Fuu Jiun Hwang

All Publications

• Two-photon 3D imaging of optically stimulated neural activity at 100 Hz. Light, science & applications Xu, D., Hwang, F. J., Ding, J. B., Peng, L. 2026; 15 (1)

  Abstract

  Understanding how neurons integrate synaptic inputs requires imaging techniques capable of capturing rapid, three-dimensional dendritic events. These processes occur on millisecond timescales and submicron spatial scales, exceeding the speed of conventional two-photon microscopy (2PM). We developed dual-view Bessel two-photon projection microscopy (dv-B2PM), a high-speed volumetric imaging approach that achieves 100 Hz whole-volume acquisition with synaptic-level resolution. dv-B2PM simultaneously records two orthogonal projections of the same 3D volume, preserving spatial information while minimizing ambiguity from structural overlap. Combining dv-B2PM with two-photon glutamate uncaging, we visualized 3D Ca²⁺ dynamics in neurons following localized stimulation. Multi-timescale analysis revealed dendrite-to-soma Ca²⁺ signal propagation, back propagated Ca²⁺ signal from the soma, and multi-frequency (5-40 Hz) Ca²⁺ transients activated along apical dendrites at speeds from ten of microns per second to millimeters per seconds. These findings demonstrate dv-B2PM as a powerful tool for direct visualization of 3D calcium dynamics associated with dendritic integration across extended neuronal structures, bridging the gap between optical imaging and the dynamic biophysics of neuronal integration.

  View details for DOI 10.1038/s41377-026-02395-2

  View details for PubMedID 42393079

  View details for PubMedCentralID PMC13328388

• Remodelling of corticostriatal axonal boutons during motor learning. Nature Sheng, M., Lu, D., Roth, R. H., Hwang, F. J., Sheng, K., Ding, J. B. 2025

  Abstract

  Motor skill learning induces long-lasting synaptic plasticity at dendritic spines1-4 and at the outputs of motor cortical neurons to the striatum5,6. However, little is known about corticostriatal axon activity and structural plasticity during learning in the adult brain. Here, using longitudinal in vivo two-photon imaging, we tracked thousands of corticostriatal axonal boutons in the dorsolateral striatum of awake mice. We found that learning a new motor skill dynamically regulated these boutons. The activities of motor corticostriatal axonal boutons exhibited selectivity for rewarded movements (RM) and unrewarded movements (UM). Notably, boutons on the same axonal branches showed diverse responses during behaviour. Motor learning significantly increased the proportion of RM boutons and reduced the heterogeneity of bouton activities. Moreover, motor learning induced profound structural dynamism in boutons. By combining structural and functional imaging, we saw that newly formed axonal boutons were more likely to exhibit selectivity for RM and were stabilized during motor learning, whereas UM boutons were selectively eliminated. These findings reveal a novel form of plasticity in corticostriatal axons and show that motor learning drives dynamic bouton reorganization to support motor skill acquisition and execution.

  View details for DOI 10.1038/s41586-025-09336-w

  View details for PubMedID 40739352

  View details for PubMedCentralID 2844762

• Motor learning drives region-specific transcriptomic remodeling in the motor cortex and dorsal striatum. bioRxiv : the preprint server for biology Sun, Y., Roth, R. H., Hwang, F. J., Wang, S., Ding, J. B. 2025

  Abstract

  Motor learning depends on coordinated activity across the motor cortex (M1) and dorsal striatum (dSTR), yet the molecular mechanisms driving learning-related synaptic and circuit remodeling remain unclear. Here, we combine activity-dependent genetic labeling (TRAP) with single-cell RNA sequencing to generate an unbiased, cell type-resolved transcriptional atlas of behaviorally engaged populations during a forelimb reaching task. We identify diverse activated neurons across M1 and dSTR, including a striking enrichment of Htr3a-expressing interneurons (Htr3a INs) in M1 that are selectively recruited during skilled reaching, as confirmed by two-photon calcium imaging. Corticostriatal projection neurons and striatal spiny projection neurons show subtype- and region-specific transcriptional remodeling involving genes linked to synaptic function, translation, and metabolism. Glial cells-including astrocytes, oligodendrocytes, and microglia-exhibit similarly robust, stage- and region-dependent gene regulation. These findings provide a comprehensive molecular framework for motor learning and highlight coordinated, cell type-specific transcriptional programs in neurons and glia that shape the encoding and retrieval of motor memory.

