Glutamate receptor recruitment to new synapses in vivo

体内谷氨酸受体募集到新突触

• 批准号: 7583713
• 负责人: PHILIP ERIC WASHBOURNE
• 金额: $ 30.68万
• 依托单位: UNIVERSITY OF OREGON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-01-01 至 2013-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7583713
• 关键词: Actins; Adhesives; Affect; Afferent Neurons; Autistic Disorder; Axon; Behavior; Behavioral; Binding; Biochemical; Biochemistry; Biological Assay; Cell Adhesion Molecules; Cell membrane; Cells; Chemical Synapse; Coculture Techniques; Cognition; Communication; Data; Dendrites; Development; Electrophysiology (science); Elements; Embryo; Event; F-Actin; Family; Family member; Gene Family; Genetic; Glutamate Receptor; Glutamates; Hippocampus (Brain); Image; Individual; Kinetics; Knock-in Mouse; Knockout Mice; Lead; Life; Light; Mediating; Mental Retardation; Mental disorders; Methods; Minor; Modeling; Molecular; Motor; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; Nature; Nervous System Physiology; Neuraxis; Neuroglia; Neurons; Neurotransmitter Receptor; Neurotransmitters; Organism; Preparation; Presynaptic Terminals; Property; Proteins; Rattus; Recruitment Activity; Reflex action; Research; Schizophrenia; Sensorimotor functions; Site; Spectrin; Speed; Spinal; Spinal Cord; Surface; Synapses; Synaptic Vesicles; System; Techniques; Testing; Thinking; Time; Touch sensation; Travel; Vertebral column; Vesicle; Work; Zebrafish; base; behavior test; developmental disease; granule cell; immunological synapse; in vitro Assay; in vivo; interdisciplinary approach; knock-down; member; nervous system development; nervous system disorder; novel; postsynaptic; public health relevance; research study; response; scaffold; synaptogenesis

DESCRIPTION (provided by applicant): Chemical synapses are the primary means for transmitting information from one neuron to the next. Synapses are initially formed during development of the nervous system, and formation of appropriate synapses is crucial for establishment of neuronal circuits that underlie behavior and cognition. Minor irregularities during synapse formation can lead to developmental disorders such as autism, mental retardation and may contribute to psychological disorders. Most synapses in the vertebrate central nervous system (CNS) depend on the neurotransmitter glutamate, and thus glutamatergic synapses have been an important focus of study in trying to unravel these and other neurological disorders. A novel family of cell adhesion molecules (CAMs), the Synaptic Cell Adhesion Molecules (SynCAMs), has recently been proposed to mediate the formation of synapses. However, this work is based on experiments in neuronal culture, and knock-out mouse data so far does not corroborate this. Furthermore, it remains unknown how the SynCAMs, and CAMs in general, bring about the recruitment of synaptic elements to new adhesive contacts. We propose to test a model describing specific mechanisms through which SynCAM family members 1 and 2 can recruit both synaptic vesicles (SVs) to the presynaptic terminal and glutamate receptors to the postsynaptic specialization. We hypothesize that an interaction between SynCAM1 or 2 and CASK in axons can directly tether SV precursors to the site of SynCAM/SynCAM interaction. We also propose that binding of DAL-1 to SynCAM1 or 2 in dendrites results in formation of an actin/spectrin subsynaptic scaffold. These cytoskeletal elements then serve two functions: 1) strengthening of the adhesive nature of the synapse and morphological remodeling to generate a spine and 2) recruitment of NMDA type glutamate receptor transport packets via an actin-dependent transport mechanism. We propose to use a variety of techniques including biochemistry, immunolabeling, live-imaging, electrophysiology and behavioral tests, because a multidisciplinary approach will comprehensively test our model. We also propose to use various neuronal preparations for our experiments including cultured hippocampal neurons, cultured cerebellar granule cells (CGCs) and spinal cord neurons in zebrafish embryos in vivo. Testing our hypothesis in zebrafish will shed light on whether these proteins and their interactions are required for forming a specific circuit in a developing embryo that is required for a sensorimotor reflex. Our approach gives us the unprecedented opportunity to determine the mechanisms of glutamatergic synapse formation using behavioral, electrophysiological, genetic and biochemical approaches in both neuronal cultures and in a living vertebrate. PUBLIC HEALTH RELEVANCE: Synapses are sites at which nerve cells communicate with each other. Communication of nerve cells is absolutely necessary for nervous system function, ranging from simple reflexes to expressing philosophical thoughts. Errors during the formation of synapses are thought to be at the basis of nervous system disorders such as autism, mental retardation and schizophrenia. We propose to study two members of a family of genes, the Synaptic Cell Adhesion Molecules (SynCAMs) 1 and 2, which are thought to mediate the formation of synapses during development. Almost nothing is yet known about how SynCAMs make a contact site between two nerve cells become a place for active communication. We will investigate how these molecules carry out synapse formation by testing their function in rat nerve cells grown in culture and by testing their function in developing zebrafish embryos. These two systems allow us to determine the nature of the molecular interactions and to determine the importance of these interactions for the formation of synapses in a living organism, respectively. Our research will help understand the mechanisms by which synapses form and bring us closer to identifying the molecular deficits in individuals with autism and mental retardation.

