GABAergic inhibition of granule cells in the temporal dentate gyrus of pilocarpine-induced epileptic rats. In situ hybridization for GAD65 mRNA and immunocytochemistry for parvalbumin and somatostatin confirmed the loss of inhibitory interneurons. In epileptic rats, granule cells had prolonged EPSPs, and they discharged more action potentials than controls. Although the conductances of evoked IPSPs recorded in normal ACSF were not significantly reduced and paired-pulse responses showed enhanced inhibition of granule cells from epileptic rats, more direct measures of granule cell inhibition revealed significant deficiencies. In granule cells from epileptic rats, evoked monosynaptic IPSP conductances were <40% of controls, and the frequency of GABA(A) receptor-mediated spontaneous and miniature IPSCs (mIPSCs) was <50% of controls. Within 3-7 d after pilocarpine-induced status epilepticus, miniature IPSC frequency had decreased, and it remained low, without functional evidence of compensatory synaptogenesis by GABAergic axons in chronically epileptic rats. Both parvalbumin- and somatostatin-immunoreactive interneuron numbers and the frequency of both fast- and slow-rising GABA(A) receptor-mediated mIPSCs were reduced, suggesting that loss of inhibitory synaptic input to granule cells occurred at both proximal/somatic and distal/dendritic sites. Reduced granule cell inhibition in the temporal dentate gyrus preceded the onset of spontaneous recurrent seizures by days to weeks, so it may contribute, but is insufficient, to cause epilepsy.

  View details for Web of Science ID 000181776900051

  View details for PubMedID 12657704

• Axon sprouting in a model of temporal lobe epilepsy creates a predominantly excitatory feedback circuit JOURNAL OF NEUROSCIENCE Buckmaster, P. S., Zhang, G. F., Yamawaki, R. 2002; 22 (15): 6650-6658

  Abstract

  The most common type of epilepsy in adults is temporal lobe epilepsy. After epileptogenic injuries, dentate granule cell axons (mossy fibers) sprout and form new synaptic connections. Whether this synaptic reorganization strengthens recurrent inhibitory circuits or forms a novel recurrent excitatory circuit is unresolved. We labeled individual granule cells in vivo, reconstructed sprouted mossy fibers at the EM level, and identified postsynaptic targets with GABA immunocytochemistry in the pilocarpine model of temporal lobe epilepsy. Granule cells projected an average of 1.0 and 1.1 mm of axon into the granule cell and molecular layers, respectively. Axons formed an average of one synapse every 7 microm in the granule cell layer and every 3 microm in the molecular layer. Most synapses were with spines (76 and 98% in the granule cell and molecular layers, respectively). Almost all of the synapses were with GABA-negative structures (93 and 96% in the granule cell and molecular layers, respectively). By integrating light microscopic and EM data, we estimate that sprouted mossy fibers form an average of over 500 new synapses per granule cell, but <25 of the new synapses are with GABAergic interneurons. These findings suggest that almost all of the synapses formed by mossy fibers in the granule cell and molecular layers are with other granule cells. Therefore, after epileptogenic treatments that kill hilar mossy cells, mossy fiber sprouting does not simply replace one recurrent excitatory circuit with another. Rather, it replaces a distally distributed and disynaptic excitatory feedback circuit with one that is local and monosynaptic.

  View details for Web of Science ID 000177182700040

  View details for PubMedID 12151544

• Evoked responses of the dentate gyrus during seizures in developing gerbils with inherited epilepsy JOURNAL OF NEUROPHYSIOLOGY Buckmaster, P. S., Wong, E. H. 2002; 88 (2): 783-793

  Abstract

  When they are 1-2 mo old, domesticated Mongolian gerbils begin having initially mild seizures which become more severe with age. To evaluate the development of this increasing seizure severity, we obtained field potential responses of the dentate gyrus to paired-pulse stimulation of the perforant path during seizures. In 18 gerbils that were 1.5-8.0 mo old, 73 seizures were analyzed. We measured population spike amplitude, the slope of the field excitatory postsynaptic potential (fEPSP), and the population spike amplitude ratio (2nd/1st) to evaluate excitatory and inhibitory synaptic processes. In gerbils <2 mo old, exposure to a novel environment was followed by an increase in population spike amplitude and then by seizure onset, but population spike amplitude ratio and fEPSP slope remained at baseline levels, and multiple population spikes were never evoked. As previously reported for chronically epileptic gerbils, these findings provide little evidence of a disinhibitory seizure-initiating mechanism in the dentate gyrus when young gerbils begin having seizures. In young gerbils evoked responses changed little during the behaviorally mild seizures. In contrast, most seizures in older gerbils included generalized convulsions, postictal depression, and evoked responses that changed dramatically. In older gerbils, shortly after seizure onset the dentate gyrus became hyperexcitable. Population spike amplitude and fEPSP slope peaked, and multiple population spikes were evoked, suggesting that mechanisms for seizure amplification and spread are more developed in older gerbils. Next, dentate gyrus excitability decreased precipitously, and population spike amplitude and fEPSP slope diminished. This period of hypoexcitability began before the end of the seizure, suggesting it may contribute to seizure termination. After the convulsive phase of the seizure, older gerbils remained motionless during a period of postictal depression, and population spike amplitude remained suppressed until the abrupt switch to normal exploratory activity. These findings suggest that the mechanisms of postictal depression may suppress granule cell excitability. The population spike amplitude ratio peaked after the convulsive phase and then gradually returned to the baseline level an average of 12 min after seizure onset, suggesting that granule cell inhibition recovers within minutes after a spontaneous seizure. Although it is unclear whether the seizure-related changes in evoked responses are a cause or an effect of increased seizure severity in older gerbils, their analysis provides clues about developmental changes in the mechanisms of seizure spread and termination.

