Wenbiao Gan, Ph.D.

In vivo studies of microglial function in synaptic plasticity
Gan W.
2011 ;36:S40-S40, Neuropsychopharmacology
Background: The focus of this presentation is to discuss the potential roles of microglia in regulating synaptic development and plasticity in the brain. Microglia are the resident immune cells of the central nervous system and display highly motile processes occupying a non-overlapping territory. Under physiological conditions, microglia may monitor the brain's microenvironment for damage signals and participate in the development and plasticity of neural circuits. Under pathological conditions, microglia undergo a series of morphological and functional changes, and may engage in containing tissue damage, phagocytosis and clearance of cellular debris, and/or the secretion of proinflammatory factors. Although microglia have been implicated in a multitude of physiological and pathological processes in the central nervous system, direct evidence of their roles in synaptic structure and functions remains elusive. Methods: Hampering efforts to delineate the role of microglia is the lack of tools to specifically perturb microglial function in vivo. To overcome this difficulty, we have recently generated mice with a targeted gene insertion allowing for the expression of tamoxifeninducible Cre recombinase in CX₃CR1 expressing microglial cells. By crossing CX₃CR1-CreER mice with mice harboring floxed alleles of the diphtheria toxin receptor (iDTR) under the control of the ubiquitous Rosa26 promoter, we have been able to specifically and efficiently ablate microglia in an inducible fashion. By ablating microglial cells and perturbing their functions in the living mice, we hope to elucidate the role of microglia in synapse development and learning-dependent synaptic plasticity. Results: Our preliminary results suggest that deletion of CX₃CR1 expressing microglial cells may cause a decrease in the turnover of postsynaptic dendritic spines in the living mouse cortex. Conclusions: Our findings indicate that the CX₃CR1-CreER mouse line provide a molecular handle for the in vivo manipulation of microglia including deletion and support an important role of microglia in synapse development and plasticity
— id: 147763, year: 2011, vol: 36, page: S40, stat: Journal Article,

Transcranial two-photon imaging of the living mouse brain
Grutzendler, Jaime; Yang, Guang; Pan, Feng; Parkhurst, Christopher N; Gan, Wen-Biao
2011 ;2011(9):?-?, Cold Spring Harbor protocols
INTRODUCTION This protocol describes imaging of the living mouse brain through a thinned skull using two-photon microscopy. This transcranial two-photon laser-scanning microscope (TPLSM) imaging method allows high-resolution imaging of fluorescently labeled neurons, microglia, astrocytes, and blood vessels, as well as subcellular structures such as dendritic spines and axonal varicosities. The surgical procedure that is required to allow imaging thins the cranium so that it becomes optically transparent. Once learned, the surgery can be performed in approximately 30 min, and imaging can follow immediately. The procedure can be repeated multiple times, allowing brain cells and tissues to be studied in the same animals over short or long time intervals, depending on the design of the experiment. Two-photon imaging through a thinned and intact skull avoids side effects caused by skull removal and is a minimally invasive method for studying the living mouse brain under physiological and pathological conditions
— id: 137133, year: 2011, vol: 2011, page: ?, stat: Journal Article,

Glucocorticoids are critical regulators of dendritic spine development and plasticity in vivo
Liston, Conor; Gan, Wen-Biao
2011 Sep 20;108(38):16074-16079, Proceedings of the National Academy of Sciences of the United States of America
Glucocorticoids are a family of hormones that coordinate diverse physiological processes in responding to stress. Prolonged glucocorticoid exposure over weeks has been linked to dendritic atrophy and spine loss in fixed tissue studies of adult brains, but it is unclear how glucocorticoids may affect the dynamic processes of dendritic spine formation and elimination in vivo. Furthermore, relatively few studies have examined the effects of stress and glucocorticoids on spines during the postnatal and adolescent period, which is characterized by rapid synaptogenesis followed by protracted synaptic pruning. To determine whether and to what extent glucocorticoids regulate dendritic spine development and plasticity, we used transcranial two-photon microscopy to track the formation and elimination of dendritic spines in vivo after treatment with glucocorticoids in developing and adult mice. Corticosterone, the principal murine glucocorticoid, had potent dose-dependent effects on dendritic spine dynamics, increasing spine turnover within several hours in the developing barrel cortex. The adult barrel cortex exhibited diminished baseline spine turnover rates, but these rates were also enhanced by corticosterone. Similar changes occurred in multiple cortical areas, suggesting a generalized effect. However, reducing endogenous glucocorticoid activity by dexamethasone suppression or corticosteroid receptor antagonists caused a substantial reduction in spine turnover rates, and the former was reversed by corticosterone replacement. Notably, we found that chronic glucocorticoid excess led to an abnormal loss of stable spines that were established early in life. Together, these findings establish a critical role for glucocorticoids in the development and maintenance of dendritic spines in the living cortex
— id: 137844, year: 2011, vol: 108, page: 16074, stat: Journal Article,

A Multisite Study of the Clinical Diagnosis of Different Autism Spectrum Disorders
Lord C; Petkova E; Hus V; Gan W; Lu F; Martin DM; Ousley O; Guy L; Bernier R; Gerdts J; Algermissen M; Whitaker A; Sutcliffe JS; Warren Z; Klin A; Saulnier C; Hanson E; Hundley R; Piggot J; Fombonne E; Steiman M; Miles J; Kanne SM; Goin-Kochel RP; Peters SU; Cook EH; Guter S; Tjernagel J; Green-Snyder LA; Bishop S; Esler A; Gotham K; Luyster R; Miller F; Olson J; Richler J; Risi S
2011 Nov 7;:?-? #, Archives of general psychiatry
CONTEXT: Best-estimate clinical diagnoses of specific autism spectrum disorders (autistic disorder, pervasive developmental disorder-not otherwise specified, and Asperger syndrome) have been used as the diagnostic gold standard, even when information from standardized instruments is available. OBJECTIVE: To determine whether the relationships between behavioral phenotypes and clinical diagnoses of different autism spectrum disorders vary across 12 university-based sites. DESIGN: Multisite observational study collecting clinical phenotype data (diagnostic, developmental, and demographic) for genetic research. Classification trees were used to identify characteristics that predicted diagnosis across and within sites. SETTING: Participants were recruited through 12 university-based autism service providers into a genetic study of autism. PARTICIPANTS: A total of 2102 probands (1814 male probands) between 4 and 18 years of age (mean [SD] age, 8.93 [3.5] years) who met autism spectrum criteria on the Autism Diagnostic Interview-Revised and the Autism Diagnostic Observation Schedule and who had a clinical diagnosis of an autism spectrum disorder. Main Outcome Measure Best-estimate clinical diagnoses predicted by standardized scores from diagnostic, cognitive, and behavioral measures. RESULTS: Although distributions of scores on standardized measures were similar across sites, significant site differences emerged in best-estimate clinical diagnoses of specific autism spectrum disorders. Relationships between clinical diagnoses and standardized scores, particularly verbal IQ, language level, and core diagnostic features, varied across sites in weighting of information and cutoffs. CONCLUSIONS: Clinical distinctions among categorical diagnostic subtypes of autism spectrum disorders were not reliable even across sites with well-documented fidelity using standardized diagnostic instruments. Results support the move from existing subgroupings of autism spectrum disorders to dimensional descriptions of core features of social affect and fixated, repetitive behaviors, together with characteristics such as language level and cognitive function
— id: 142976, year: 2011, vol: , page: ?, stat: Journal Article,

