Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 63
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
2.
Trends Cell Biol ; 2024 Jun 08.
Artigo em Inglês | MEDLINE | ID: mdl-38853082

RESUMO

Astrocytes are multifaceted glial cell types that perform structural, functional, metabolic, and homeostatic roles in the brain. Recent studies have revealed mechanisms underlying the diversity of bidirectional communication modes between astrocytes and neurons - the fundamental organizing principle shaping synaptic properties at tripartite synapses. These astrocyte-neuron interactions are critical for the proper functioning of synapses and neural circuits. This review focuses on molecular mechanisms that direct these interactions, highlighting the versatile roles of multiple adhesion-based paths that likely modulate them, often in a context-dependent manner. It also describes how astrocyte-mediated processes go awry in certain brain disorders and provides a timely insight on the pivotal roles of astrocyte-neuron interactions in synaptic integrity and their relevance to understanding and treating neurological disorders.

3.
Proc Natl Acad Sci U S A ; 121(26): e2322978121, 2024 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-38900791

RESUMO

MDGA (MAM domain containing glycosylphosphatidylinositol anchor) family proteins were previously identified as synaptic suppressive factors. However, various genetic manipulations have yielded often irreconcilable results, precluding precise evaluation of MDGA functions. Here, we found that, in cultured hippocampal neurons, conditional deletion of MDGA1 and MDGA2 causes specific alterations in synapse numbers, basal synaptic transmission, and synaptic strength at GABAergic and glutamatergic synapses, respectively. Moreover, MDGA2 deletion enhanced both N-methyl-D-aspartate (NMDA) receptor- and α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor-mediated postsynaptic responses. Strikingly, ablation of both MDGA1 and MDGA2 abolished the effect of deleting individual MDGAs that is abrogated by chronic blockade of synaptic activity. Molecular replacement experiments further showed that MDGA1 requires the meprin/A5 protein/PTPmu (MAM) domain, whereas MDGA2 acts via neuroligin-dependent and/or MAM domain-dependent pathways to regulate distinct postsynaptic properties. Together, our data demonstrate that MDGA paralogs act as unique negative regulators of activity-dependent postsynaptic organization at distinct synapse types, and cooperatively contribute to adjustment of excitation-inhibition balance.


Assuntos
Hipocampo , Sinapses , Transmissão Sináptica , Animais , Sinapses/metabolismo , Camundongos , Hipocampo/metabolismo , Hipocampo/citologia , Transmissão Sináptica/fisiologia , Neurônios/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Receptores de N-Metil-D-Aspartato/genética , Camundongos Knockout , Receptores de AMPA/metabolismo , Receptores de AMPA/genética , Moléculas de Adesão Celular Neuronais/metabolismo , Moléculas de Adesão Celular Neuronais/genética , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Células Cultivadas
6.
Nat Commun ; 15(1): 1624, 2024 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-38388459

RESUMO

LAR-RPTPs are evolutionarily conserved presynaptic cell-adhesion molecules that orchestrate multifarious synaptic adhesion pathways. Extensive alternative splicing of LAR-RPTP mRNAs may produce innumerable LAR-RPTP isoforms that act as regulatory "codes" for determining the identity and strength of specific synapse signaling. However, no direct evidence for this hypothesis exists. Here, using targeted RNA sequencing, we detected LAR-RPTP mRNAs in diverse cell types across adult male mouse brain areas. We found pronounced cell-type-specific patterns of two microexons, meA and meB, in Ptprd mRNAs. Moreover, diverse neural circuits targeting the same neuronal populations were dictated by the expression of different Ptprd variants with distinct inclusion patterns of microexons. Furthermore, conditional ablation of Ptprd meA+ variants at presynaptic loci of distinct hippocampal circuits impaired distinct modes of synaptic transmission and objection-location memory. Activity-triggered alterations of the presynaptic Ptprd meA code in subicular neurons mediates NMDA receptor-mediated postsynaptic responses in CA1 neurons and objection-location memory. Our data provide the evidence of cell-type- and/or circuit-specific expression patterns in vivo and physiological functions of LAR-RPTP microexons that are dynamically regulated.