  View details for DOI 10.1152/jn.00067.2002

  View details for Web of Science ID 000177276100022

  View details for PubMedID 12163530

• Axon arbors and synaptic connections of a vulnerable population of interneurons in the dentate gyrus in vivo JOURNAL OF COMPARATIVE NEUROLOGY Buckmaster, P. S., Yamawaki, R., Zhang, G. F. 2002; 445 (4): 360-373

  Abstract

  The predominant gamma-aminobutyric acid (GABA)ergic neuron class in the hilus of the dentate gyrus consists of spiny somatostatinergic interneurons. We examined the axon projections and synaptic connections made by spiny hilar interneurons labeled with biocytin in gerbils in vivo. Axon length was 152-497 mm/neuron. Sixty to 85% of the axon concentrated in the outer two thirds of the molecular layer of the dentate gyrus. The septotemporal span of the axon arbor extended over 48-82% of the total hippocampal length, which far exceeds the septotemporal span of axons of granule cells whose complete axon arbors extended over 15-29%. A three-dimensionally reconstructed 216-microm-long spiny hilar interneuron axon segment in the outer third of the molecular layer formed an average of 1 synapse every 5.1 microm. Of the 42 symmetric (inhibitory) synapses formed by the reconstructed segment, 88% were with spiny dendrites of presumed granule cells, and 67% were with dendritic spines that also receive an asymmetric (excitatory) contact from an unlabeled axon terminal. Postembedding GABA-immunocytochemistry revealed that 55% of the GABAergic synapses in the outer third of the molecular layer were with spines. Therefore, in the outer molecular layer, spiny hilar interneurons form synaptic contacts that appear to be positioned to exert inhibitory control near sites of excitatory synaptic input from the entorhinal cortex to granule cell dendritic spines. These findings demonstrate far-reaching, yet highly specific, connectivity of individual interneurons and suggest that the loss of spiny hilar interneurons, as occurs in temporal lobe epilepsy, may contribute to hyperexcitability in the hippocampus.

  View details for DOI 10.1002/cne.10183

  View details for Web of Science ID 000174520200006

  View details for PubMedID 11920713

• BDNF overexpression increases dendrite complexity in hippocampal dentate gyrus NEUROSCIENCE Tolwani, R. J., Buckmaster, P. S., Varma, S., Cosgaya, J. M., Wu, Y., Suri, C., Shooter, E. M. 2002; 114 (3): 795-805

  Abstract

  There is increasing evidence that brain-derived neurotrophic factor (BDNF) modulates synaptic and morphological plasticity in the developing and mature nervous system. Plasticity may be modulated partially by BDNF's effects on dendritic structure. Utilizing transgenic mice where BDNF overexpression was controlled by the beta-actin promoter, we evaluated the effects of long-term overexpression of BDNF on the dendritic structure of granule cells in the hippocampal dentate gyrus. BDNF transgenic mice provided the opportunity to investigate the effects of modestly increased BDNF levels on dendrite structure in the complex in vivo environment. While the elevated BDNF levels were insufficient to change levels of TrkB receptor isoforms or downstream TrkB signaling, they did increase dendrite complexity of dentate granule cells. These cells showed an increased number of first order dendrites, of total dendritic length and of total number of branch points. These results suggest that dendrite structure of granule cells is tightly regulated and is sensitive to modest increases in levels of BDNF. This is the first study to evaluate the effects of BDNF overexpression on dendrite morphology in the intact hippocampus and extends previous in vitro observations that BDNF influences synaptic plasticity by increasing complexity of dendritic arbors.

  View details for Web of Science ID 000178439300025

  View details for PubMedID 12220579

• Heightened seizure severity in somatostatin knockout mice EPILEPSY RESEARCH Buckmaster, P. S., Otero-Corchon, V., Rubinstein, M., Low, M. J. 2002; 48 (1-2): 43-56

  Abstract

  Patients and experimental models of temporal lobe epilepsy display loss of somatostatinergic neurons in the dentate gyrus. To determine if loss of the peptide somatostatin contributes to epileptic seizures we examined kainate-evoked seizures and kindling in somatostatin knockout mice. Somatostatin knockout mice were not observed to experience spontaneous seizures. Timm staining, acetylcholinesterase histochemistry, and immunocytochemistry for NPY, calbindin, calretinin, and parvalbumin revealed no compensatory changes or developmental abnormalities in the dentate gyrus of somatostatin knockout mice. Optical fractionator counting of Nissl-stained hilar neurons showed similar numbers of neurons in wild type and somatostatin knockout mice. Mice were treated systemically with kainic acid to evoke limbic seizures. Somatostatin knockout mice tended to have a shorter average latency to stage 5 seizures, their average maximal behavioral seizure score was higher, and they tended to be more likely to die than controls. In response to kindling by daily electrical stimulation of the perforant path, to more specifically challenge the dentate gyrus, mean afterdischarge duration in somatostatin knockout mice was slightly longer, but the number of treatments to five stage 4-5 seizures was similar to controls. Although we cannot exclude the possibility of undetected compensatory mechanisms in somatostatin knockout mice, these findings suggest that somatostatin may be mildly anticonvulsant, but its loss alone is unlikely to account for seizures in temporal lobe epilepsy.

  View details for Web of Science ID 000173916400005

  View details for PubMedID 11823109

• Absence of temporal lobe epilepsy pathology in dogs with medically intractable epilepsy JOURNAL OF VETERINARY INTERNAL MEDICINE Buckmaster, P. S., Smith, M. O., Buckmaster, C. L., LeCouteur, R. A., Dudek, F. E. 2002; 16 (1): 95-99

  Abstract

  Epilepsy is a common neurological problem in dogs. In some dogs, seizures cannot be controlled adequately with anticonvulsant medication. Temporal lobe epilepsy is the most common type of epilepsy in adult humans, it is frequently resistant to anticonvulsant therapy, and it is commonly associated with characteristic neuropathological abnormalities in the hippocampal dentate gyrus. We sought to test the hypothesis that dogs with medically intractable epilepsy have temporal lobe epilepsy. The hippocampi of 6 dogs that were euthanized because of chronic, recurrent seizures were compared with those of 8 nonepileptic controls. In control and epileptic dogs, stereological cell counting showed similar numbers of neurons in the hilus of the dentate gyrus, somatostatin immunoreactivity identified numerous immunopositive neurons in the hilus, and Timm staining showed the normal pattern of granule cell axon projections. These findings demonstrate a lack of hilar neuron loss and granule cell axon reorganization, suggesting that temporal lobe epilepsy is not a common cause of medically intractable epilepsy in dogs.

  View details for Web of Science ID 000173430700013

  View details for PubMedID 11822812

• Intracellular recording and labeling of mossy cells and proximal CA3 pyramidal cells in macaque monkeys JOURNAL OF COMPARATIVE NEUROLOGY Buckmaster, P. S., Amaral, D. G. 2001; 430 (2): 264-281

  Abstract

  Little is known about the morphological characteristics and intracellular electrophysiological properties of neurons in the primate hippocampus and dentate gyrus. We have therefore begun a program of studies using intracellular recording and biocytin labeling in hippocampal slices from macaque monkeys. In the current study, we investigated mossy cells and proximal CA3 pyramidal cells. As in rats, macaque mossy cells display fundamentally different traits than proximal CA3 pyramidal cells. Interestingly, macaque mossy cells and CA3 pyramidal neurons display some morphological differences from those in rats. Macaque monkey mossy cells extend more dendrites into the molecular layer of the dentate gyrus, have more elaborate thorny excrescences on their proximal dendrites, and project more axon collaterals into the CA3 region. In macaques, three types of proximal CA3 pyramidal cells are found: classical pyramidal cells, neurons with their dendrites confined to the CA3 pyramidal cell layer, and a previously undescribed cell type, the "dentate" CA3 pyramidal cell, whose apical dendrites extend into and ramify within the hilus, granule cell layer, and molecular layer of the dentate gyrus. The basic electrophysiological properties of mossy cells and proximal CA3 cells are similar to those reported for the rodent. Mossy cells have a higher frequency of large amplitude spontaneous depolarizing postsynaptic potentials, and proximal CA3 pyramidal cells are more likely to discharge bursts of action potentials. Although mossy cells and CA3 pyramidal cells in macaque monkeys display many morphological and electrophysiological features described in rodents, these findings highlight significant species differences, with more heterogeneity and the potential for richer interconnections in the primate hippocampus.