Transient Effects of Anesthetics on Dendritic Spines and Filopodia in the Living Mouse Cortex
Yang G; Chang PC; Bekker A; Blanck TJ; Gan WB
2011 Oct;115(4):718-726, Anesthesiology
BACKGROUND:: Anesthetics are widely used to induce unconsciousness, pain relief, and immobility during surgery. It remains unclear whether the use of anesthetics has significant and long-lasting effects on synapse development and plasticity in the brain. To address this question, the authors examined the formation and elimination of dendritic spines, postsynaptic sites of excitatory synapses, in the developing mouse cortex during and after anesthetics exposure. METHODS:: Transgenic mice expressing yellow fluorescence protein in layer 5 pyramidal neurons were used in this study. Mice at 1 month of age underwent ketamine-xylazine and isoflurane anesthesia over a period of hours. The elimination and formation rates of dendritic spines and filopodia, the precursors of spines, were followed over hours to days in the primary somatosensory cortex using transcranial two-photon microscopy. Four to five animals were examined under each experimental condition. Student t test and Mann-Whitney U test were used to analyze the data. RESULTS:: Administration of either ketamine-xylazine or isoflurane rapidly altered dendritic filopodial dynamics but had no significant effects on spine dynamics. Ketamine-xylazine increased filopodial formation whereas isoflurane decreased filopodial elimination during 4 h of anesthesia. Both effects were transient and disappeared within a day after the animals woke up. CONCLUSION:: Studies suggest that exposure to anesthetics transiently affects the dynamics of dendritic filopodia but has no significant effect on dendritic spine development and plasticity in the cortex of 1-month-old mice
— id: 137134, year: 2011, vol: 115, page: 718, stat: Journal Article,

In vivo imaging neuronal recovery in neuroinflammation
Gan Wanbiao; Christopher, Parkhurst; Scott, Hayes; Gan Wen-Biao
2010 NOV 15 ;228(1-2):145-145, Journal of neuroimmunology
— id: 120558, year: 2010, vol: 228, page: 145, stat: Journal Article,

Dendritic spine instability and insensitivity to modulation by sensory experience in a mouse model of fragile X syndrome
Pan, Feng; Aldridge, Georgina M; Greenough, William T; Gan, Wen-Biao
2010 Oct 12;107(41):17768-17773, Proceedings of the National Academy of Sciences of the United States of America
Fragile X syndrome (FXS) is the most common inherited form of mental retardation and is caused by transcriptional inactivation of the X-linked fragile X mental retardation 1 (FMR1) gene. FXS is associated with increased density and abnormal morphology of dendritic spines, the postsynaptic sites of the majority of excitatory synapses. To better understand how lack of the FMR1 gene function affects spine development and plasticity, we examined spine formation and elimination of layer 5 pyramidal neurons in the whisker barrel cortex of Fmr1 KO mice with a transcranial two-photon imaging technique. We found that the rates of spine formation and elimination over days to weeks were significantly higher in both young and adult KO mice compared with littermate controls. The heightened spine turnover in KO mice was due to the existence of a larger pool of 'short-lived' new spines in KO mice than in controls. Furthermore, we found that the formation of new spines and the elimination of existing ones were less sensitive to modulation by sensory experience in KO mice. These results indicate that the loss of Fmr1 gene function leads to ongoing overproduction of transient spines in the primary somatosensory cortex. The insensitivity of spine formation and elimination to sensory alterations in Fmr1 KO mice suggest that the developing synaptic circuits may not be properly tuned by sensory stimuli in FXS
— id: 113804, year: 2010, vol: 107, page: 17768, stat: Journal Article,

Microglia dynamics and function in the CNS
Parkhurst, Christopher N; Gan, Wen-Biao
2010 Oct;20(5):595-600, Current opinion in neurobiology
Microglial cells constitute the resident immune cell population of the mammalian central nervous system. One striking feature of these cells is their highly dynamic nature under both normal and pathological brain conditions. The highly branched processes of resting microglia display a constitutive mobility and undergo rapid directional movement towards sites of acute tissue disruption. Microglia can be converted by a large number of different stimuli to a chronically activated state by signaling through both purinergic and Toll-like receptor systems, among others. Recent work has uncovered some of the mechanisms underlying microglia dynamics and shed new light into the functional significance of this enigmatic member of the glial cell family
— id: 138196, year: 2010, vol: 20, page: 595, stat: Journal Article,

Thinned-skull cranial window technique for long-term imaging of the cortex in live mice
Yang, Guang; Pan, Feng; Parkhurst, Christopher N; Grutzendler, Jaime; Gan, Wen-Biao
2010 Jan;5(2):201-208, Nature Protocols
Imaging neurons, glia and vasculature in the living brain has become an important experimental tool for understanding how the brain works. Here we describe in detail a protocol for imaging cortical structures at high optical resolution through a thinned-skull cranial window in live mice using two-photon laser scanning microscopy (TPLSM). Surgery can be performed within 30-45 min and images can be acquired immediately thereafter. The procedure can be repeated multiple times allowing longitudinal imaging of the cortex over intervals ranging from days to years. Imaging through a thinned-skull cranial window avoids exposure of the meninges and the cortex, thus providing a minimally invasive approach for studying structural and functional changes of cells under normal and pathological conditions in the living brain
— id: 106596, year: 2010, vol: 5, page: 201, stat: Journal Article,

Dendritic spine dynamics
Bhatt, D Harshad; Zhang, Shengxiang; Gan, Wen-Biao
2009 ;71:261-282, Annual review of physiology
Dendritic spines are the postsynaptic components of most excitatory synapses in the mammalian brain. Spines accumulate rapidly during early postnatal development and undergo a substantial loss as animals mature into adulthood. In past decades, studies have revealed that the number and size of dendritic spines are regulated by a variety of gene products and environmental factors, underscoring the dynamic nature of spines and their importance to brain plasticity. Recently, in vivo time-lapse imaging of dendritic spines in the cerebral cortex suggests that, although spines are highly plastic during development, they are remarkably stable in adulthood, and most of them last throughout life. Therefore, dendritic spines may provide a structural basis for lifelong information storage, in addition to their well-established role in brain plasticity. Because dendritic spines are the key elements for information acquisition and retention, understanding how spines are formed and maintained, particularly in the intact brain, will likely provide fundamental insights into how the brain possesses the extraordinary capacity to learn and to remember
— id: 100626, year: 2009, vol: 71, page: 261, stat: Journal Article,