Assuntos
Sinapses , Transmissão Sináptica , Camundongos , Animais , Masculino , Transmissão Sináptica/fisiologia , Sinapses/metabolismo , Transdução de Sinais , Neurônios/metabolismo , Moléculas de Adesão Celular/metabolismo , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/metabolismo , RNA Mensageiro/metabolismo
7.
Curr Opin Neurobiol ; 81: 102728, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37236068

RESUMO

Postsynaptic GABAergic receptors interact with various membrane and intracellular proteins to mediate inhibitory synaptic transmission. They form structural and/or signaling synaptic protein complexes that perform a variety of postsynaptic functions. In particular, the key GABAergic synaptic scaffold, gephyrin, and its interacting partners govern downstream signaling pathways that are essential for GABAergic synapse development, transmission, and plasticity. In this review, we discuss recent researches on GABAergic synaptic signaling pathways. We also outline the main outstanding issues that need to be addressed in this field and highlight the association of dysregulated GABAergic synaptic signaling with the onset of various brain disorders.


Assuntos
Receptores de GABA-A , Sinapses , Receptores de GABA-A/metabolismo , Sinapses/fisiologia , Transmissão Sináptica/fisiologia , Proteínas de Transporte/metabolismo , Plasticidade Neuronal/fisiologia
8.
Sci Rep ; 13(1): 6547, 2023 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-37085584

RESUMO

Neuroinflammation impacts the brain and cognitive behavior through microglial activation. In this study, we determined the temporal sequence from microglial activation to synaptic dysfunction and cognitive behavior induced by neuroinflammation in mice. We found that LPS injection activated microglia within a short period, followed by impairments in GABAergic synapses, and that these events led to long-term cognitive impairment. We demonstrated that, 3 days after LPS injection, microglia in the hippocampus were significantly activated due to the LPS-induced inflammation in association with alterations in cellular morphology, microglial density, and expression of phagocytic markers. GABAergic synaptic impairments were detected at 4-6 days after LPS treatment, a time when microglia activity had returned to normal. Consequently, memory impairment persisted for 6 days after injection of LPS. Our results suggest that neuroinflammation induces microglia activation, GABAergic synaptic deficits and prolonged memory impairment over a defined temporal sequence. Our observations provide insight into the temporal sequence of neuroinflammation-associated brain pathologies. Moreover, the specific loss of inhibitory synapses accompanying the impaired inhibitory synaptic transmission provides mechanistic insight that may explain the prolonged cognitive deficit observed in patients with neuroinflammation. Thus, this study provides essential clues regarding early intervention strategies against brain pathologies accompanying neuroinflammation.


Assuntos
Disfunção Cognitiva , Microglia , Camundongos , Animais , Microglia/metabolismo , Lipopolissacarídeos/metabolismo , Doenças Neuroinflamatórias , Disfunção Cognitiva/metabolismo , Hipocampo/metabolismo , Inflamação/patologia , Transtornos da Memória/metabolismo , Camundongos Endogâmicos C57BL
9.
Nat Commun ; 13(1): 4112, 2022 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-35840571

RESUMO

SLITRK2 is a single-pass transmembrane protein expressed at postsynaptic neurons that regulates neurite outgrowth and excitatory synapse maintenance. In the present study, we report on rare variants (one nonsense and six missense variants) in SLITRK2 on the X chromosome identified by exome sequencing in individuals with neurodevelopmental disorders. Functional studies showed that some variants displayed impaired membrane transport and impaired excitatory synapse-promoting effects. Strikingly, these variations abolished the ability of SLITRK2 wild-type to reduce the levels of the receptor tyrosine kinase TrkB in neurons. Moreover, Slitrk2 conditional knockout mice exhibited impaired long-term memory and abnormal gait, recapitulating a subset of clinical features of patients with SLITRK2 variants. Furthermore, impaired excitatory synapse maintenance induced by hippocampal CA1-specific cKO of Slitrk2 caused abnormalities in spatial reference memory. Collectively, these data suggest that SLITRK2 is involved in X-linked neurodevelopmental disorders that are caused by perturbation of diverse facets of SLITRK2 function.