  View details for Web of Science ID 000166232700010

  View details for PubMedID 11135261

• Somatostatin-immunoreactive interneurons contribute to lateral inhibitory circuits in the dentate gyrus of control and epileptic rats HIPPOCAMPUS Boyett, J. M., Buckmaster, P. S. 2001; 11 (4): 418-422

  Abstract

  Lateral inhibition, a feature of neuronal circuitry that enhances signaling specificity, has been demonstrated in the rat dentate gyrus. However, neither the underlying neuronal circuits, nor the ways in which these circuits are altered in temporal lobe epilepsy, are completely understood. This study examines the potential contribution of one class of inhibitory interneurons to lateral inhibitory circuits in the dentate gyrus of both control and epileptic rats. The retrograde tracer wheat germ ag-glutinin-apo-horse radish peroxidase-gold (WGA-apo-HRP-gold) was injected into the septal dentate gyrus. Neurons double-labeled for glutamic acid decarboxylase (GAD) and the retrograde tracer are concentrated in the hilus and may contribute to lateral inhibition. Neurons double-labeled for somatostatin and the retrograde tracer account for at least 28% of GAD-positive neurons with axon projections appropriate for generating lateral inhibition in control rats. Despite an overall loss of somatostatin-expressing cells in epileptic animals, the number of somatostatin-positive interneurons with axon projections appropriate for generating lateral inhibition is similar to that seen in controls. These findings suggest that somatostatinergic interneurons participate in lateral inhibitory circuits in the dentate gyrus of both control and epileptic rats, and that surviving somatostatinergic interneurons might sprout new axon collaterals in epileptic animals.

  View details for Web of Science ID 000170601900009

  View details for PubMedID 11530846

• Testing the disinhibition hypothesis of epileptogenesis in vivo and during spontaneous seizures JOURNAL OF NEUROSCIENCE Buckmaster, P. S., Jongen-Relo, A. L., Davari, S. B., Wong, E. H. 2000; 20 (16): 6232-6240

  Abstract

  The "disinhibition" hypothesis contends that (1) seizures begin when granule cells in the dentate gyrus of the dorsal hippocampus are disinhibited and (2) disinhibition occurs because GABAergic interneurons are excessively inhibited by other GABAergic interneurons. We tested the disinhibition hypothesis using the experimental model that inspired it-naturally epileptic Mongolian gerbils. To determine whether there is an excess of GABAergic interneurons in the dentate gyrus of epileptic gerbils, as had been reported previously, GABA immunocytochemistry, in situ hybridization of GAD67 mRNA, and the optical fractionator method were used. There were no significant differences in the numbers of GABAergic interneurons. To determine whether granule cells in epileptic gerbils were disinhibited during the interictal period, IPSPs were recorded in vivo with hippocampal circuits intact in urethane-anesthetized gerbils. The reversal potentials and conductances of IPSPs in granule cells in epileptic versus control gerbils were similar. To determine whether the level of inhibitory control in the dentate gyrus transiently decreases before seizure onset, field potential responses to paired-pulse perforant path stimulation were obtained from the dorsal hippocampus while epileptic gerbils experienced spontaneous seizures. Evidence of reduced inhibition was found after, but not before, seizure onset, indicating that seizures are not triggered by disinhibition in this region. However, seizure-induced depression of inhibition may amplify and promote the spread of seizure activity to other brain regions. These findings do not support the disinhibition hypothesis and suggest that in this model of epilepsy seizures initiate by another mechanism or at a different site.

  View details for Web of Science ID 000088676400040

  View details for PubMedID 10934273

• Highly specific neuron loss preserves lateral inhibitory circuits in the dentate gyrus of kainate-induced epileptic rats JOURNAL OF NEUROSCIENCE Buckmaster, P. S., Jongen-Relo, A. L. 1999; 19 (21): 9519-9529

  Abstract

  Patients with temporal lobe epilepsy display neuron loss in the hilus of the dentate gyrus. This has been proposed to be epileptogenic by a variety of different mechanisms. The present study examines the specificity and extent of neuron loss in the dentate gyrus of kainate-treated rats, a model of temporal lobe epilepsy. Kainate-treated rats lose an average of 52% of their GAD-negative hilar neurons (putative mossy cells) and 13% of their GAD-positive cells (GABAergic interneurons) in the dentate gyrus. Interneuron loss is remarkably specific; 83% of the missing GAD-positive neurons are somatostatin-immunoreactive. Of the total neuron loss in the hilus, 97% is attributed to two cell types-mossy cells and somatostatinergic interneurons. The retrograde tracer wheat germ agglutinin (WGA)-apoHRP-gold was used to identify neurons with appropriate axon projections for generating lateral inhibition. Previously, it was shown that lateral inhibition between regions separated by 1 mm persists in the dentate gyrus of kainate-treated rats with hilar neuron loss. Retrogradely labeled GABAergic interneurons are found consistently in sections extending 1 mm septotemporally from the tracer injection site in control and kainate-treated rats. Retrogradely labeled putative mossy cells are found up to 4 mm from the injection site, but kainate-treated rats have fewer than controls, and in several kainate-treated rats virtually all of these cells are missing. These findings support hypotheses of temporal lobe epileptogenesis that involve mossy cell and somatostatinergic neuron loss and suggest that lateral inhibition in the dentate gyrus does not require mossy cells but, instead, may be generated directly by GABAergic interneurons.