Experience-dependent dendritic spine dynamics in the mouse cortex
Gan, WB; Yang, G; Pan, F
2009 FEB ;65(1):S7-S7, Neuroscience research
— id: 106960, year: 2009, vol: 65, page: S7, stat: Journal Article,

Ballistic delivery of dyes for structural and functional studies of the nervous system
Gan, Wen-Biao; Grutzendler, Jaime; Wong, Rachel O; Lichtman, Jeff W
2009 Apr;2009(4):pdb.prot5202-pdb.prot5202, Cold Spring Harbor protocols
This protocol describes detailed procedures for rapid labeling of cells in a variety of preparations by means of particle-mediated ballistic (i.e., Gene Gun) delivery of fluorescent dyes. The method has been used for rapid labeling of cells with either lipid- or water-soluble dyes, in a variety of preparations at different ages. Tissue preparations include fixed mouse brain slices (described here), cell cultures, and tissue explants. This ballistic labeling technique is useful for studying neuronal connectivity, function, and pathology in the nervous system of living as well as fixed specimens
— id: 112400, year: 2009, vol: 2009, page: pdb.prot5202, stat: Journal Article,

Ankyrin Repeat-rich Membrane Spanning/Kidins220 protein regulates dendritic branching and spine stability in vivo
Wu, Synphen H; Arevalo, Juan Carlos; Sarti, Federica; Tessarollo, Lino; Gan, Wen-Biao; Chao, Moses V
2009 Aug;69(9):547-557, Developmental Neurobiology
The development of nervous system connectivity depends upon the arborization of dendritic fields and the stabilization of dendritic spine synapses. It is well established that neuronal activity and the neurotrophin BDNF modulate these correlated processes. However, the downstream mechanisms by which these extrinsic signals regulate dendritic development and spine stabilization are less well known. Here we report that a substrate of BDNF signaling, the Ankyrin Repeat-rich Membrane Spanning (ARMS) protein or Kidins220, plays a critical role in the branching of cortical and hippocampal dendrites and in the turnover of cortical spines. In the barrel somatosensory cortex and the dentate gyrus, regions where ARMS/Kidins220 is highly expressed, no difference in the complexity of dendritic arbors was observed in 1-month-old adolescent ARMS/Kidins220(+/-) mice compared to wild-type littermates. However, at 3 months of age, young adult ARMS/Kidins220(+/-) mice exhibited decreased dendritic complexity. This suggests that ARMS/Kidins220 does not play a significant role in the initial formation of dendrites but, rather, is involved in the refinement or stabilization of the arbors later in development. In addition, at 1 month of age, the rate of spine elimination was higher in ARMS/Kidins220(+/-) mice than in wild-type mice, suggesting that ARMS/Kidins220(+/-) levels regulate spine stability. Taken together, these data suggest that ARMS/Kidins220 is important for the growth of dendritic arbors and spine stability during an activity- and BDNF-dependent period of development. (c) 2009 Wiley Periodicals, Inc. Develop Neurobiol 2009
— id: 100607, year: 2009, vol: 69, page: 547, stat: Journal Article,

Stably maintained dendritic spines are associated with lifelong memories
Yang, Guang; Pan, Feng; Gan, Wen-Biao
2009 Dec 17;462(7275):920-924, Nature
Changes in synaptic connections are considered essential for learning and memory formation. However, it is unknown how neural circuits undergo continuous synaptic changes during learning while maintaining lifelong memories. Here we show, by following postsynaptic dendritic spines over time in the mouse cortex, that learning and novel sensory experience lead to spine formation and elimination by a protracted process. The extent of spine remodelling correlates with behavioural improvement after learning, suggesting a crucial role of synaptic structural plasticity in memory formation. Importantly, a small fraction of new spines induced by novel experience, together with most spines formed early during development and surviving experience-dependent elimination, are preserved and provide a structural basis for memory retention throughout the entire life of an animal. These studies indicate that learning and daily sensory experience leave minute but permanent marks on cortical connections and suggest that lifelong memories are stored in largely stably connected synaptic networks
— id: 105966, year: 2009, vol: 462, page: 920, stat: Journal Article,

Various dendritic abnormalities are associated with fibrillar amyloid deposits in Alzheimer's disease
Grutzendler, Jaime; Helmin, Kathryn; Tsai, Julia; Gan, Wen-Biao
2007 Feb;1097:30-39, Annals of the New York Academy of Sciences
Dystrophic neurites are associated with fibrillar amyloid deposition in Alzheimer's disease (AD), but the frequency and types of changes in synaptic structures near amyloid deposits have not been well characterized. Using high-resolution confocal microscopy to image lipophilic dye-labeled dendrites and thioflavin-S-labeled amyloid plaques, we systematically analyzed the structural changes of dendrites associated with amyloid deposition in both a transgenic mouse model of AD (PSAPP) and in human postmortem brain. We found that in PSAPP mice, dendritic branches passing through or within 40 mum from amyloid deposits displayed various dendritic abnormalities such as loss of dendritic spines, shaft atrophy, bending, abrupt branch endings, varicosity formation, and sprouting. Similar structural alterations of dendrites were seen in postmortem human AD tissue, with spine loss as the most common abnormality in both PSAPP mice and human AD brains. These results demonstrate that fibrillar amyloid deposits and their surrounding microenvironment are toxic to dendrites and likely contribute to significant disruption of neuronal circuits in AD
— id: 71869, year: 2007, vol: 1097, page: 30, stat: Journal Article,

Various dendritic abnormalities are associated with fibrillar amyloid deposits in Alzheimer's disease
Grutzendler, Jaime; Helmin, Kathryn; Tsai, Julia; Gan, Wen-Biao
Imaging and the aging brain Malden, MA, US: Blackwell Publishing, 2007,
(from the chapter) Dystrophic neurites are associated with fibrillar amyloid deposition in Alzheimer's disease (AD), but the frequency and types of changes in synaptic structures near amyloid deposits have not been well characterized. Using high-resolution confocal microscopy to image lipophilic dye-labeled dendrites and thioflavin-S-labeled amyloid plaques, we systematically analyzed the structural changes of dendrites associated with amyloid deposition in both a transgenic mouse model of AD (PSAPP) and in human postmortem brain. We found that in PSAPP mice, dendritic branches passing through or within 40 p.m. from amyloid deposits displayed various dendritic abnormalities such as loss of dendritic spines, shaft atrophy, bending, abrupt branch endings, varicosity formation, and sprouting. Similar structural alterations of dendrites were seen in postmortem human AD tissue, with spine loss as the most common abnormality in both PSAPP mice and human AD brains. These results demonstrate that fibrillar amyloid deposits and their surrounding microenvironment are toxic to dendrites and likely contribute to significant disruption of neuronal circuits in AD.
— id: 4483, year: 2007, vol: , page: 30, stat: Chapter,