Assuntos
Transtornos do Neurodesenvolvimento , Sinapses , Animais , Cognição , Hipocampo/fisiologia , Camundongos , Camundongos Knockout , Transtornos do Neurodesenvolvimento/genética , Transtornos do Neurodesenvolvimento/metabolismo , Sinapses/metabolismo
10.
J Inflamm Res ; 15: 3053-3063, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35645573

RESUMO

Background: Peripheral inflammation-triggered mild neuroinflammation impacts the brain and behavior through microglial activation. In this study, we performed an unbiased analysis of the vulnerability of different brain areas to neuroinflammation induced by systemic inflammation. Methods: We injected mice with a single low dose of LPS to induce mild inflammation and then analyzed microglial activation in 34 brain regions by immunohistochemical methods and whole-brain imaging using multi-slide scanning microscopy. We also conducted quantitative RT-PCR to measure the levels of inflammatory cytokines in selected brain regions of interest. Results: We found that microglia in different brain regions are differentially activated by mild, LPS-induced inflammation relative to the increase in microglia numbers or increased CD68 expression. The increased number of microglia induced by mild inflammation was not attributable to infiltration of peripheral immune cells. In addition, microglia residing in brain regions, in which a single low-dose injection of LPS produced microglial changes, preferentially generated pro-inflammatory cytokines. Conclusion: Our results suggest that mild neuroinflammation induces regionally different microglia activation, producing pro-inflammatory cytokines. Our observations provide insight into induction of possible region-specific neuroinflammation-associated brain pathologies through microglial activation.

11.
Trends Neurosci ; 45(7): 517-528, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35577622

RESUMO

Multiple synaptic adhesion proteins are thought to collectively define the properties of specific synapses and thereby shape the architectures of neural circuits. Growing evidence supports a molecular model in which a set of central hub proteins interacts with a vast number of other proteins to organize multifarious synaptic adhesion pathways. However, several fundamental open questions remain, partly owing to drawbacks in current approaches and interpretations. In this opinion, we provide an overview of synaptic adhesion pathways, underscoring open questions to be addressed in future work, and highlighting approaches for advancing understanding of synaptic adhesion processes.


Assuntos
Sinapses , Humanos , Sinapses/metabolismo
12.
Biol Psychiatry ; 91(9): 821-831, 2022 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-35219498

RESUMO

BACKGROUND: IQSEC3, a gephyrin-binding GABAergic (gamma-aminobutyric acidergic) synapse-specific guanine nucleotide exchange factor, was recently reported to regulate activity-dependent GABAergic synapse maturation, but the underlying signaling mechanisms remain incompletely understood. METHODS: We generated mice with conditional knockout (cKO) of Iqsec3 to examine whether altered synaptic inhibition influences hippocampus-dependent fear memory formation. In addition, electrophysiological recordings, immunohistochemistry, and behavioral assays were used to address our question. RESULTS: We found that Iqsec3-cKO induces a specific reduction in GABAergic synapse density, GABAergic synaptic transmission, and maintenance of long-term potentiation in the hippocampal CA1 region. In addition, Iqsec3-cKO mice exhibited impaired fear memory formation. Strikingly, Iqsec3-cKO caused abnormally enhanced activation of ribosomal P70-S6K1-mediated signaling in the hippocampus but not in the cortex. Furthermore, inhibiting upregulated S6K1 signaling by expressing dominant-negative S6K1 in the hippocampal CA1 of Iqsec3-cKO mice completely rescued impaired fear learning and inhibitory synapse density but not deficits in long-term potentiation maintenance. Finally, upregulated S6K1 signaling was rescued by IQSEC3 wild-type, but not by an ARF-GEF (adenosine diphosphate ribosylation factor-guanine nucleotide exchange factor) inactive IQSEC3 mutant. CONCLUSIONS: Our results suggest that IQSEC3-mediated balanced synaptic inhibition in hippocampal CA1 is critical for the proper formation of hippocampus-dependent fear memory.