  View details for Web of Science ID 000083177900039

  View details for PubMedID 10531454

• Neuron loss and axon reorganization in the dentate gyrus of cats infected with the feline immunodeficiency virus JOURNAL OF COMPARATIVE NEUROLOGY Mitchell, T. W., Buckmaster, P. S., Hoover, E. A., Whalen, L. R., Dudek, F. E. 1999; 411 (4): 563-577

  Abstract

  The pathophysiological bases of cognitive, motor, and behavioral abnormalities in patients infected with the human immunodeficiency virus (HIV-1) remain largely unknown. To test the possibility that changes in hippocampal neuronal structure may contribute to these neurologic abnormalities, we examined the brains of cats infected with the feline immunodeficiency virus (FIV), an animal model of HIV-1 infection. We evaluated the dentate gyrus by using Timm's staining to estimate the extent of granule cell axon reorganization and by using Nissl staining, immunocytochemistry, and the optical fractionator method to estimate changes in the number of different neuronal subtypes. FIV-infected cats had abnormally high amounts of Timm's staining in the inner molecular layer and granule cell layer and loss of Nissl-stained, somatostatin-immunoreactive, and parvalbumin-immunoreactive neurons in the hilus. An inverse correlation existed between hilar neuron numbers and extent of aberrant Timm's staining. Increased Timm's staining and hilar neuron loss occurred throughout the septotemporal axis of the hippocampus. This type of neuronal loss and synaptic reorganization may provide an anatomic basis for some of the neurologic symptoms found in FIV-infected cats and HIV-infected humans.

  View details for Web of Science ID 000081734900003

  View details for PubMedID 10421868

• In vivo intracellular analysis of granule cell axon reorganization in epileptic rats JOURNAL OF NEUROPHYSIOLOGY Buckmaster, P. S., Dudek, F. E. 1999; 81 (2): 712-721

  Abstract

  In vivo intracellular recording and labeling in kainate-induced epileptic rats was used to address questions about granule cell axon reorganization in temporal lobe epilepsy. Individually labeled granule cells were reconstructed three dimensionally and in their entirety. Compared with controls, granule cells in epileptic rats had longer average axon length per cell; the difference was significant in all strata of the dentate gyrus including the hilus. In epileptic rats, at least one-third of the granule cells extended an aberrant axon collateral into the molecular layer. Axon projections into the molecular layer had an average summed length of 1 mm per cell and spanned 600 microm of the septotemporal axis of the hippocampus-a distance within the normal span of granule cell axon collaterals. These findings in vivo confirm results from previous in vitro studies. Surprisingly, 12% of the granule cells in epileptic rats, and none in controls, extended a basal dendrite into the hilus, providing another route for recurrent excitation. Consistent with recurrent excitation, many granule cells (56%) in epileptic rats displayed a long-latency depolarization superimposed on a normal inhibitory postsynaptic potential. These findings demonstrate changes, occurring at the single-cell level after an epileptogenic hippocampal injury, that could result in novel, local, recurrent circuits.

  View details for Web of Science ID 000078832800029

  View details for PubMedID 10036272

• Recurrent spontaneous motor seizures after repeated low-dose systemic treatment with kainate: assessment of a rat model of temporal lobe epilepsy EPILEPSY RESEARCH Hellier, J. L., Patrylo, P. R., Buckmaster, P. S., Dudek, F. E. 1998; 31 (1): 73-84

  Abstract

  Human temporal lobe epilepsy is associated with complex partial seizures that can produce secondarily generalized seizures and motor convulsions. In some patients with temporal lobe epilepsy, the seizures and convulsions occur following a latent period after an initial injury and may progressively increase in frequency for much of the patient's life. Available animal models of temporal lobe epilepsy are produced by acute treatments that often have high mortality rates and/or are associated with a low proportion of animals developing spontaneous chronic motor seizures. In this study, rats were given multiple low-dose intraperitoneal (i.p.) injections of kainate in order to minimize the mortality rate usually associated with single high-dose injections. We tested the hypothesis that these kainate-treated rats consistently develop a chronic epileptic state (i.e. long-term occurrence of spontaneous, generalized seizures and motor convulsions) following a latent period after the initial treatment. Kainate (5 mg/kg per h, i.p.) was administered to rats every hour for several hours so that class III-V seizures were elicited for > or = 3 h, while control rats were treated similarly with saline. This treatment protocol had a relatively low mortality rate (15%). After acute treatment, rats were observed for the occurrence of motor seizures for 6-8 h/week. Nearly all of the kainate-treated rats (97%) had two or more spontaneous motor seizures months after treatment. With this observation protocol, the average latency for the first spontaneous motor seizure was 77+/-38 (+/-S.D.) days after treatment. Although variability was observed between rats, seizure frequency initially increased with time after treatment, and nearly all of the kainate-treated rats (91%) had spontaneous motor seizures until the time of euthanasia (i.e. 5-22 months after treatment). Therefore, multiple low-dose injections of kainate, which cause recurrent motor seizures for > or = 3 h, lead to the development of a chronic epileptic state that is characterized by (i) a latent period before the onset of chronic motor seizures, and (ii) a high but variable seizure frequency that initially increases with time after the first chronic seizure. This modification of the kainate-treatment protocol is efficient and relatively simple, and the properties of the chronic epileptic state appear similar to severe human temporal lobe epilepsy. Furthermore, the observation that seizure frequency initially increased as a function of time after kainate treatment supports the hypothesis that temporal lobe epilepsy can be a progressive syndrome.

  View details for Web of Science ID 000074539900007

  View details for PubMedID 9696302

• Axonal sprouting in hippocampus of cats infected with feline immunodeficiency virus (FIV) JOURNAL OF ACQUIRED IMMUNE DEFICIENCY SYNDROMES Mitchell, T. W., Buckmaster, P. S., Hoover, E. A., Whalen, L. R., Dudek, F. E. 1998; 17 (1): 1-8

  Abstract

  Neurologic dysfunction and neuropathology are common findings in patients infected with HIV and in cats infected with feline immunodeficiency virus (FIV). The pathogenesis of lentivirus-associated alterations in the central nervous system (CNS) is multifactorial. Because seizures, alterations in memory, and behavioral changes are clinical manifestations in adults and children infected with HIV, we explored the possibility that changes in neuronal structure may occur in the hippocampus. To do this, we examined the dentate gyrus of FIV-infected cats, an animal model of HIV infection. Neuropathologic findings included gliosis within the hilus of the dentate gyrus and granule cell axonal sprouting. Using the Timm's method, which labels axons of dentate gyrus granule cells, abnormally high amounts of staining were observed in the inner one third of the molecular layer in 45% of FIV-infected cats (n = 11) and in none of the controls (n = 19). Prominent axonal sprouting was seen in three FIV-infected cats that were infected as kittens, suggesting that younger cats may be more susceptible. Axon reorganization of the dentate granule cells has been hypothesized to underlie complex partial seizure activity in human temporal lobe epilepsy. These results suggest that FIV infection causes granule cell axon reorganization in the hippocampus of cats. A similar neuropathogenetic mechanism may contribute to neurologic dysfunction in HIV-infected patients.