Choice of cranial window type for in vivo imaging affects dendritic spine turnover in the cortex
Xu, Hua-Tai; Pan, Feng; Yang, Guang; Gan, Wen-Biao
2007 May;10(5):549-551, Nature neuroscience
Determining the degree of synapse formation and elimination is essential for understanding the structural basis of brain plasticity and pathology. We show that in vivo imaging of dendritic spine dynamics through an open-skull glass window, but not a thinned-skull window, is associated with high spine turnover and substantial glial activation during the first month after surgery. These findings help to explain existing discrepancies in the degree of dendritic spine plasticity observed in the mature cortex.
— id: 73017, year: 2007, vol: 10, page: 549, stat: Journal Article,

Recent advances in basic neurosciences and brain disease: from synapses to behavior
Bi, Guo-Qiang; Bolshakov, Vadim; Bu, Guojun; Cahill, Catherine M; Chen, Zhou-Feng; Collingridge, Graham L; Cooper, Robin L; Coorssen, Jens R; El-Husseini, Alaa; Galhardo, Vasco; Gan, Wen-Biao; Gu, Jianguo; Inoue, Kazuhide; Isaac, John; Iwata, Koichi; Jia, Zhengping; Kaang, Bong-Kiun; Kawamata, Mikito; Kida, Satoshi; Klann, Eric; Kohno, Tatsuro; Li, Min; Li, Xiao-Jiang; MacDonald, John F; Nader, Karim; Nguyen, Peter V; Oh, Uhtaek; Ren, Ke; Roder, John C; Salter, Michael W; Song, Weihong; Sugita, Shuzo; Tang, Shao-Jun; Tao, Yuanxiang; Wang, Yu Tian; Woo, Newton; Woodin, Melanie A; Yan, Zhen; Yoshimura, Megumu; Xu, Ming; Xu, Zao C; Zhang, Xia; Zhen, Mei; Zhuo, Min
2006 ;2:38-38, Molecular pain
Understanding basic neuronal mechanisms hold the hope for future treatment of brain disease. The 1st international conference on synapse, memory, drug addiction and pain was held in beautiful downtown Toronto, Canada on August 21-23, 2006. Unlike other traditional conferences, this new meeting focused on three major aims: (1) to promote new and cutting edge research in neuroscience; (2) to encourage international information exchange and scientific collaborations; and (3) to provide a platform for active scientists to discuss new findings. Up to 64 investigators presented their recent discoveries, from basic synaptic mechanisms to genes related to human brain disease. This meeting was in part sponsored by Molecular Pain, together with University of Toronto (Faculty of Medicine, Department of Physiology as well as Center for the Study of Pain). Our goal for this meeting is to promote future active scientific collaborations and improve human health through fundamental basic neuroscience researches. The second international meeting on Neurons and Brain Disease will be held in Toronto (August 29-31, 2007)
— id: 112401, year: 2006, vol: 2, page: 38, stat: Journal Article,

Two-photon imaging of synaptic plasticity and pathology in the living mouse brain
Grutzendler, Jaime; Gan, Wen-Biao
2006 Oct;3(4):489-496, NeuroRx
Two-photon microscopy (TPM) has become an increasingly important tool for imaging the structure and function of brain cells in living animals. TPM imaging studies of neuronal structures over intervals ranging from seconds to years have begun to provide important insights into the structural plasticity of synapses and the modulating effects of experience in the intact brain. TPM has also started to reveal how neuronal connections are altered in animal models of neurodegeneration, acute brain injury, and cerebrovascular disease. Here, we review some of these studies with special emphasis on the degree of structural dynamism of postsynaptic dendritic spines in the adult mouse brain as well as synaptic pathology in mouse models of Alzheimer's disease and cerebral ischemia. We also discuss technical considerations that are critical for the acquisition and interpretation of data from TPM in vivo
— id: 112403, year: 2006, vol: 3, page: 489, stat: Journal Article,

The P2Y12 receptor regulates microglial activation by extracellular nucleotides
Haynes, Sharon E; Hollopeter, Gunther; Yang, Guang; Kurpius, Dana; Dailey, Michael E; Gan, Wen-Biao; Julius, David
2006 Dec;9(12):1512-1519, Nature neuroscience
Microglia are primary immune sentinels of the CNS. Following injury, these cells migrate or extend processes toward sites of tissue damage. CNS injury is accompanied by release of nucleotides, serving as signals for microglial activation or chemotaxis. Microglia express several purinoceptors, including a G(i)-coupled subtype that has been implicated in ATP- and ADP-mediated migration in vitro. Here we show that microglia from mice lacking G(i)-coupled P2Y(12) receptors exhibit normal baseline motility but are unable to polarize, migrate or extend processes toward nucleotides in vitro or in vivo. Microglia in P2ry(12)(-/-) mice show significantly diminished directional branch extension toward sites of cortical damage in the living mouse. Moreover, P2Y(12) expression is robust in the 'resting' state, but dramatically reduced after microglial activation. These results imply that P2Y(12) is a primary site at which nucleotides act to induce microglial chemotaxis at early stages of the response to local CNS injury
— id: 112402, year: 2006, vol: 9, page: 1512, stat: Journal Article,

ATP mediates rapid microglial response to local brain injury in vivo
Davalos, Dimitrios; Grutzendler, Jaime; Yang, Guang; Kim, Jiyun V; Zuo, Yi; Jung, Steffen; Littman, Dan R; Dustin, Michael L; Gan, Wen-Biao
2005 Jun;8(6):752-758, Nature neuroscience
Parenchymal microglia are the principal immune cells of the brain. Time-lapse two-photon imaging of GFP-labeled microglia demonstrates that the fine termini of microglial processes are highly dynamic in the intact mouse cortex. Upon traumatic brain injury, microglial processes rapidly and autonomously converge on the site of injury without cell body movement, establishing a potential barrier between the healthy and injured tissue. This rapid chemotactic response can be mimicked by local injection of ATP and can be inhibited by the ATP-hydrolyzing enzyme apyrase or by blockers of G protein-coupled purinergic receptors and connexin channels, which are highly expressed in astrocytes. The baseline motility of microglial processes is also reduced significantly in the presence of apyrase and connexin channel inhibitors. Thus, extracellular ATP regulates microglial branch dynamics in the intact brain, and its release from the damaged tissue and surrounding astrocytes mediates a rapid microglial response towards injury
— id: 56024, year: 2005, vol: 8, page: 752, stat: Journal Article,