  View details for DOI 10.1101/2025.07.11.664268

  View details for PubMedID 40791319

  View details for PubMedCentralID PMC12338536

• Neuroendocrine circuit for sleep-dependent growth hormone release. Cell Ding, X., Hwang, F. J., Silverman, D., Zhong, P., Li, B., Ma, C., Lu, L., Jiang, G., Zhang, Z., Huang, X., Tu, X., Tian, Z. M., Ding, J., Dan, Y. 2025

  Abstract

  Sleep is known to promote tissue growth and regulate metabolism, partly by enhancing growth hormone (GH) release, but the underlying circuit mechanism is unknown. We demonstrate how GH release, which is enhanced during both rapid eye movement (REM) and non-REM (NREM) sleep, is regulated by sleep-wake-dependent activity of distinct hypothalamic neurons expressing GH-releasing hormone (GHRH) and somatostatin (SST). SST neurons in the arcuate nucleus suppress GH release by inhibiting nearby GHRH neurons that stimulate GH release, whereas periventricular SST neurons inhibit GH release by projecting to the median eminence. GH release is associated with strong surges of both GHRH and SST activity during REM sleep but moderately increased GHRH and decreased SST activity during NREM sleep. Furthermore, we identified a negative feedback pathway in which GH enhances the excitability of locus coeruleus neurons and increases wakefulness. These results elucidate a circuit mechanism underlying bidirectional interactions between sleep and hormone regulation.

  View details for DOI 10.1016/j.cell.2025.05.039

  View details for PubMedID 40562026

• Thalamic integration of basal ganglia and cerebellar circuits during motor learning. bioRxiv : the preprint server for biology Roth, R. H., Muniak, M. A., Huang, C. J., Hwang, F. J., Sun, Y., Min, C., Mao, T., Ding, J. B. 2024

  Abstract

  The ability to control movement and learn new motor skills is one of the fundamental functions of the brain. The basal ganglia (BG) and the cerebellum (CB) are two key brain regions involved in controlling movement, and neuronal plasticity within these two regions is crucial for acquiring new motor skills. However, how these regions interact to produce a cohesive unified motor output remains elusive. Here, we discovered that a subset of neurons in the motor thalamus receive converging synaptic inputs from both BG and CB. By performing multi-site fiber photometry in mice learning motor tasks, we found that motor thalamus neurons integrate BG and CB signals and show distinct movement-related activity. Lastly, we found a critical role of these thalamic neurons and their BG and CB inputs in motor learning and control. These results identify the thalamic convergence of BG and CB and its crucial role in integrating movement signals.

  View details for DOI 10.1101/2024.10.31.621388

  View details for PubMedID 39554076

  View details for PubMedCentralID PMC11565971

• Multiplexed neurochemical sensing with sub-nM sensitivity across 2.25 mm2 area. Biosensors & bioelectronics Mintz Hemed, N., Hwang, F. J., Zhao, E. T., Ding, J. B., Melosh, N. A. 2024; 261: 116474

  Abstract

  Multichannel arrays capable of real-time sensing of neuromodulators in the brain are crucial for gaining insights into new aspects of neural communication. However, measuring neurochemicals, such as dopamine, at low concentrations over large areas has proven challenging. In this research, we demonstrate a novel approach that leverages the scalability and processing power offered by microelectrode array devices integrated with a functionalized, high-density microwire bundle, enabling electrochemical sensing at an unprecedented scale and spatial resolution. The sensors demonstrate outstanding selective molecular recognition by incorporating a selective polymeric membrane. By combining cutting-edge commercial multiplexing, digitization, and data acquisition hardware with a bio-compatible and highly sensitive neurochemical interface array, we establish a powerful platform for neurochemical analysis. This multichannel array has been successfully utilized in vitro and ex vivo systems. Notably, our results show a sensing area of 2.25 mm2 with an impressive detection limit of 820 pM for dopamine. This new approach paves the way for investigating complex neurochemical processes and holds promise for advancing our understanding of brain function and neurological disorders.