  View details for Web of Science ID 000071451900001

  View details for PubMedID 9436752

• Neuron loss, granule cell axon reorganization, and functional changes in the dentate gyrus of epileptic kainate-treated rats JOURNAL OF COMPARATIVE NEUROLOGY Buckmaster, P. S., Dudek, F. E. 1997; 385 (3): 385-404

  Abstract

  We sought to describe quantitatively the morphological and functional changes that occur in the dentate gyrus of kainate-treated rats, an experimental model of temporal lobe epilepsy. Adult rats were treated systemically with kainic acid, and, months later, after displaying spontaneous recurrent motor seizures, their dentate gyri were examined. Histological, immunocytochemical, and quantitative stereological techniques were used to estimate numbers of neurons per dentate gyrus of various classes and to estimate the extent of granule cell axon reorganization along the septotemporal axis of the hippocampus in control rats and epileptic kainate-treated rats. Compared with control rats, epileptic kainate-treated rats had fewer Nissl-stained hilar neurons and fewer somatostatin-immunoreactive neurons. There was a correlation between the extent of hilar neuron loss and the extent of somatostatin-immunoreactive neuron loss. However, functional inhibition in the dentate gyrus, assessed with paired-pulse responses to perforant-pathway stimulation, revealed enhanced, and not the expected reduced, inhibition in epileptic kainate-treated rats. Numbers of parvalbumin- and cholecystokinin-immunoreactive neurons were similar in control rats and in most kainate-treated rats. A minority (36%) of the epileptic kainate-treated rats had fewer parvalbumin- and cholecystokinin-immunoreactive neurons than control rats, and those few (8%) with extreme loss in these interneuron classes showed markedly hyperexcitable dentate gyrus field-potential responses to orthodromic stimulation. Compared with control rats, epileptic kainate-treated rats had larger proportions of their granule cell and molecular layers infiltrated with Timm stain. There was a correlation between the extent of abnormal Timm staining and the extent of hilar neuron loss. Granule cell axon reorganization and dentate gyrus neuron loss were more severe in temporal vs. septal hippocampus. These findings from the dentate gyrus of epileptic kainate-treated rats are strikingly similar to those reported for human temporal lobe epilepsy, and they suggest that neuron loss and axon reorganization in the temporal hippocampus may be important in epileptogenesis.

  View details for Web of Science ID A1997XU67500004

  View details for PubMedID 9300766

• Network properties of the dentate gyrus in epileptic rats with hilar neuron loss and granule cell axon reorganization JOURNAL OF NEUROPHYSIOLOGY Buckmaster, P. S., Dudek, F. E. 1997; 77 (5): 2685-2696

  Abstract

  Neuron loss in the hilus of the dentate gyrus and granule cell axon reorganization have been proposed as etiologic factors in human temporal lobe epilepsy. To explore these possible epileptogenic mechanisms, electrophysiological and anatomic methods were used to examine the dentate gyrus network in adult rats that had been treated systemically with kainic acid. All kainate-treated rats, but no age-matched vehicle-treated controls, were observed to have spontaneous recurrent motor seizures beginning weeks to months after exposure to kainate. Epileptic kainate-treated rats and control animals were anesthetized for field potential recording from the dentate gyrus in vivo. Epileptic kainate-treated rats displayed spontaneous positivities ("dentate electroencephalographic spikes") with larger amplitude and higher frequency than those in control animals. After electrophysiological recording, rats were perfused and their hippocampi were processed for Nissl and Timm staining. Epileptic kainate-treated rats displayed significant hilar neuron loss and granule cell axon reorganization. It has been hypothesized that hilar neuron loss reduces lateral inhibition in the dentate gyrus, thereby decreasing seizure threshold. To assess lateral inhibition, simultaneous recordings were obtained from the dentate gyrus in different hippocampal lamellae, separated by 1 mm. The perforant path was stimulated with paired-pulse paradigms, and population spike amplitudes were measured. Responses were obtained from one lamella while a recording electrode in a distant lamella leaked saline or the gamma-aminobutyric acid-A receptor antagonist bicuculline. Epileptic kainate-treated and control rats both showed significantly more paired-pulse inhibition when a lateral lamella was hyperexcitable. To assess seizure threshold in the dentate gyrus, two techniques were used. Measurement of stimulus threshold for evoking maximal dentate activation revealed significantly higher thresholds in epileptic kainate-treated rats compared with controls. In contrast, epileptic kainate-treated rats were more likely than controls to discharge spontaneous bursts of population spikes and to display stimulus-triggered afterdischarges when a focal region of the dentate gyrus was disinhibited with bicuculline. These spontaneous bursts and afterdischarges were confined to the disinhibited region and did not spread to other septotemporal levels of the dentate gyrus. Epileptic kainate-treated rats that displayed spontaneous bursts and/or afterdischarges had significantly larger percentages of Timm staining in the granule cell and molecular layers than epileptic kainate-treated rats that failed to show spontaneous bursts or afterdischarges. In summary, this study reveals functional abnormalities in the dentate gyri of epileptic kainate-treated rats; however, lateral inhibition persists, suggesting that vulnerable hilar neurons are not necessary for generating lateral inhibition in the dentate gyrus.

  View details for Web of Science ID A1997WZ56300031

  View details for PubMedID 9163384

• Ultrastructural localization of neurotransmitter immunoreactivity in mossy cell axons and their synaptic targets in the rat dentate gyrus HIPPOCAMPUS Wenzel, H. J., Buckmaster, P. S., Anderson, N. L., Wenzel, M. E., Schwartzkroin, P. A. 1997; 7 (5): 559-570

  Abstract

  Electrophysiologically identified and intracellularly biocytin-labeled mossy cells in the dentate hilus of the rat were studied using electron microscopy and postembedding immunogold techniques. Ultrathin sections containing a labeled mossy cell or its axon collaterals were reacted with antisera against the excitatory neurotransmitter glutamate and against the inhibitory neurotransmitter gamma-aminobutyric acid (GABA). From single- and double-immunolabeled preparations, we found that 1) mossy cell axon terminals made asymmetric contacts onto postsynaptic targets in the hilus and stratum moleculare of the dentate gyrus and showed immunoreactivity primarily for glutamate, but never for GABA; 2) in the hilus, glutamate-positive mossy cell axon terminals targeted GABA-positive dendritic shafts of hilar interneurons and GABA-negative dendritic spines; and 3) in the inner molecular layer, the mossy cell axon formed asymmetric synapses with dendritic spines associated with GABA-negative (presumably granule cell) dendrites. The results of this study support the view that excitatory (glutamatergic) mossy cell terminals contact GABAergic interneurons and non-GABAergic neurons in the hilar region and GABA-negative granule cells in the stratum moleculare. This pattern of connectivity is consistent with the hypothesis that mossy cells provide excitatory feedback to granule cells in a dentate gyrus associational network and also activate local hilar inhibitory elements.

