Molecular probes to visualize endogenous synaptic proteins in vivo

体内内源性突触蛋白可视化的分子探针

• 批准号: 8598703
• 负责人: DONALD B ARNOLD
• 金额: $ 35.88万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-15 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8598703
• 关键词: Affinity; Animals; Antibodies; Autistic Disorder; Behavioral; Binding; Biological Assay; Brain; Cells; DLG4 gene; Data; Disease; Event; Excitatory Synapse; Fibronectins; GAD65 enzyme; Genes; Genome; Goals; Grant; Human; In Vitro; Individual; Inhibitory Synapse; Label; Libraries; Life; Location; Maps; Mental Retardation; Messenger RNA; Methods; Molecular; Molecular Probes; Monitor; Morphology; Mus; Neurons; Organism; Output; Partner in relationship; Plasmids; Procedures; Production; Proteins; Publications; Recombinant Antibody; Research; Schizophrenia; Site; Specificity; Structure; Synapses; System; Testing; Time; Transcriptional Regulation; Transgenic Animals; Transgenic Mice; Vertebral column; Work; cell type; density; gephyrin; in vivo; novel; postsynaptic; presynaptic; public health relevance; recombinase; tool

DESCRIPTION (provided by applicant): The purpose of the research proposed in this grant is to generate tools that will allow endogenous synaptic proteins to be visualized in neurons in vivo. Previously we have generated recombinant antibodies known as FingRs (Fibronectin intrabodies generated with mRNA display) that can be expressed in living neurons where they label endogenous target proteins noninvasively and with high fidelity. Furthermore, a transcriptional regulation system controls the expression levels of FingRs so that they are expressed at precisely the same level as their endogenous counterparts, insuring low background. In this grant we will generate transgenic mice that express fluorescent protein-fused FingRs that recognize synaptic proteins. Synaptic proteins in individual neurons in the brains of these mice can be visualized in vivo in real time. In turn, this will allow events at the molecular level to be correlated with events at the cellular, circuit and whole animal level. In particular, by mapping the locations and amounts of synaptic proteins in neurons, it will be possible to monitor the strength of both synaptic inputs and outputs in living neurons. The ability to monitor synaptic strength will be very useful for studying diseases associated with aberrant synaptic connectivity including schizophrenia, mental retardation and autism.

描述（由申请人提供）：本次资助中提出的研究目的是生成能够在体内神经元中可视化内源性突触蛋白的工具。此前，我们已经生成了称为 FingR（通过 mRNA 展示生成的纤连蛋白胞内抗体）的重组抗体，这些抗体可以在活神经元中表达，并以非侵入性和高保真度标记内源性靶蛋白。此外，转录调控系统控制 FingR 的表达水平，使它们的表达水平与其内源对应物完全相同，从而确保低背景。在这笔资助中，我们将培育出表达融合荧光蛋白的 FingR 的转基因小鼠，该 FingR 可识别突触蛋白。这些小鼠大脑中单个神经元的突触蛋白可以在体内实时可视化。反过来，这将允许在 分子水平与细胞、回路和整个动物水平的事件相关。特别是，通过绘制神经元中突触蛋白的位置和数量，将有可能监测活体神经元中突触输入和输出的强度。能力 监测突触强度对于研究与异常突触连接相关的疾病（包括精神分裂症、智力低下和自闭症）非常有用。