  View details for DOI 10.1016/j.bios.2024.116474

  View details for PubMedID 38870827

• A positively tuned voltage indicator for extended electrical recordings in the brain. Nature methods Evans, S. W., Shi, D., Chavarha, M., Plitt, M. H., Taxidis, J., Madruga, B., Fan, J. L., Hwang, F., van Keulen, S. C., Suomivuori, C., Pang, M. M., Su, S., Lee, S., Hao, Y. A., Zhang, G., Jiang, D., Pradhan, L., Roth, R. H., Liu, Y., Dorian, C. C., Reese, A. L., Negrean, A., Losonczy, A., Makinson, C. D., Wang, S., Clandinin, T. R., Dror, R. O., Ding, J. B., Ji, N., Golshani, P., Giocomo, L. M., Bi, G., Lin, M. Z. 2023; 20 (7): 1104-1113

  Abstract

  Genetically encoded voltage indicators (GEVIs) enable optical recording of electrical signals in the brain, providing subthreshold sensitivity and temporal resolution not possible with calcium indicators. However, one- and two-photon voltage imaging over prolonged periods with the same GEVI has not yet been demonstrated. Here, we report engineering of ASAP family GEVIs to enhance photostability by inversion of the fluorescence-voltage relationship. Two of the resulting GEVIs, ASAP4b and ASAP4e, respond to 100-mV depolarizations with ≥180% fluorescence increases, compared with the 50% fluorescence decrease of the parental ASAP3. With standard microscopy equipment, ASAP4e enables single-trial detection of spikes in mice over the course of minutes. Unlike GEVIs previously used for one-photon voltage recordings, ASAP4b and ASAP4e also perform well under two-photon illumination. By imaging voltage and calcium simultaneously, we show that ASAP4b and ASAP4e can identify place cells and detect voltage spikes with better temporal resolution than commonly used calcium indicators. Thus, ASAP4b and ASAP4e extend the capabilities of voltage imaging to standard one- and two-photon microscopes while improving the duration of voltage recordings.

  View details for DOI 10.1038/s41592-023-01913-z

  View details for PubMedID 37429962

• Dichotomous regulation of striatal plasticity by dynorphin. Molecular psychiatry Yang, R., Tuan, R. R., Hwang, F., Bloodgood, D. W., Kong, D., Ding, J. B. 2022

  Abstract

  Modulation of corticostriatal plasticity alters the information flow throughout basal ganglia circuits and represents a fundamental mechanism for motor learning, action selection, and reward. Synaptic plasticity in the striatal direct- and indirect-pathway spiny projection neurons (dSPNs and iSPNs) is regulated by two distinct networks of GPCR signaling cascades. While it is well-known that dopamine D2 and adenosine A2a receptors bi-directionally regulate iSPN plasticity, it remains unclear how D1 signaling modulation of synaptic plasticity is counteracted by dSPN-specific Gi signaling. Here, we show that striatal dynorphin selectively suppresses long-term potentiation (LTP) through Kappa Opioid Receptor (KOR) signaling in dSPNs. Both KOR antagonism and conditional deletion of dynorphin in dSPNs enhance LTP counterbalancing with different levels of D1 receptor activation. Behaviorally, mice lacking dynorphin in D1 neurons show comparable motor behavior and reward-based learning, but enhanced flexibility during reversal learning. These findings support a model in which D1R and KOR signaling bi-directionally modulate synaptic plasticity and behavior in the direct pathway.

  View details for DOI 10.1038/s41380-022-01885-0

  View details for PubMedID 36460726

• Motor learning selectively strengthens cortical and striatal synapses of motor engram neurons. Neuron Hwang, F., Roth, R. H., Wu, Y., Sun, Y., Kwon, D. K., Liu, Y., Ding, J. B. 2022

  Abstract

  Learning and consolidation of new motor skills require plasticity in the motor cortex and striatum, two key motor regions of the brain. However, how neurons undergo synaptic changes and become recruited during motor learning to form a memory engram remains unknown. Here, we train mice on a motor learning task and use a genetic approach to identify and manipulate behavior-relevant neurons selectively in the primary motor cortex (M1). We find that the degree of M1 engram neuron reactivation correlates with motor performance. We further demonstrate that learning-induced dendritic spine reorganization specifically occurs in these M1 engram neurons. In addition, we find that motor learning leads to an increase in the strength of M1 engram neuron outputs onto striatal spiny projection neurons (SPNs) and that these synapses form clusters along SPN dendrites. These results identify a highly specific synaptic plasticity during the formation of long-lasting motor memory traces in the corticostriatal circuit.