  View details for Web of Science ID A1997YB74400011

  View details for PubMedID 9347352

• Electrophysiological correlates of seizure sensitivity in the dentate gyrus of epileptic juvenile and adult gerbils JOURNAL OF NEUROPHYSIOLOGY Buckmaster, P. S., Tam, E., Schwartzkroin, P. A. 1996; 76 (4): 2169-2180

  Abstract

  1. Naturally occurring inherited epilepsy is common among Mongolian gerbils, providing an opportunity to identify neurological factors that correlate with seizure behavior. In the present study we examine the ontogeny of seizure behavior and compare the electrophysiology and anatomy of the dentate gyrus in epileptic and nonepileptic gerbils. 2. Behavioral seizure testing revealed that young gerbils do not begin having seizures until they are 2 mo of age, at which time seizure incidence across animals is at its highest level. Most seizure-positive juvenile gerbils became epileptic adults, but 30% outgrew their epileptic condition. 3. The number of somatostatin- and parvalbumin-immunoreactive neurons in the dentate gyrus and Ammon's horn was counted, with the use of quantitative stereological techniques, in juvenile and adult gerbils. No significant differences were detected between epileptic and nonepileptic groups. 4. In dentate gyrus field potential responses to perforant path stimulation, adult epileptic gerbils showed enhanced paired-pulse inhibition at short (30 ms) interstimulus intervals and enhanced facilitation at intermediate (70 ms) intervals compared with nonepileptic controls. These differences were most pronounced when stimuli were delivered at faster (1.0 Hz) rather than slower (0.1 Hz) rates. 5. Compared with seizure-negative juveniles, seizure-positive juveniles showed the same pattern of paired-pulse response abnormalities as epileptic adults. However, seizure-positive juveniles had a lower threshold for maximal dentate activation than epileptic adults. 6. These results demonstrate similar functional abnormalities in the dentate gyri of epileptic adult gerbils and in juvenile gerbils before they experience multiple seizures. Such findings suggest that abnormalities in functional inhibition of the dentate gyrus network precede and therefore might contribute to overt seizure activity.

  View details for Web of Science ID A1996VN61300003

  View details for PubMedID 8899592

• Axon arbors and synaptic connections of hippocampal mossy cells in the rat in vivo JOURNAL OF COMPARATIVE NEUROLOGY Buckmaster, P. S., Wenzel, H. J., Kunkel, D. D., Schwartzkroin, P. A. 1996; 366 (2): 270-292

  View details for Web of Science ID A1996TX43500008

• Possible mechanisms of seizure-related cell damage in the dentate hilus. Epilepsy research. Supplement Schwartzkroin, P. A., Buckmaster, P. S., Strowbridge, B. W., Kunkel, D. D., Owens, J., Pokorný, J. 1996; 12: 317-324

  View details for PubMedID 9302531

• Neurobiology of hippocampal interneurons: A workshop review HIPPOCAMPUS Buckmaster, P. S., Soltesz, I. 1996; 6 (3): 330-339

  View details for Web of Science ID A1996VA30500009

  View details for PubMedID 8841830

• Physiological and morphological heterogeneity of dentate gyrus-hilus interneurons in the gerbil hippocampus in vivo. European journal of neuroscience Buckmaster, P. S., Schwartzkroin, P. A. 1995; 7 (10): 1393-1402

  Abstract

  A variety of morphological types of dentate gyrus/hilus interneurons have been described, but little is known about their corresponding physiological characteristics. To address this issue, intracellular responses to current injection and perforant path stimulation were obtained from putative dentate interneurons in anaesthetized adult gerbils. Our sample of interneurons showed heterogeneity in their intrinsic physiological characteristics and spike thresholds to perforant path stimulation, suggesting the existence of distinct physiologically-defined classes. 'Fast-spiking' interneurons had a low threshold to perforant path stimulation, whereas 'slow-spiking' interneurons responded with predominantly inhibitory potentials. In several cases, cells were intracellularly labelled with biocytin for visualization. Interneurons with different physiological traits had distinct morphological features. These results confirm that, as in hippocampus proper, morphologically identifiable interneurons in the dentate hilus show electrophysiological features that are likely to reflect functionally specific roles in informational processing.

  View details for PubMedID 8542057

• PHYSIOLOGICAL AND MORPHOLOGICAL HETEROGENEITY OF DENTATE GYRUS-HILUS INTERNEURONS IN THE GERBIL HIPPOCAMPUS IN-VIVO EUROPEAN JOURNAL OF NEUROSCIENCE Buckmaster, P. S., Schwartzkroin, P. A. 1995; 7 (6): 1393-1402

  Abstract

  A variety of morphological types of dentate gyrus/hilus interneurons have been described, but little is known about their corresponding physiological characteristics. To address this issue, intracellular responses to current injection and perforant path stimulation were obtained from putative dentate interneurons in anaesthetized adult gerbils. Our sample of interneurons showed heterogeneity in their intrinsic physiological characteristics and spike thresholds to perforant path stimulation, suggesting the existence of distinct physiologically-defined classes. 'Fast-spiking' interneurons had a low threshold to perforant path stimulation, whereas 'slow-spiking' interneurons responded with predominantly inhibitory potentials. In several cases, cells were intracellularly labelled with biocytin for visualization. Interneurons with different physiological traits had distinct morphological features. These results confirm that, as in hippocampus proper, morphologically identifiable interneurons in the dentate hilus show electrophysiological features that are likely to reflect functionally specific roles in informational processing.

  View details for Web of Science ID A1995RF77500030

  View details for PubMedID 7582114

• INTERNEURONS AND INHIBITION IN THE DENTATE GYRUS OF THE RAT IN-VIVO JOURNAL OF NEUROSCIENCE Buckmaster, P. S., Schwartzkroin, P. A. 1995; 15 (1): 774-789

  Abstract

  Inhibitory cells are critically involved in shaping normal hippocampal function and are thought to be important elements in the development of hippocampal pathologies. However, there is relatively little information about the extent and pattern of axonal arborization of hippocampal interneurons and, therefore, about the sphere of influence of these cells. What we do know about these cells is based largely on in vitro slice studies, in which interneuronal interactions may be severely attenuated. The present study was carried out to provide a more realistic picture of interneuron influence. Intracellular recordings were obtained from dentate interneurons in the intact brain of anesthetized rats, and cells were intracellularly labeled with biocytin. The axonal arbors of two classes of dentate interneurons were traced through the hippocampus; each was found to extend long distances (up to half of the total septotemporal length of the hippocampus) perpendicular to the hippocampal lamellae and to target preferential strata. These results suggest that dentate interneurons have far-reaching effects on target cells in distant hippocampal lamellae. One implication of this finding is that dentate neurons should receive more inhibitory synaptic drive in vivo than in slice preparations, in which many inhibitory axon collaterals are amputated. Synaptic responses to perforant path stimulation were examined in granule cells, mossy cells, and CA3 pyramidal cells in vivo, for comparison with previously published results from hippocampal slice studies. In vivo, all cell types showed excitatory synaptic responses that were brief and limited by robust IPSPs that were larger in amplitude and conductance than responses to comparable stimuli recorded in vitro. This difference could not be explained by a change in the intrinsic physiological properties of the cells in the slice preparation, because those parameters were similar in vivo and in vitro. We conclude that dentate gyrus inhibitory interneurons can affect the excitability of neurons in distant areas of the hippocampus, and that these distant influences cannot be appreciated in conventional in vitro preparations.