Assuntos
Medo , Fatores de Troca do Nucleotídeo Guanina , Hipocampo , Sinapses , Animais , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Hipocampo/metabolismo , Potenciação de Longa Duração , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Sinapses/metabolismo , Regulação para Cima
13.
Proc Natl Acad Sci U S A ; 119(4)2022 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-35074912

RESUMO

Balanced synaptic inhibition, controlled by multiple synaptic adhesion proteins, is critical for proper brain function. MDGA1 (meprin, A-5 protein, and receptor protein-tyrosine phosphatase mu [MAM] domain-containing glycosylphosphatidylinositol anchor protein 1) suppresses synaptic inhibition in mammalian neurons, yet the molecular mechanisms underlying MDGA1-mediated negative regulation of GABAergic synapses remain unresolved. Here, we show that the MDGA1 MAM domain directly interacts with the extension domain of amyloid precursor protein (APP). Strikingly, MDGA1-mediated synaptic disinhibition requires the MDGA1 MAM domain and is prominent at distal dendrites of hippocampal CA1 pyramidal neurons. Down-regulation of APP in presynaptic GABAergic interneurons specifically suppressed GABAergic, but not glutamatergic, synaptic transmission strength and inputs onto both the somatic and dendritic compartments of hippocampal CA1 pyramidal neurons. Moreover, APP deletion manifested differential effects in somatostatin- and parvalbumin-positive interneurons in the hippocampal CA1, resulting in distinct alterations in inhibitory synapse numbers, transmission, and excitability. The infusion of MDGA1 MAM protein mimicked postsynaptic MDGA1 gain-of-function phenotypes that involve the presence of presynaptic APP. The overexpression of MDGA1 wild type or MAM, but not MAM-deleted MDGA1, in the hippocampal CA1 impaired novel object-recognition memory in mice. Thus, our results establish unique roles of APP-MDGA1 complexes in hippocampal neural circuits, providing unprecedented insight into trans-synaptic mechanisms underlying differential tuning of neuronal compartment-specific synaptic inhibition.


Assuntos
Precursor de Proteína beta-Amiloide/metabolismo , Hipocampo/metabolismo , Hipocampo/fisiopatologia , Moléculas de Adesão de Célula Nervosa/genética , Inibição Neural , Sinapses/metabolismo , Precursor de Proteína beta-Amiloide/genética , Região CA1 Hipocampal , Proteínas de Transporte , Dendritos/metabolismo , Neurônios GABAérgicos/metabolismo , Interneurônios , Modelos Biológicos , Moléculas de Adesão de Célula Nervosa/química , Moléculas de Adesão de Célula Nervosa/metabolismo , Inibição Neural/genética , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Células Piramidais/metabolismo , Receptores de GABA-B/metabolismo , Transmissão Sináptica
14.
Proc Natl Acad Sci U S A ; 119(3)2022 01 18.
Artigo em Inglês | MEDLINE | ID: mdl-35022233

RESUMO

Synaptic cell-adhesion molecules (CAMs) organize the architecture and properties of neural circuits. However, whether synaptic CAMs are involved in activity-dependent remodeling of specific neural circuits is incompletely understood. Leucine-rich repeat transmembrane protein 3 (LRRTM3) is required for the excitatory synapse development of hippocampal dentate gyrus (DG) granule neurons. Here, we report that Lrrtm3-deficient mice exhibit selective reductions in excitatory synapse density and synaptic strength in projections involving the medial entorhinal cortex (MEC) and DG granule neurons, accompanied by increased neurotransmitter release and decreased excitability of granule neurons. LRRTM3 deletion significantly reduced excitatory synaptic innervation of hippocampal mossy fibers (Mf) of DG granule neurons onto thorny excrescences in hippocampal CA3 neurons. Moreover, LRRTM3 loss in DG neurons significantly decreased mossy fiber long-term potentiation (Mf-LTP). Remarkably, silencing MEC-DG circuits protected against the decrease in the excitatory synaptic inputs onto DG and CA3 neurons, excitability of DG granule neurons, and Mf-LTP in Lrrtm3-deficient mice. These results suggest that LRRTM3 may be a critical factor in activity-dependent synchronization of the topography of MEC-DG-CA3 excitatory synaptic connections. Collectively, our data propose that LRRTM3 shapes the target-specific structural and functional properties of specific hippocampal circuits.


