Mechanisms of Central Neuron Synaptogenesis

中枢神经元突触发生的机制

• 批准号: 7057818
• 负责人: ANN M CRAIG
• 金额: $ 21.36万
• 依托单位: UNIVERSITY OF BRITISH COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-06-01 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7057818
• 关键词: GABA receptor; RNA interference; cell adhesion molecules; fibroblasts; gamma aminobutyrate; glutamate receptor; glutamates; hippocampus; immunocytochemistry; intermolecular interaction; laboratory rat; membrane proteins; nerve /myelin protein; neurons; point mutation; protein isoforms; protein localization; protein protein interaction; protein structure function; synaptic vesicles; synaptogenesis; tissue /cell culture; transfection /expression vector

DESCRIPTION (provided by applicant): The long-term goal of this research is to understand how central neuron synapses form. Our current focus is to test the function of neurexins in synapse formation between hippocampal neurons. Beta-neurexins bind to neuroligins-1, a component of glutamate postsynaptic sites. Since neurexins bind synaptotagmin and presynaptic PDZ proteins including CASK and Mints, and neuroligins bind postsynaptic PDZ proteins including PSD-95, it has been suggested that the trans-synaptic neurexin-neuroligin interaction is involved in initiation of glutamatergic synapses. Furthermore, neuroligins expressed on the surface of fibroblasts can induce clustering of synaptic vesicles in contacting axons, both glutamate and GABA axons. In preliminary experiments, we found that neurexin expressed on the surface of fibroblasts induces clustering of the excitatory postsynaptic scaffolding protein PSD-95 and NMDA glutamate receptors, and of the inhibitory postsynaptic scaffolding protein gephyrin and GABA receptors, in contacting dendrites. Other proteins distantly related to neurexins including agrin cannot cluster either glutamate or GABA postsynaptic components. A putative role for neurexins at GABA synapses is supported by the expression and presynaptic localization of neurexins in GABA as well as glutamate axons. Thus the present state of knowledge suggests a central role for neurexins in both glutamate and GABA synapse formation, but our knowledge is incomplete. We propose to test the hypothesis that different isoforms of neurexins, and perhaps neuroligins, promote differentiation of glutamate and GABA synapses. We will define the residues of neurexin-lb necessary for inducing clustering of GABA and glutamate postsynaptic components, and test the possibility that different neuroligin isoforms mediate these effects. We will test the other neurexin isoforms la, 2a/b, and 3a/b for similar inducing and binding activity. We will determine the subcellular distributions of different neuroligin isoforms, and test their ability to induce presynaptic differentiation in GABA and glutamate axons. Finally, we will use two approaches to knock out the function of neurexins in hippocampal cultures and determine whether neurexins are essential for any aspect of synaptogenesis. These experiments address fundamental mechanisms of brain development with implications for therapeutic approaches to regeneration following neurological disorders or trauma.

描述（由申请人提供）：这项研究的长期目标是了解中央神经元突触的形成方式。我们目前的重点是测试神经素在海马神经元之间突触形成中的功能。 β-系蛋白与谷氨酸后突触部位的成分Neuroligins-1结合。由于神经毒素结合突触毒素和突触前PDZ蛋白，包括木桶和薄荷蛋白，而神经素结合了包括PSD-95在内的突触后PDZ蛋白，因此已经提出了反式突破性神经神经素 - 神经素 - 神经素 - 蛋白质 - 尿素 - 素素的相互作用涉及Glutameargic Stynapses的启动。此外，在成纤维细胞表面表达的神经素可以在接触轴突（谷氨酸和GABA轴突）中诱导突触囊泡的聚类。在初步实验中，我们发现在成纤维细胞表面表达的神经毒素会诱导兴奋性的突触后支架蛋白PSD-95和NMDA谷氨酸受体的聚类，以及抑制性的突触后触觉后骨织蛋白的蛋白质gephyrin和Gaba受体。与包括agrin的神经毒素相关的其他蛋白质不能聚集谷氨酸或GABA突触后成分。神经毒素在GABA突触中的推定作用得到了神经毒素在GABA和谷氨酸轴突中的表达和突触前定位的支持。因此，当前的知识状态表明神经毒素在谷氨酸和GABA突触形成中的核心作用，但我们的知识并不完整。我们建议检验以下假设：神经毒素和神经素的不同同工型促进谷氨酸和GABA突触的分化。我们将定义为诱导GABA和谷氨酸后突触成分聚类所必需的神经LB的残基，并测试不同神经素同工型介导这些作用的可能性。我们将测试其他神经毒素同工型LA，2A/B和3A/B，以实现类似的诱导和结合活性。我们将确定不同神经素同工型的亚细胞分布，并测试它们在GABA和谷氨酸轴突中诱导突触前分化的能力。最后，我们将使用两种方法来淘汰神经毒素在海马培养物中的功能，并确定神经毒素是否对于突触发生的任何方面都是必不可少的。这些实验介绍了大脑发育的基本机制，对神经系统疾病或创伤后再生的治疗方法的影响。