Defective neuromuscular synapses in mice lacking amyloid precursor protein (APP) and APP-Like protein 2
Wang, Pei; Yang, Guang; Mosier, Dennis R; Chang, Paul; Zaidi, Tahire; Gong, Yan-Dao; Zhao, Nan-Ming; Dominguez, Bertha; Lee, Kuo-Fen; Gan, Wen-Biao; Zheng, Hui
2005 Feb 2;25(5):1219-1225, Journal of neuroscience
Biochemical and genetic studies place the amyloid precursor protein (APP) at the center stage of Alzheimer's disease (AD) pathogenesis. Although mutations in the APP gene lead to dominant inheritance of familial AD, the normal function of APP remains elusive. Here, we report that the APP family of proteins plays an essential role in the development of neuromuscular synapses. Mice deficient in APP and its homolog APP-like protein 2 (APLP2) exhibit aberrant apposition of presynaptic marker proteins with postsynaptic acetylcholine receptors and excessive nerve terminal sprouting. The number of synaptic vesicles at presynaptic terminals is dramatically reduced. These structural abnormalities are accompanied by defective neurotransmitter release and a high incidence of synaptic failure. Our results identify APP/APLP2 as key regulators of structure and function of developing neuromuscular synapses
— id: 112406, year: 2005, vol: 25, page: 1219, stat: Journal Article,

Reduced synaptic vesicle density and active zone size in mice lacking amyloid precursor protein (APP) and APP-like protein 2
Yang, Guang; Gong, Yan-Dao; Gong, Kai; Jiang, Wu-Ling; Kwon, Elaine; Wang, Pei; Zheng, Hui; Zhang, Xiu-Fang; Gan, Wen-Biao; Zhao, Nan-Ming
2005 Aug 12-19;384(1-2):66-71, Neuroscience letters
Although abnormal processing of amyloid precursor protein (APP) leads to early onset of Alzheimer's disease, the normal function of this protein is poorly understood. APP is widely expressed in axons, dendrites, and synapses in both central and peripheral nervous systems. Mice homozygous for APP or its homologue APP-like protein 2 (APLP2) null mutation (KO) are viable, but double mutants for APP and APLP2 deletions (DKO) are early postnatal lethal. To investigate the role of APP in synapse development, we compared the ultrastructure of submandibular ganglion synapses between DKO and littermate APLP2 KO mice at birth. Using serial electron microscopy, we found that the size of presynaptic boutons and the number of active zones per bouton were comparable in both strains of animals. However, the synaptic vesicle density, active zone size, and docked vesicle number per active zone were significantly reduced in DKO compared to those in APLP2 KO. These results indicate that the APP family of proteins plays an important role in regulating the formation and function of inter-neuronal synapses
— id: 112405, year: 2005, vol: 384, page: 66, stat: Journal Article,

A model of mini-embolic stroke offers measurements of the neurovascular unit response in the living mouse
Zhang, Zheng Gang; Zhang, Li; Ding, Guangliang; Jiang, Quan; Zhang, Rui Lan; Zhang, Xueguo; Gan, Wen-Biao; Chopp, Michael
2005 Dec;36(12):2701-2704, Stroke
BACKGROUND AND PURPOSE: To measure cerebral vascular and neuronal responses after stroke in the living mouse, we generated a mouse model of embolic stroke localized to the parietal cortex. METHODS: Male C57/6J or male transgenic mice (2 to 3 months old) expressing yellow fluorescent protein (YFP) were used in the present study. A single fibrin-rich clot (8 mm in length) was injected into a branch of the right middle cerebral artery (MCA). MRI measurements were performed to measure ischemic lesion. Using confocal and 2-photon microscopy, changes in the embolus, dendrites, and dendritic spines were measured in the living mouse. RESULTS: Eight of 11 mice (73%) had the embolus localized to a branch of the right MCA in the parietal cortex. Expansion of the embolus within the artery was observed 24 hours after stroke. The presence of ischemic lesion in the parietal cortex was verified by MRI measurements, and histopathological analysis revealed that these mice (n=8) had a cortical infarct volume of 4.9+/-3.6% of the contralateral hemisphere. In the living mouse, substantial loss of YFP-labeled axonal and dendritic structures as well as the formation of abnormal dendritic bulbs were detected in the ischemic boundary regions 24 hours after stroke compared with that 1 hour after stroke. CONCLUSIONS: This model offers a novel approach to study the neurovascular unit in cerebral cortex after stroke in the living mouse
— id: 112404, year: 2005, vol: 36, page: 2701, stat: Journal Article,

Development of long-term dendritic spine stability in diverse regions of cerebral cortex
Zuo, Yi; Lin, Aerie; Chang, Paul; Gan, Wen-Biao
2005 Apr 21;46(2):181-189, Neuron
Synapse formation and elimination occur throughout life, but the magnitude of such changes at distinct developmental stages remains unclear. Using transgenic mice overexpressing yellow fluorescent protein and transcranial two-photon microscopy, we repeatedly imaged dendritic spines on the apical dendrites of layer 5 pyramidal neurons. In young adolescent mice (1-month-old), 13%-20% of spines were eliminated and 5%-8% formed over 2 weeks in barrel, motor, and frontal cortices, indicating a cortical-wide spine loss during this developmental period. As animals mature, there is also a substantial loss of dendritic filopodia involved in spinogenesis. In adult mice (4-6 months old), 3%-5% of spines were eliminated and formed over 2 weeks in various cortical regions. Over 18 months, only 26% of spines were eliminated and 19% formed in adult barrel cortex. Thus, after a concurrent loss of spines and spine precursors in diverse regions of young adolescent cortex, spines become stable and a majority of them can last throughout life
— id: 55610, year: 2005, vol: 46, page: 181, stat: Journal Article,

Long-term sensory deprivation prevents dendritic spine loss in primary somatosensory cortex
Zuo, Yi; Yang, Guang; Kwon, Elaine; Gan, Wen-Biao
2005 Jul 14;436(7048):261-265, Nature
A substantial decrease in the number of synapses occurs in the mammalian brain from the late postnatal period until the end of life. Although experience plays an important role in modifying synaptic connectivity, its effect on this nearly lifelong synapse loss remains unknown. Here we used transcranial two-photon microscopy to visualize postsynaptic dendritic spines in layer I of the barrel cortex in transgenic mice expressing yellow fluorescent protein. We show that in young adolescent mice, long-term sensory deprivation through whisker trimming prevents net spine loss by preferentially reducing the rate of ongoing spine elimination, not by increasing the rate of spine formation. This effect of deprivation diminishes as animals mature but still persists in adulthood. Restoring sensory experience after adolescent deprivation accelerates spine elimination. Similar to sensory manipulation, the rate of spine elimination decreases after chronic blockade of NMDA (N-methyl-D-aspartate) receptors with the antagonist MK801, and accelerates after drug withdrawal. These studies of spine dynamics in the primary somatosensory cortex suggest that experience plays an important role in the net loss of synapses over most of an animal's lifespan, particularly during adolescence
— id: 56375, year: 2005, vol: 436, page: 261, stat: Journal Article,