Assuntos
Sincronização Cortical/fisiologia , Hipocampo/fisiologia , Proteínas de Membrana/metabolismo , Rede Nervosa/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Sinapses/fisiologia , Animais , Região CA3 Hipocampal/metabolismo , Giro Denteado/metabolismo , Córtex Entorrinal/metabolismo , Potenciação de Longa Duração , Proteínas de Membrana/deficiência , Camundongos Knockout , Fibras Musgosas Hipocampais/metabolismo , Proteínas do Tecido Nervoso/deficiência , Neurônios/metabolismo , Pseudópodes/metabolismo , Transmissão Sináptica/fisiologia
15.
Cell Rep ; 36(3): 109417, 2021 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-34289353

RESUMO

Activity-dependent GABAergic synapse plasticity is important for normal brain functions, but the underlying molecular mechanisms remain incompletely understood. Here, we show that Npas4 (neuronal PAS-domain protein 4) transcriptionally regulates the expression of IQSEC3, a GABAergic synapse-specific guanine nucleotide-exchange factor for ADP-ribosylation factor (ARF-GEF) that directly interacts with gephyrin. Neuronal activation by an enriched environment induces Npas4-mediated upregulation of IQSEC3 protein specifically in CA1 stratum oriens layer somatostatin (SST)-expressing GABAergic interneurons. SST+ interneuron-specific knockout (KO) of Npas4 compromises synaptic transmission in these GABAergic interneurons, increases neuronal activity in CA1 pyramidal neurons, and reduces anxiety behavior, all of which are normalized by the expression of wild-type IQSEC3, but not a dominant-negative ARF-GEF-inactive mutant, in SST+ interneurons of Npas4-KO mice. Our results suggest that IQSEC3 is a key GABAergic synapse component that is directed by Npas4 and ARF activity, specifically in SST+ interneurons, to orchestrate excitation-to-inhibition balance and control anxiety-like behavior.


Assuntos
Ansiedade/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Comportamento Animal , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Hipocampo/metabolismo , Interneurônios/metabolismo , Somatostatina/metabolismo , Animais , Neurônios GABAérgicos/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout , Regiões Promotoras Genéticas/genética , Ligação Proteica , Sinapses/metabolismo , Transmissão Sináptica , Regulação para Cima
16.
iScience ; 24(2): 102037, 2021 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-33532714

RESUMO

Gephyrin is critical for the structure, function, and plasticity of inhibitory synapses. Gephyrin mutations have been linked to various neurological disorders; however, systematic analyses of the functional consequences of these mutations are lacking. Here, we performed molecular dynamics simulations of gephyrin to predict how six reported point mutations might change the structural stability and/or function of gephyrin. Additional in silico analyses revealed that the A91T and G375D mutations reduce the binding free energy of gephyrin oligomer formation. Gephyrin A91T and G375D displayed altered clustering patterns in COS-7 cells and nullified the inhibitory synapse-promoting effect of gephyrin in cultured neurons. However, only the G375D mutation reduced gephyrin interaction with GABAA receptors and neuroligin-2 in mouse brain; it also failed to normalize deficits in GABAergic synapse maintenance and neuronal hyperactivity observed in hippocampal dentate gyrus-specific gephyrin-deficient mice. Our results provide insights into biochemical, cell-biological, and network-activity effects of the pathogenic G375D mutation.