  View details for Web of Science ID A1995QB81200026

  View details for PubMedID 7823179

• HIPPOCAMPAL MOSSY CELL-FUNCTION - A SPECULATIVE VIEW HIPPOCAMPUS Buckmaster, P. S., Schwartzkroin, P. A. 1994; 4 (4): 393-402

  View details for Web of Science ID A1994PP27000001

  View details for PubMedID 7874231

• HYPEREXCITABILITY IN THE DENTATE GYRUS OF THE EPILEPTIC MONGOLIAN GERBIL EPILEPSY RESEARCH Buckmaster, P. S., Schwartzkroin, P. A. 1994; 18 (1): 23-28

  Abstract

  There are anatomical differences in the hippocampi of seizure-resistant (SR) vs. seizure-sensitive (SS) Mongolian gerbils. SS gerbils have more GABAergic neurons and more GABAergic axon terminals contacting inhibitory basket cells in the dentate gyrus. Hence, it has been hypothesized that inhibition of basket cells causes disinhibition of granule cells and results in hyperexcitability in SS gerbils. To test this hypothesis we measured the level of excitability in the dentate gyrus of anesthetized adult SS and SR gerbils, using a paired-pulse stimulus paradigm to evoke population spikes. Population spikes of SS and SR gerbils both showed strong paired-pulse inhibition at short interstimulus intervals and paired-pulse facilitation at intermediate interstimulus intervals. However, compared to SR gerbils, SS gerbils had significantly stronger paired-pulse facilitation which persisted at longer interstimulus intervals. These results show that SS gerbils have hyperexcitable dentate gyri and support the view that less effective inhibition is an underlying basis for this hyperexcitability.

  View details for Web of Science ID A1994NN49900003

  View details for PubMedID 8088254

• SOMATOSTATIN-IMMUNOREACTIVITY IN THE HIPPOCAMPUS OF MOUSE, RAT, GUINEA-PIG, AND RABBIT HIPPOCAMPUS Buckmaster, P. S., Kunkel, D. D., Robbins, R. J., Schwartzkroin, P. A. 1994; 4 (2): 167-180

  Abstract

  The hippocampi of species commonly used for in vitro physiologic studies were examined to determine if there were species-specific and regional differences in somatostatin immunoreactivity. The distributions of somatostatin-immunoreactive somata and fiber plexuses were determined, and the concentration of somatostatin along the septotemporal axis of the hippocampus was measured using a radioimmunoassay. There are many similarities in the patterns of somatostatin immunoreactivity in the hippocampi of mice, rats, guinea pigs, and rabbits. All species had a relatively even distribution of somatostatin-positive perikarya across three fields of the hippocampus (dentate gyrus, CA3, and CA1-2), a similar distribution of somatostatin-immunoreactive perikarya across the strata of the CA1-2 field and the dentate gyrus; and more somatostatin-positive cells in temporal than in septal hippocampus. However, there are species-specific differences in the distribution of somatostatin-immunoreactive perikarya across the strata of CA3. In addition, unlike the other species examined, mice appeared not to have a somatostatin-immunoreactive fiber plexus in the molecular layer of the dentate gyrus. The functional significance of these differences remains to be determined.

  View details for Web of Science ID A1994PA05700005

  View details for PubMedID 7951691

• A COMPARISON OF RAT HIPPOCAMPAL MOSSY CELLS AND CA3C PYRAMIDAL CELLS JOURNAL OF NEUROPHYSIOLOGY Buckmaster, P. S., Strowbridge, B. W., Schwartzkroin, P. A. 1993; 70 (4): 1281-1299

  Abstract

  1. There is a long-standing debate about whether the large spiny cells in the hilar region of the hippocampus should be classified as pyramidal cells of Ammon's horn or as a distinct cell type of the dentate gyrus. The rationale for grouping these hilar neurons (termed "mossy cells") with pyramidal cells of Ammon's horn is shared characteristics. In the present study we have compared the morphological and physiological characteristics of mossy cells and nearby CA3c pyramidal cells with the use of a rat hippocampal slice preparation. 2. Biocytin-labeled neurons were examined on the basis of soma area, location, shape, number of primary dendrites, extent of dendritic spines, dendritic location, and axon trajectories. Mossy cells had larger soma areas than CA3c pyramidal cells, and they had more large complex spines (thorny excrescences) on their proximal dendrites and somata than CA3c pyramidal cells. Mossy cell dendritic trees and axon collaterals ramified in different regions of the hippocampus than dendrites and axons of CA3c pyramidal cells. 3. Intrinsic physiological properties, and spontaneous and evoked synaptic properties, were measured and compared. Mossy cells had significantly higher input resistances, smaller amplitude burst afterhyperpolarizations, smaller amplitude action potentials, less spike-frequency adaptation, and more anomalous rectification than CA3c pyramidal cells. 4. Mossy cells had spontaneous excitatory postsynaptic potentials (EPSPs) that were significantly higher in frequency and larger in amplitude than CA3c pyramidal cells. A larger proportion of mossy cells than CA3c pyramidal cells responded to perforant path stimulation with depolarizing postsynaptic potentials without any apparent hyperpolarization. Conversely, a smaller proportion of mossy cells than CA3c pyramidal cells responded to perforant path stimulation with inhibitory postsynaptic potentials (IPSPs), and spontaneous IPSPs were more difficult to detect in mossy cells. 5. The intrinsic physiological properties of mossy cells endow these cells with potent excitatory mechanisms but relatively fewer inhibitory control processes than CA3c pyramidal cells. Recordings of spontaneous and evoked PSPs suggest that mossy cells receive more excitatory input and less inhibitory input than CA3c pyramidal cells. These intrinsic and synaptic properties of mossy cells may explain this cell type's exceptional vulnerability to excitotoxic damage by intense afferent stimulation. 6. In summary, mossy cells were significantly different from CA3c pyramidal cells in many of their morphological, intrinsic physiological, and synaptic properties.(ABSTRACT TRUNCATED AT 400 WORDS)