Axon branch removal at developing synapses by axosome shedding
Bishop, Derron L; Misgeld, Thomas; Walsh, Mark K; Gan, Wen-Biao; Lichtman, Jeff W
2004 Nov 18;44(4):651-661, Neuron
In many parts of the developing nervous system, the number of axonal inputs to each postsynaptic cell is dramatically reduced. This synapse elimination has been extensively studied at the neuromuscular junction, but how axons are lost is unknown. Here, we combine time-lapse imaging of fluorescently labeled axons and serial electron microscopy to show that axons at neuromuscular junctions are removed by an unusual cellular mechanism. As axons disappear, they shed numerous membrane bound remnants. These 'axosomes' contain a high density of synaptic organelles and are formed by engulfment of axon tips by Schwann cells. After this engulfment, the axosome's contents mix with the cytoplasm of the glial cell. Axosome shedding might underlie other forms of axon loss and may provide a pathway for interactions between axons and glia
— id: 57697, year: 2004, vol: 44, page: 651, stat: Journal Article,

Age-associated synapse elimination in mouse parasympathetic ganglia
Coggan, Jay S; Grutzendler, Jaime; Bishop, Derron L; Cook, Melissa R; Gan, Wenbiao; Heym, Jason; Lichtman, Jeff W
2004 Aug;60(2):214-226, Journal of neurobiology
Little is known about the effects of aging on synapses in the mammalian nervous system. We examined the innervation of individual mouse submandibular ganglion (SMG) neurons for evidence of age-related changes in synapse efficacy and number. For approximately 85% of adult life expectancy (30 months) the efficacy of synaptic transmission, as determined by excitatory postsynaptic potential (EPSP) amplitudes, remains constant. Similarly, the number of synapses contacting individual SMG neurons is also unchanged. After 30 months of age, however, some neurons (23%) dramatically lose synaptic input exhibiting both smaller EPSP amplitude and fewer synaptic boutons. Attenuation of both the amplitude and frequency of miniature EPSPs was also observed in neurons from aged animals. Electron micrographs revealed that, although there were many vesicle-laden preganglionic axonal processes in the vicinity of the postsynaptic membrane, the number of synaptic contacts was significantly lower in old animals. These results demonstrate primary, age-associated synapse elimination with functional consequences that cannot be explained by pre- or postsynaptic cell death
— id: 112399, year: 2004, vol: 60, page: 214, stat: Journal Article,

Solid-phase synthesis of styryl dyes and their application as amyloid sensors
Li, Qian; Lee, Jun-Seok; Ha, Chanki; Park, Chan Beum; Yang, Guang; Gan, Wen Biao; Chang, Young-Tae
2004 Nov 26;43(46):6331-6335, Angewandte Chemie. International edition in English
— id: 112407, year: 2004, vol: 43, page: 6331, stat: Journal Article,

In vivo imaging of prion amyloid deposits
Sadowski, M; Pankiewicz, J; Scholtzova, H; Tsai, J; Carp, RI; Meeker, HC; Debnath, M; Mathis, CA; Shao, L; Klunk, WE; Gan, WB; Wisniewski, T
2004 JUL ;25(10):S280-S281, Neurobiology of aging
— id: 47732, year: 2004, vol: 25, page: S280, stat: Journal Article,

Detection of prion amyloid deposits in vivo
Sadowski, Marcin; Pankiewicz, Joanna; Scholtzova, Henrieta; Tsai, Julia; Carp, Richard I.; Meeker, Cliff H.; Gan, Wen-Biao; Klunk, William E.; Mathis, Chester A.; Shao, Li; Debnath, Manik; Wisniewski, Thomas
2004 ;62(7, Suppl. 5):A446-A447, Neurology
— id: 97609, year: 2004, vol: 62, page: A446, stat: Journal Article,

Targeting prion amyloid deposits in vivo
Sadowski, Marcin; Pankiewicz, Joanna; Scholtzova, Henrieta; Tsai, Julia; Li, Yongsheng; Carp, Richard I; Meeker, Harry C; Gambetti, Pierluigi; Debnath, Manik; Mathis, Chester A; Shao, Li; Gan, Wen-Biao; Klunk, William E; Wisniewski, Thomas
2004 Jul;63(7):775-784, Journal of neuropathology & experimental neurology
The diagnosis of prion diseases in humans is challenging due to a lack of specific and sensitive non-invasive tests. Many forms of human prion disease including variant Creutzfeldt-Jakob disease (vCJD), Gerstmann-Straussler-Scheinker (GSS) syndrome, and 10% of sporadic CJD cases are associated with amyloid deposition. Several positron emission tomography (PET) ligands have recently been developed to directly image beta-amyloid associated with Alzheimer disease. One of them, methoxy-X04, is a fluorescent derivative of Congo red with high binding affinity toward amyloid fibrils and good blood-brain barrier permeability. Using methoxy-X04, we investigated whether amyloid-targeting ligands can be also employed for direct imaging of amyloid deposits associated with some prion diseases. Such a method could potentially become a novel diagnostic approach for these conditions. Studies were performed on MB mice infected with the 87V mouse-adapted scrapie strain. Labeling of PrP amyloid plaques in brains of presymptomatic and symptomatic mice was demonstrated using in vivo transcranial two-photon microscopy after systemic administration of methoxy-X04. During real-time imaging, PrP amyloid deposits could be clearly distinguished 15 min after intravenous administration of methoxy-X04. The ligand showed rapid clearance from brain areas that did not contain amyloid deposits. PrP amyloid deposits could also be detected by direct application of methoxy-X04 on cerebellar sections from GSS patients. These results suggest that methoxy-X04 or similar derivatives could be used as PET imaging agents to improve the diagnosis of human prion diseases associated with amyloid deposition
— id: 44512, year: 2004, vol: 63, page: 775, stat: Journal Article,

Fibrillar amyloid deposition leads to local synaptic abnormalities and breakage of neuronal branches
Tsai, Julia; Grutzendler, Jaime; Duff, Karen; Gan, Wen-Biao
2004 Nov;7(11):1181-1183, Nature neuroscience
Amyloid plaques are a hallmark of Alzheimer disease, but their importance in its pathogenesis is controversial. By neuronal labeling and transcranial two-photon imaging, we show in a transgenic mouse model of Alzheimer disease that dendrites passing through or near fibrillar amyloid deposits undergo spine loss and shaft atrophy, and nearby axons develop large varicosities, together leading to neurite breakage and large-scale, permanent disruption of neuronal connections. Thus, fibrillar amyloid deposition is more detrimental to neuronal circuitry than previously thought, underscoring the importance of prevention and early clearance of plaques
— id: 48078, year: 2004, vol: 7, page: 1181, stat: Journal Article,