17.
Prog Neurobiol ; 200: 101983, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33422662

RESUMO

Trans-synaptic cell-adhesion molecules are critical for governing various stages of synapse development and specifying neural circuit properties via the formation of multifarious signaling pathways. Recent studies have pinpointed the putative roles of trans-synaptic cell-adhesion molecules in mediating various cognitive functions. Here, we review the literature on the roles of a diverse group of central synaptic organizers, including neurexins (Nrxns), leukocyte common antigen-related receptor protein tyrosine phosphatases (LAR-RPTPs), and their associated binding proteins, in regulating properties of specific type of synapses and neural circuits. In addition, we highlight the findings that aberrant synaptic adhesion signaling leads to alterations in the structures, transmission, and plasticity of specific synapses across diverse brain areas. These results seem to suggest that proper trans-synaptic signaling pathways by Nrxns, LAR-RPTPs, and their interacting network is likely to constitute central molecular complexes that form the basis for cognitive functions, and that these complexes are heterogeneously and complexly disrupted in many neuropsychiatric and neurodevelopmental disorders.


Assuntos
Fenômenos Fisiológicos do Sistema Nervoso , Transdução de Sinais , Encéfalo/metabolismo , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/metabolismo , Sinapses/metabolismo
18.
Mol Autism ; 11(1): 87, 2020 10 30.
Artigo em Inglês | MEDLINE | ID: mdl-33126897

RESUMO

BACKGROUND: Recent progress in genomics has contributed to the identification of a large number of autism spectrum disorder (ASD) risk genes, many of which encode synaptic proteins. Our understanding of ASDs has advanced rapidly, partly owing to the development of numerous animal models. Extensive characterizations using a variety of behavioral batteries that analyze social behaviors have shown that a subset of engineered mice that model mutations in genes encoding Shanks, a family of excitatory postsynaptic scaffolding proteins, exhibit autism-like behaviors. Although these behavioral assays have been useful in identifying deficits in simple social behaviors, alterations in complex social behaviors remain largely untested. METHODS: Two syndromic ASD mouse models-Shank2 constitutive knockout [KO] mice and Shank3 constitutive KO mice-were examined for alterations in social dominance and social cooperative behaviors using tube tests and automated cooperation tests. Upon naïve and salient behavioral experience, expression levels of c-Fos were analyzed as a proxy for neural activity across diverse brain areas, including the medial prefrontal cortex (mPFC) and a number of subcortical structures. FINDINGS: As previously reported, Shank2 KO mice showed deficits in sociability, with intact social recognition memory, whereas Shank3 KO mice displayed no overt phenotypes. Strikingly, the two Shank KO mouse models exhibited diametrically opposed alterations in social dominance and cooperative behaviors. After a specific social behavioral experience, Shank mutant mice exhibited distinct changes in number of c-Fos+ neurons in the number of cortical and subcortical brain regions. CONCLUSIONS: Our results underscore the heterogeneity of social behavioral alterations in different ASD mouse models and highlight the utility of testing complex social behaviors in validating neurodevelopmental and neuropsychiatric disorder models. In addition, neural activities at distinct brain regions are likely collectively involved in eliciting complex social behaviors, which are differentially altered in ASD mouse models.


Assuntos
Comportamento Cooperativo , Proteínas dos Microfilamentos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Predomínio Social , Animais , Encéfalo/patologia , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Neurônios/metabolismo
19.
STAR Protoc ; 1(2): 100095, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-33111124

RESUMO

Clustering of synaptic vesicles along the neuronal axons is a critical mechanism underpinning proper synaptic transmission. Here, we provide a detailed protocol for analyzing the distribution of synaptic vesicles in presynaptic boutons of cultured neurons. The protocol covers preparation of cultured neurons, expression of synaptic vesicle-enriched proteins, and quantification procedures. Utilizing neurons from postnatal transgenic mice, this method can be applied to investigate the roles of synaptic genes in regulating vesicle dynamics at synaptic sites. For complete details on the use and execution of this protocol, please refer to Han et al. (2020a).