  View details for Web of Science ID A1993MC01300001

  View details for PubMedID 8283200

• BRAIN-STEM AUDITORY EVOKED-POTENTIAL INTERWAVE INTERVALS ARE PROLONGED IN VITAMIN-B6-DEFICIENT CATS JOURNAL OF NUTRITION Buckmaster, P. S., Holliday, T. A., Bai, S. C., Rogers, Q. R. 1993; 123 (1): 20-26

  Abstract

  Vitamin B-6 deficiency has been reported to produce behavioral, neurophysiological and neuropathological abnormalities in a variety of species. In this investigation we used brainstem auditory evoked potentials (BAEP) to determine if vitamin B-6 deficiency in cats affected peripheral and brainstem auditory pathways. Brainstem auditory evoked potentials were recorded from growing cats as they developed vitamin B-6 deficiency, which was confirmed using clinical, hematological and urinary criteria. The BAEP interwave intervals measured from early (wave 1 or 1N) to late waves (5N) or from middle (wave 3) to late waves increased significantly, whereas interwave intervals from early to middle waves did not differ significantly. These results indicate that vitamin B-6 deficiency affects one or more structures of the brainstem that generate the later parts of the BAEP. The finding of prolonged interwave intervals in vitamin B-6-deficient animals is consistent with slowed axonal conduction velocity secondary to defective myelination. Recording BAEP provided a noninvasive means of detecting effects of vitamin B-6 deficiency on specific parts of the central nervous system.

  View details for Web of Science ID A1993KG33100002

  View details for PubMedID 8421226

• MOSSY CELL AXONAL PROJECTIONS TO THE DENTATE GYRUS MOLECULAR LAYER IN THE RAT HIPPOCAMPAL SLICE HIPPOCAMPUS Buckmaster, P. S., Strowbridge, B. W., Kunkel, D. D., SCHMIEGE, D. L., Schwartzkroin, P. A. 1992; 2 (4): 349-362

  Abstract

  The ipsilateral associational pathway connects different septotemporal levels of the dentate gyrus. Neurons of the dentate hilus project hundreds of micrometers from the cells of origin to the inner molecular layer. The authors hypothesized that mossy cells, the major cell type of the hilus, also project locally to the inner molecular layer. Within a 400 microns slice, mossy cells were (1) recorded intracellularly while the inner molecular layer was stimulated to test for antidromic responses, and (2) labeled with biocytin and examined with light and electron microscopy for axonal projections into the molecular layer. The authors found that mossy cells can be antidromically activated by inner molecular layer stimulation and that axonal projections to the molecular layer can be visualized within a 400 microns hippocampal slice. In 13 of 19 intracellularly labeled and electrophysiologically characterized mossy cells, collaterals could be traced into the molecular layer. These results suggest that mossy cells contribute to the ipsilateral associational pathway and also participate in local recurrent circuitry to influence granule cell activity.

  View details for Web of Science ID A1992JR88600002

  View details for PubMedID 1284975

• POTENTIATION OF SPONTANEOUS SYNAPTIC ACTIVITY IN RAT MOSSY CELLS NEUROSCIENCE LETTERS Strowbridge, B. W., Buckmaster, P. S., Schwartzkroin, P. A. 1992; 142 (2): 205-210

  Abstract

  Recent studies have demonstrated the vulnerability of dentate mossy cells to seizure-induced damage. One source of potentially damaging synaptic input are spontaneously active granule cell terminals ('mossy terminals'.) We sought to test whether there were activity-dependent changes in the spontaneous excitatory input to mossy cells. Using the in vitro slice preparation, we examined the frequency and amplitude of spontaneous excitatory postsynaptic potentials (EPSPs) after intracellular current injection designed to mimic the extreme depolarization these neurons receive during repetitive afferent stimulation. In 4 of 7 neurons, depolarization with trains of current pulses resulted in a significant and persistent increase in frequency of spontaneous synaptic depolarizations (to an average of 178% of the initial baseline rate). In 3 of these affected neurons, an increased frequency of large amplitude, fast-rising EPSPs accounted for the majority of this change. Injection of hyperpolarizing current pulses failed to alter spontaneous activity in 3 other mossy cells. These results suggest spontaneous synaptic input to mossy cells in plastic and can be potentiated by depolarization of a single postsynaptic mossy cell. The ability of mossy cells to potentiate their excitatory input may be relevant to their vulnerability to excitotoxic injury during repetitive afferent stimulation.

  View details for Web of Science ID A1992JK73700024

  View details for PubMedID 1454217

• FASTIGIAL NUCLEUS ACTIVITY IN THE ALERT MONKEY DURING SLOW EYE AND HEAD MOVEMENTS JOURNAL OF NEUROPHYSIOLOGY Buttner, U., FUCHS, A. F., MARKERTSCHWAB, G., Buckmaster, P. 1991; 65 (6): 1360-1371

  Abstract

  1. Single units were recorded extracellularly from the fastigial nucleus of three macaque monkeys. Two untrained animals were subjected to whole-body yaw rotations in the light and dark and to full-field horizontal optokinetic stimuli provided by a drum with vertical stripes. The third also was subjected to sinusoidal yaw rotations but, in addition, was trained to follow a small spot, which moved in various ways relative to the animal, to reveal possible smooth pursuit and vestibular sensitivities. 2. On the basis of their responses to vestibular and optokinetic stimuli and their responses during smooth pursuit, fastigial neurons could be divided functionally into a rostral and a caudal group. 3. Most rostral neurons exhibited an increased firing for contralateral head rotations and ipsilateral optokinetic stimuli. A few had the opposite combination of directional preferences. The average firing rates increased monotonically both with contralateral head velocity and ipsilateral drum velocity and decreased monotonically for the oppositely directed movements. There was no change in firing rate for either spontaneous saccades or smooth pursuit of a small moving spot. 4. In contrast, neurons in the caudal fastigial nuclei not only have a robust vestibular sensitivity, but respond during smooth pursuit as well. Most discharge during contralateral head velocity and contralateral smooth pursuit so that they exhibit very little modulation during the vestibuloocular reflex (VOR) or when the rotating animal is fixating a target stationary in the world (SIW). The remaining neurons discharge during contralateral head rotations but ipsilateral eye rotations; these units exhibit their greatest modulation during the SIW condition. 5. Because they respond during quite different behavioral situations, it seems likely that rostral fastigial neurons are involved with descending control of the somatic musculature, whereas the caudal neurons are involved in oculomotor control. The sparse anatomic and lesion data that is available is consistent with this idea.

  View details for Web of Science ID A1991FQ59000009

  View details for PubMedID 1875245