Imaging and therapeutic approaches for beta-sheet structures in prion and Alzheimer's diseases
Wisniewski, T; Pankiewicz, J; Scholtzova, H; Fernando, G; Chabalgoity, JA; Ji, Y; Wadghiri, YZ; Gan, WB; Tang, CY; Turnbull, DH; Mathis, CA; Kascsak, R; Klunk, WE; Carp, RI; Frangione, B; Sigurdsson, EM; Sadowski, M
2004 ;25(2):S30-S31, Neurobiology of aging
— id: 97595, year: 2004, vol: 25, page: S30, stat: Journal Article,

In vivo imaging of amyloid plaques in AD and prion disease model mice
Wisniewski, T; Sigurdsson, EM; Wadghiri, YZ; Carp, R; Tang, CY; Turnbull, DH; Mathis, C; Klunk, WE; Gan, WB; Sadowski, M
2004 APR ;25(12):S29-S29, Neurobiology of aging
— id: 42446, year: 2004, vol: 25, page: S29, stat: Journal Article,

Glutamate-dependent stabilization of presynaptic terminals
Gan, Wen-Biao
2003 Jun 5;38(5):677-678, Neuron
Dissecting the mechanisms underlying synapse formation and elimination is fundamental to understand how the nervous system is constructed and subsequently modified. Two studies by Tashiro et al. and by Hashimoto and Kano in this issue of Neuron provide new insights into the roles of neurotransmitter glutamate release in regulating the motility of hippocampal mossy fiber filopodia and synaptic competition among climbing fibers
— id: 39201, year: 2003, vol: 38, page: 677, stat: Journal Article,

Synaptic dynamism measured over minutes to months: age-dependent decline in an autonomic ganglion
Gan, Wen-Biao; Kwon, Elaine; Feng, Guoping; Sanes, Joshua R; Lichtman, Jeff W
2003 Sep;6(9):956-960, Nature neuroscience
Naturally occurring rearrangements of synaptic terminals are common in the nervous systems of young mammals, but little is known about their incidence in adults. Using transgenic mice that express yellow fluorescent protein (YFP) in axons, we repeatedly imaged nerve terminals in the parasympathetic submandibular ganglion. We found that the pattern of synaptic branches underwent significant rearrangements over several weeks in young adult mice. In older mice, rearrangements were less common, and synaptic patterns on individual neurons were recognizable for many months to years. Axonal branches frequently retracted or extended on a time scale of minutes in young adult mice, but seldom in mature animals. These results provide direct evidence for a decrease in plasticity of interneuronal connections as animals make the transition from young adulthood to middle age. The long-term stability of synaptic patterns could provide a structural basis for the persistence of memory in the adult nervous system
— id: 112408, year: 2003, vol: 6, page: 956, stat: Journal Article,

Rapid labeling of neuronal populations by ballistic delivery of fluorescent dyes
Grutzendler, Jaime; Tsai, Julia; Gan, Wen-Biao
2003 May;30(1):79-85, Methods
Particle-mediated ballistic delivery of fluorescent dyes has been recently used to label neuronal populations in a rapid and efficient fashion. Here we describe detailed protocols for this technique as well as recent improvements in its implementation. This technique allows rapid labeling of entire neurons in a Golgi-like manner after membranes of individual neurons are contacted by particles coated with lipophilic dyes. Neurons can be labeled by dyes of different colors at controlled densities to facilitate the study of structural interactions between cells. Furthermore, in conjunction with other histochemical labeling methods, the technique can be used to study changes in neuronal structures associated with pathologic processes in animal models or postmortem human brain. In addition to lipophilic dyes, water-soluble molecules such as calcium indicators can also be delivered efficiently with this technique. The method of ballistic delivery of indicators thus provides new avenues to probe the structure and function of the nervous system
— id: 39245, year: 2003, vol: 30, page: 79, stat: Journal Article,

Apolipoprotein E isoform-specific regulation of dendritic spine morphology in apolipoprotein E transgenic mice and Alzheimer's disease patients
Ji, Y; Gong, Y; Gan, W; Beach, T; Holtzman, D M; Wisniewski, T
2003 ;122(2):305-315, Neuroscience
Dendritic spines are postsynaptic sites of excitatory input in the mammalian nervous system. Apolipoprotein (apo) E participates in the transport of plasma lipids and in the redistribution of lipids among cells. A role for apoE is implicated in regeneration of synaptic circuitry after neural injury. The apoE4 allele is a major risk factor for late-onset familial and sporadic Alzheimer's disease (AD) and is associated with a poor outcome after brain injury. ApoE isoforms are suggested to have differential effects on neuronal repair mechanisms. In vitro studies have demonstrated the neurotrophic properties of apoE3 on neurite outgrowth. We have investigated the influence of apoE genotype on neuronal cell dendritic spine density in mice and in human postmortem tissue. In order to compare the morphology of neurons developing under different apoE conditions, gene gun labeling studies of dendritic spines of dentate gyrus (DG) granule cells of the hippocampus were carried out in wild-type (WT), human apoE3, human apoE4 expressing transgenic mice and apoE knockout (KO) mice; the same dendritic spine parameters were also assessed in human postmortem DG from individuals with and without the apoE4 gene. Quantitative analysis of dendritic spine length, morphology, and number was carried out on these mice at 3 weeks, 1 and 2 years of age. Human apoE3 and WT mice had a higher density of dendritic spines than human E4 and apoE KO mice in the 1 and 2 year age groups (P<0.0001), while at 3 weeks there were no differences between the groups. These age dependent differences in the effects of apoE isoforms on neuronal integrity may relate to the increased risk of dementia in aged individuals with the apoE4 allele. Significantly in human brain, apoE4 dose correlated inversely with dendritic spine density of DG neurons cell in the hippocampus of both AD (P=0.0008) and aged normal controls (P=0.0015). Our findings provide one potential explanation for the increased cognitive decline seen in aged and AD patients expressing apoE4
— id: 46270, year: 2003, vol: 122, page: 305, stat: Journal Article,

Rapid labeling of neuronal structures in post-mortem human brain by ballistic delivery of lipophilic dyes
Grutzendler, J; Gong, YD; Gan, WB; Wisniewski, T
2002 Jul-Aug;23(1):1762-, Neurobiology of aging
— id: 32435, year: 2002, vol: 23, page: 1762, stat: Journal Article,