Assuntos
Axônios/fisiologia , Cultura Primária de Células/métodos , Vesículas Sinápticas/fisiologia , Animais , Células Cultivadas , Análise por Conglomerados , Camundongos , Camundongos Endogâmicos C57BL , Proteínas do Tecido Nervoso/metabolismo , Neurônios/citologia , Terminações Pré-Sinápticas/fisiologia , Sinapses/metabolismo , Transmissão Sináptica/fisiologia
20.
J Neurosci ; 40(44): 8438-8462, 2020 10 28.
Artigo em Inglês | MEDLINE | ID: mdl-33037075

RESUMO

Neurexins (Nrxns) and LAR-RPTPs (leukocyte common antigen-related protein tyrosine phosphatases) are presynaptic adhesion proteins responsible for organizing presynaptic machineries through interactions with nonoverlapping extracellular ligands. Here, we report that two members of the LAR-RPTP family, PTPσ and PTPδ, are required for the presynaptogenic activity of Nrxns. Intriguingly, Nrxn1 and PTPσ require distinct sets of intracellular proteins for the assembly of specific presynaptic terminals. In addition, Nrxn1α showed robust heparan sulfate (HS)-dependent, high-affinity interactions with Ig domains of PTPσ that were regulated by the splicing status of PTPσ. Furthermore, Nrxn1α WT, but not a Nrxn1α mutant lacking HS moieties (Nrxn1α ΔHS), inhibited postsynapse-inducing activity of PTPσ at excitatory, but not inhibitory, synapses. Similarly, cis expression of Nrxn1α WT, but not Nrxn1α ΔHS, suppressed the PTPσ-mediated maintenance of excitatory postsynaptic specializations in mouse cultured hippocampal neurons. Lastly, genetics analyses using male or female Drosophila Dlar and Dnrx mutant larvae identified epistatic interactions that control synapse formation and synaptic transmission at neuromuscular junctions. Our results suggest a novel synaptogenesis model whereby different presynaptic adhesion molecules combine with distinct regulatory codes to orchestrate specific synaptic adhesion pathways.SIGNIFICANCE STATEMENT We provide evidence supporting the physical interactions of neurexins with leukocyte common-antigen related receptor tyrosine phosphatases (LAR-RPTPs). The availability of heparan sulfates and alternative splicing of LAR-RPTPs regulate the binding affinity of these interactions. A set of intracellular presynaptic proteins is involved in common for Nrxn- and LAR-RPTP-mediated presynaptic assembly. PTPσ triggers glutamatergic and GABAergic postsynaptic differentiation in an alternative splicing-dependent manner, whereas Nrxn1α induces GABAergic postsynaptic differentiation in an alternative splicing-independent manner. Strikingly, Nrxn1α inhibits the glutamatergic postsynapse-inducing activity of PTPσ, suggesting that PTPσ and Nrxn1α might control recruitment of a different pool of postsynaptic machinery. Drosophila orthologs of Nrxns and LAR-RPTPs mediate epistatic interactions in controlling synapse structure and strength at neuromuscular junctions, underscoring the physiological significance in vivo.


Assuntos
Proteínas de Ligação ao Cálcio/fisiologia , Antígenos Comuns de Leucócito/fisiologia , Moléculas de Adesão de Célula Nervosa/fisiologia , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Drosophila/genética , Drosophila melanogaster , Potenciais Pós-Sinápticos Excitadores/fisiologia , Espaço Extracelular/metabolismo , Feminino , Células HEK293 , Humanos , Larva , Masculino , Camundongos , Conformação Molecular , Moléculas de Adesão de Célula Nervosa/metabolismo , Gravidez , Terminações Pré-Sinápticas/metabolismo , Ratos , Proteínas Tirosina Fosfatases Semelhantes a Receptores/genética , Transmissão Sináptica/fisiologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...