Viral tracing of genetically defined neural circuitry

基因定义的神经回路的病毒追踪

基本信息

批准号：
8206536
负责人：
KEVIN T BEIER
金额：
$ 1.48万
依托单位：
HARVARD MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-02-01 至 2012-05-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): My doctoral dissertation research integrates virology, genetics, and neurobiology with the aim of creating novel neural circuitry tracers. While general information flow in the retina from photon detection to action potentials through the optic nerve has been relatively well characterized, synaptic communication remains rather poorly understood. The dissection of this circuitry requires neuronal tracers that have the ability to label cells that are synaptically connected. Historically, neural tracers such as wheat germ agglutinin (WGA), nanobeads, and fluorescent dyes have aided this investigation. These reagents have the inherent limitations of dilution and questionable tracer specificity. These problems can be obviated either by creating a genetic model in which each cell in a circuit produces the tracer, or by using viruses, which are self- replicating entities. Using the murine retina, I aim to develop new tracing methods capable of labeling distinct circuit types in vivo. I will first develop a tool that permits viral Infection in small populations of a chosen cell type. Employing the specificity of the TVA/ASLV-A Interaction used by Wickersham and colleagues, a modified rabies virus containing the ASLV-A extracellular domain will be used to target specific TVA-expressing amacrine cells using this tool. The synaptic connectivity of these cells can be verified by physiological analyses. Lastly, using the rabies virus glycoprotein as a template, I will design a retrograde tracer that labels entire circuits permanently and specifically. Containing both Cre and GFP, it will have capabilities similar to viral transsynaptic tracers without the deleterious effects of metabolic alterations. This research will provide neuroscientists with a new tool for examining lineage tracing and two novel, distinct methods for tracing neuronal circuits in the CNS that will provide synaptic connectivity information in neural circuits, and will shed light on the relatively simple yet elegant circuitry of the retina. Fixing problems associated with aberrant neural circuitry, such as memory loss, blindness, and neurodeneratlon requires a fundamental understanding of the normal activites of these circuits. Therefore, by creating a tool to better dissect circuitry, my goal is to help elucidate the vastness of cognitive computation and the pathologies associated with abnormal circuitry function.

描述（由申请人提供）：我的博士学位论文研究整合了病毒学，遗传学和神经生物学，目的是创建新型的神经电路示踪剂。虽然视网膜从光子检测到通过视神经的动作电位的一般信息流的表征相对较好，但突触通信仍然相当多。该电路的解剖需要具有标记突触连接的细胞的神经元示踪剂。从历史上看，诸如小麦胚芽凝集素（WGA），纳米角和荧光染料等神经示踪剂有助于这项研究。这些试剂具有稀释和可疑的示踪特异性的固有局限性。可以通过创建一个遗传模型来消除这些问题，在该模型中，电路中的每个细胞都会产生示踪剂，或者使用自我复制实体的病毒。我使用鼠视网膜，旨在开发能够在体内标记不同电路类型的新方法。我将开发一种允许在所选细胞类型的小种群中允许病毒感染的工具。采用Wickersham及其同事使用的TVA/ASLV-A相互作用的特异性，使用该工具将使用含有ASLV-A细胞外域的改良狂犬病病毒来靶向表达特定TVA的无链藻细胞。这些细胞的突触连通性可以通过生理分析来验证。最后，使用狂犬病病毒糖蛋白作为模板，我将设计一个逆行示踪剂，该曲线示踪剂将整个电路永久和专门标记。它同时包含CRE和GFP，具有类似于病毒透射性示踪剂的能力，而没有代谢改变的有害作用。这项研究将为神经科学家提供一种新的工具，用于检查谱系跟踪，并为CNS中的神经元电路追踪两种新颖的，独特的方法，这些方法将在神经回路中提供突触连通性信息，并将阐明相对简单的视网膜电路相对简单而优雅的电路。解决与异常神经回路相关的问题，例如记忆丧失，失明和神经肾上腺素，需要对这些电路的正常活动物有基本的了解。因此，通过创建一种工具以更好地剖析电路，我的目标是帮助阐明认知计算的广泛性以及与异常电路函数相关的病理。