Robust trans-synaptic labeling technologies for cell type-specific quantitation of synaptic connectivity

强大的跨突触标记技术，用于突触连接的细胞类型特异性定量

• 批准号: 8935699
• 负责人: EDWARD M CALLAWAY
• 金额: $ 37.74万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-21 至 2020-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8935699
• 关键词: 16p11.2; 17p11.2; 7q11.23; Astrocytes; Autistic Disorder; Biological Assay; Brain; Cell Line; Cellular Assay; Cerebral cortex; Cholecystokinin; Collaborations; FMR1 Gene; Genotype; Glutamates; Glycoproteins; Haplotypes; Helper Viruses; Human; In Vitro; Infection; Label; Methods; Methyl-CpG-Binding Protein 2; Modeling; Mus; Neurons; Neurotransmitters; Patients; Pattern; Point Mutation; Rabies; Rabies virus; Reagent; Reporter; Schizophrenia; Serotyping; Specificity; Subfamily lentivirinae; Surface; Synapses; System; TSC2 gene; Technology; Testing; Tissues; Viral Vector; Wild Type Mouse; autism spectrum disorder; base; biosignature; cell type; cellular transduction; excitatory neuron; human subject; in vivo; induced pluripotent stem cell; inhibitory neuron; interest; mouse model; neural circuit; new technology; novel; pre-clinical; tool

Project Summary Neural circuits are composed of networks of specific cell types that are interconnected in precise patterns to give rise to normal brain function. Past studies have investigated the rates of connections that are formed by cultured neurons in vitro and these have revealed abnormalities both in a mouse model of autism (MeCP2 KO mice) and for human induced pluripotent stem cell (hiPSC)-derived neurons from schizophrenic patients (Brennand et al., 2011). But simple separation of cortical neurons into just excitatory versus inhibitory groups barely scratches the surface of the diversity of cortical cell types, the known specificity of connections between specific cell types in the abnormal or normal brain. In Project 5 we propose to develop and then implement novel tools that will allow assays of the cell-type specificity of connections that are formed in cultures of hiPSC- derived neurons. These assays are based on the use of glycoprotein deleted rabies viruses for trans-synaptic labeling of connected neurons, and conditional expression of EnvA/TVA targeting systems for directing the initial infection of rabies virus to specific cell types (Wickersham et al., 2007b; Marshel et al., 2010; Wall et al., 2010)

项目摘要 神经回路由特定细胞类型的网络组成，这些网络以精确的模式相互连接， 产生正常的大脑功能。过去的研究调查了由以下因素形成的连接率 在体外培养的神经元，这些都揭示了异常，无论是在小鼠模型的自闭症（MeCP 2 KO 小鼠）和来自精神分裂症患者的人诱导多能干细胞（hiPSC）衍生的神经元 （Brendal等人，2011年）。但是简单地将皮层神经元分为兴奋性和抑制性两组 这仅仅触及了皮层细胞类型多样性的表面，已知的皮层细胞之间连接的特异性， 异常或正常大脑中的特定细胞类型。在项目5中，我们建议开发并实施 新的工具，将允许测定在hiPSC培养物中形成的连接的细胞类型特异性， 衍生神经元这些测定是基于使用糖蛋白缺失的狂犬病病毒进行跨突触 连接的神经元的标记，以及EnvA/TVA靶向系统的条件表达，用于指导神经元的功能。 狂犬病病毒对特定细胞类型的初始感染（Wickersham等，2007 b; Marshel等人，2010; Wall等人， （2010年）