描述（由申请人提供）：化学突触是将信息从一个神经元传输到另一个神经元的主要手段。最初是在神经系统发展过程中形成的突触，适当的突触的形成对于建立行为和认知构成的神经元回路至关重要。突触形成过程中的较小违规行为会导致自闭症，智力低下的发育障碍，并可能导致心理疾病。脊椎动物中枢神经系统（CNS）中的大多数突触都取决于神经递质谷氨酸，因此谷氨酸能突触已成为试图揭示这些和其他神经系统疾病的研究的重要重点。最近已经提出了一种新型的细胞粘附分子（CAM），突触细胞粘附分子（Syncam）的家族，以介导突触的形成。但是，这项工作基于神经元培养的实验，到目前为止，敲除小鼠数据并不能证实这一点。此外，尚不清楚Syncam和CAM通常如何将突触元素募集到新的粘合剂接触中。我们建议测试一个描述特定机制的模型，该模型通过该模型可以通过该模型募集两个突触囊泡（SVS）到突触前末端和谷氨酸盐受体到突触后专业化。我们假设Syncam1或2与轴突中的木桶之间的相互作用可以直接将SV前体绑定到Syncam/Syncam相互作用的位点。我们还建议DAL-1与syncam1或2在树突中的结合会导致形成肌动蛋白/谱素次突触支架。然后，这些细胞骨架元素具有两个功能：1）通过肌动蛋白依赖性转运机制募集突触和形态重塑的粘附性和形态重塑的粘附性。我们建议使用各种技术，包括生物化学，免疫标记，现场成像，电生理学和行为测试，因为多学科方法将全面测试我们的模型。我们还建议在我们的实验中使用各种神经元制剂进行实验，包括培养的海马神经元，培养的小脑颗粒细胞（CGC）和体内斑马鱼胚胎中的脊髓神经元。在斑马鱼中检验我们的假设将阐明这些蛋白质及其相互作用是否需要在发育中的胚胎中形成特定电路，而感觉运动反射需要。我们的方法为我们提供了前所未有的机会，可以使用神经元培养物和活脊椎动物中的行为，电生理学，遗传学和生化方法来确定谷氨酸能突触形成的机制。公共卫生相关性：突触是神经细胞相互通信的部位。神经细胞的交流对于神经系统功能是绝对必要的，从简单反射到表达哲学思想。突触形成过程中的错误被认为是基于自闭症，智力低下和精神分裂症等神经系统疾病的基础。我们建议研究一个基因家族的两个成员，即突触细胞粘附分子（Syncams）1和2，它们被认为可以介导发育过程中突触的形成。关于syncam如何使两个神经细胞之间的接触位点成为主动交流的地方，几乎一无所知。我们将研究这些分子如何通过测试其在培养物中生长的大鼠神经细胞的功能以及测试其在发展斑马鱼胚胎中的功能来进行突触。这两个系统使我们能够确定分子相互作用的性质，并分别确定这些相互作用在生物体中的突触形成的重要性。我们的研究将有助于理解突触形成的机制，并使我们更加接近确定自闭症和智力低下的个体的分子缺陷。