  View details for DOI 10.1016/j.neuron.2022.06.006

  View details for PubMedID 35809573

• Phase-Locking Requires Efficient Ca<SUP>2+</SUP> Extrusion at the Auditory Hair Cell Ribbon Synapse JOURNAL OF NEUROSCIENCE Cuadra, A. E., Hwang, F., Burt, L. M., Edmonds, W. C., Chobany, A. V., Li, G. 2021; 41 (8): 1625-1635

  Abstract

  Proper perception of sounds in the environment requires auditory signals to be encoded with extraordinary temporal precision up to tens of microseconds, but how it originates from the hearing organs in the periphery is poorly understood. In particular, sound-evoked spikes in auditory afferent fibers in vivo are phase-locked to sound frequencies up to 5 kHz, but it is not clear how hair cells can handle intracellular Ca2+ changes with such high speed and efficiency. In this study, we combined patch-clamp recording and two-photon Ca2+ imaging to examine Ca2+ dynamics in hair cell ribbon synapses in the bullfrog amphibian papilla of both sexes. We found that Ca2+ clearance from single synaptic ribbons followed a double exponential function, and the weight of the fast component, but not the two time constants, was significantly reduced for prolonged stimulation, and during inhibition of the plasma membrane Ca2+ ATPase (PMCA), the mitochondrial Ca2+ uptake (MCU), or the sarcolemma/endoplasmic reticulum Ca2+ ATPase (SERCA), but not the Na+/Ca2+ exchanger (NCX). Furthermore, we found that both the basal Ca2+ level and the Ca2+ rise during sinusoidal stimulation were significantly increased by inhibition of PMCA, MCU, or SERCA. Consistently, phase-locking of synaptic vesicle releases from hair cells was also significantly reduced by blocking PMCA, MCU, or SERCA, but not NCX. We conclude that, in addition to fast diffusion mediated by mobile Ca2+ buffer, multiple Ca2+ extrusion pumps are required for phase-locking at the auditory hair cell ribbon synapse.SIGNIFICANCE STATEMENT Hair cell synapses can transmit sound-driven signals precisely in the kHz range. However, previous studies of Ca2+ handling in auditory hair cells have often been conducted in immature hair cells, with elevated extracellular Ca2+ concentration, or through steady-state stimulation that may not be physiologically relevant. Here we examine Ca2+ clearance from hair cell synaptic ribbons in a fully mature preparation at physiological concentration of external Ca2+ and at physiological temperature. By stimulating hair cells with sinusoidal voltage commands that mimic pure sound tones, we recapitulated the phase-locking of hair cell exocytosis with an in vitro approach. This allowed us to reveal the Ca2+ extrusion mechanisms that are required for phase-locking at auditory hair cell ribbon synapses.

  View details for DOI 10.1523/JNEUROSCI.1324-18.2020

  View details for Web of Science ID 000621630500002

  View details for PubMedID 33446517

  View details for PubMedCentralID PMC8115884

• Spectrally filtered passive Si photodiode array for on-chip fluorescence imaging of intracellular calcium dynamics SCIENTIFIC REPORTS Xiong, Z., Hwang, F., Sun, F., Xie, Y., Mao, D., Li, G., Xu, G. 2019; 9: 9083

  Abstract

  On-chip fluorescence imaging devices are recognized for their miniaturized and implantable nature that can benefit the study of intracellular dynamics at a variety of settings. However, it is challenging to integrate a spectral filter onto such devices (to block the excitation light) that has similar performance to the state-of-the-art emission filters used in fluorescence microscopes. In this work, we report a 100%-yield, spectrally filtered passive Si photodiode array designed for on-chip fluorescence imaging of intracellular Ca2+ dynamics. Coated with a spectral filter layer that has a high extinction ratio (>103), our array features high wavelength selectivity (>102), high linearity (R2 > 0.98), and low detection limit (45.1 μW 640/30 nm light). Employing fluorescence microscopy as the reference, we demonstrate that our array can conduct on-chip Ca2+ imaging in C2C12 cells that were chemically triggered to increase their intracellular Ca2+ levels. Importantly, our array-level data qualitatively captured the static fluorescence image of the cells and the intracellular Ca2+ dynamics, both of which are correlated with the microscope-collected data. Our results suggest the possible use of the spectrally filtered array towards a miniaturized on-chip fluorescence imaging device, which may open up new opportunities in tissue-level pharmaceutical screening and fundamental studies on cell networks.

  View details for DOI 10.1038/s41598-019-45563-8

  View details for Web of Science ID 000472597100011

  View details for PubMedID 31235791

  View details for PubMedCentralID PMC6591417