Long-term dendritic spine stability in the adult cortex
Grutzendler, Jaime; Kasthuri, Narayanan; Gan, Wen-Biao
2002 Dec 19-26;420(6917):812-816, Nature
The structural dynamics of synapses probably has a crucial role in the development and plasticity of the nervous system. In the mammalian brain, the vast majority of excitatory axo-dendritic synapses occur on dendritic specializations called 'spines'. However, little is known about their long-term changes in the intact developing or adult animal. To address this question we developed a transcranial two-photon imaging technique to follow identified spines of layer-5 pyramidal neurons in the primary visual cortex of living transgenic mice expressing yellow fluorescent protein. Here we show that filopodia-like dendritic protrusions, extending and retracting over hours, are abundant in young animals but virtually absent from the adult. In young mice, within the 'critical period' for visual cortex development, approximately 73% of spines remain stable over a one-month interval; most changes are associated with spine elimination. In contrast, in adult mice, the overwhelming majority of spines (approximately 96%) remain stable over the same interval with a half-life greater than 13 months. These results indicate that spines, initially plastic during development, become remarkably stable in the adult, providing a potential structural basis for long-term information storage
— id: 39348, year: 2002, vol: 420, page: 812, stat: Journal Article,

Apolipoprotein E expression modulates neuronal spine density in an isotype specific manner in transgenic mice
Ji, Y; Gong, YD; Gan, WB; Wisniewski, T
2002 Jul-Aug;23(1):1472-, Neurobiology of aging
— id: 32426, year: 2002, vol: 23, page: 1472, stat: Journal Article,

Imaging calcium dynamics in the nervous system by means of ballistic delivery of indicators
Kettunen, Petronella; Demas, Jay; Lohmann, Christian; Kasthuri, Narayanan; Gong, Yandao; Wong, Rachel O L; Gan, Wen-Biao
2002 Sep 15;119(1):37-43, Journal of neuroscience methods
The use of fluorescence-based calcium indicators has, over the years, unraveled important calcium-dependent mechanisms underlying neuronal function and development. However, difficulties associated with the loading of calcium indicators have limited their widespread use, particularly for the study of neuronal processing in the adult nervous system. Here, we show that in the central and peripheral nervous systems, populations of neurons and their processes, including dendritic spines and filopodia, can be labeled rapidly and efficiently by delivering calcium indicator-coated particles using a 'gene gun'. Importantly, neuronal labeling occurred both in vitro and in vivo, and across a wide range of ages and preparations. The labeled cells demonstrate spontaneous and evoked calcium transients, indicating that particle-mediated delivery is not deleterious to neuronal function. Furthermore, unlike loading with patch pipettes, cytoplasmic content is preserved following ballistic loading. This enables the study of calcium-dependent second messenger pathways without loss of signaling components. The ballistic delivery of calcium indicators thus opens up many new avenues for further exploration of the structure and function of the nervous system from single spines to neuronal networks
— id: 112409, year: 2002, vol: 119, page: 37, stat: Journal Article,

Local synaptic abnormalities associated with fibrillar beta- amyloid in APP and PS1 double transgenic mice
Tsai, J; Gan, WB; Matsuoka, Y; Duff, K
2002 ;Suppl 1(757-757):Jul-Aug 1, Neurobiology of aging
— id: 32415, year: 2002, vol: Suppl 1, page: Jul, stat: Journal Article,

Asynchronous synapse elimination in neonatal motor units: studies using GFP transgenic mice
Keller-Peck, C R; Walsh, M K; Gan, W B; Feng, G; Sanes, J R; Lichtman, J W
2001 Aug 16;31(3):381-394, Neuron
In developing muscle, synapse elimination reduces the number of motor axons that innervate each postsynaptic cell. This loss of connections is thought to be a consequence of axon branch trimming. However, branch retraction has not been observed directly, and many questions remain, such as: do all motor axons retract branches, are eliminated branches withdrawn synchronously, and are withdrawing branches localized to particular regions? To address these questions, we used transgenic mice that express fluorescent proteins in small subsets of motor axons, providing a unique opportunity to reconstruct complete axonal arbors and identify all the postsynaptic targets. We found that, during early postnatal development, each motor axon loses terminal branches, but retracting branches withdraw asynchronously and without obvious spatial bias, suggesting that local interactions at each neuromuscular junction regulate synapse elimination
— id: 141724, year: 2001, vol: 31, page: 381, stat: Journal Article,

Multicolor "DiOlistic" labeling of the nervous system using lipophilic dye combinations
Gan WB; Grutzendler J; Wong WT; Wong RO; Lichtman JW
2000 Aug;27(2):219-225, Neuron
We describe a technique for rapid labeling of a large number of cells in the nervous system with many different colors. By delivering lipophilic dye-coated particles to neuronal preparations with a 'gene gun,' individual neurons and glia whose membranes are contacted by the particles are quickly labeled. Using particles that are each coated with different combinations of various lipophilic dyes, many cells within a complex neuronal network can be simultaneously labeled with a wide variety of colors. This approach is most effective in living material but also labels previously fixed material. In living material, labeled neurons continue to show normal synaptic responses and undergo dendritic remodeling. This technique is thus useful for studying structural plasticity of neuronal circuits in living preparations. In addition, the Golgi-like labeling of neurons with many different colors provides a novel way to study neuronal connectivity
— id: 11502, year: 2000, vol: 27, page: 219, stat: Journal Article,

TrkB works at postsynaptic sites
Gan WB
1999 Nov;24(3):491-492, Neuron
— id: 8378, year: 1999, vol: 24, page: 491, stat: Journal Article,

Vital imaging and ultrastructural analysis of individual axon terminals labeled by iontophoretic application of lipophilic dye
Gan, W B; Bishop, D L; Turney, S G; Lichtman, J W
1999 Oct 30;93(1):13-20, Journal of neuroscience methods
We describe a method for in vivo confocal fluorescence imaging of synaptic terminals and subsequent electron microscopic reconstructions of the same terminals. By iontophoretically applying lipophilic dye to nerve terminals at a single neuromuscular junction with a sharp microelectrode in living neonatal mice, we were able to quickly label other synaptic terminals of the same motor unit. This vital labeling technique allows the same synapses to be imaged in living animals for several days. By using two dyes applied to separate junctions we could visualize competing axons converging at the same site. We also show that similar approaches can be used to study synaptic inputs to neurons. Following photoconversion, the dye labeled axons and synapses were easily identified and distinguished from unlabeled synapses of other axons ultrastructurally. This new labeling technique thus provides a useful means to study reorganization of synaptic structure at high temporal and spatial resolution
— id: 141723, year: 1999, vol: 93, page: 13, stat: Journal Article,

Synaptic segregation at the developing neuromuscular junction
Gan WB; Lichtman JW
1998 Nov 20;282(5393):1508-1511, Science
Throughout the developing nervous system, competition between axons causes the permanent removal of some synaptic connections. In mouse neuromuscular junctions at birth, terminal branches of different axons are intermingled. However, during the several weeks after birth, these branches progressively segregated into nonoverlapping compartments before the complete withdrawal of all but one axon. Segregation was caused by selective branch atrophy, detachment, and withdrawal; the axon branches that were nearest to the competitor's branches were removed before the more distant branches were removed. This progression suggests that the signals that mediate the competitive removal of synapses must decrease in potency over short distances
— id: 8377, year: 1998, vol: 282, page: 1508, stat: Journal Article,