Resources for Studying Neural Circuit Structure and Function with G-Deleted Rabies Viruses

研究 G 缺失狂犬病病毒神经回路结构和功能的资源

基本信息

批准号：
9130302
负责人：
EDWARD M CALLAWAY
金额：
$ 29.75万
依托单位：
SALK INSTITUTE FOR BIOLOGICAL STUDIES
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2019-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9130302
关键词：
Advertising Affect Aliquot Axon Behavior Behavioral Brain Brain region Cataloging Catalogs Cell Line Cells Chimera organism Cloning Cognition Communities Cost Sharing Dendrites Elements Ensure Experimental Designs Gated Ion Channel Gene Expression Gene Transfer Genes Genetic Engineering Genome Glycoproteins Health Helper Viruses Institutes Laboratories Lentivirus Vector Link Maps Monitor Monoclonal Antibody R24 Movement Mus Nervous System Physiology Nervous system structure Neurons Neurosciences Neurosciences Research Output Perception Play Population Production Proteins Protocols documentation Publications Rabies Rabies virus Reagent Reproducibility Research Research Personnel Resources Role Speed Structure Synapses System Technology Testing Travel Variant Viral Virus Receptors adeno-associated viral vector calcium indicator cell type cost design env Gene Products gene function genetic technology glycoprotein G improved information processing innovation interest light gated meetings memory retrieval nervous system disorder neural circuit neuronal circuitry new technology novel presynaptic presynaptic neurons promoter receptor research study response sensor targeted treatment tool uptake vector web site

项目摘要

DESCRIPTION (provided by applicant): Deciphering how neural circuits within the mammalian brain give rise to perception, cognition, and behavior is central to understanding how the nervous system functions. Neural circuits operate over a vast range of spatial and computational scales, from high-level circuits that integrate information across multiple brain regions, to microcircuits that perform simple input/output transformations within a specialized brain structure. Each level of analysis is important for formulating responses to environmental conditions. However, studying a specific neural circuit is extremely difficult, as most nervous system structures contain many types of neurons with inextricably intertwined axons and dendrites. To overcome this obstacle, the glycoprotein (G)-deleted rabies vector system was developed to identify direct synaptic inputs to a particular neuronal population. By pseudo typing the G-deleted rabies vector with a foreign envelope protein, such as EnvA, the vector selectively transduces target neurons genetically engineered to express the EnvA receptor. If these cells also express rabies glycoprotein the vector travels retrograde exactly one synaptic step and transduces direct presynaptic neurons. The rabies genome can be altered to encode any gene of interest, including fluorescent proteins to reveal the cytoarchitecture of presynaptic cells, or neuroscience tools (e.g., calcium indicators or light-gated ion channels) to monitor or manipulate circuit activity. Thus, the G-deleted rabies vector system allows the fine- scale manipulation of specific cell types within a circuit, allowing investigators to test hypotheses linking these circuts to behavior. This technology has revolutionized the study of neuronal circuits, creating high demand for these cutting-edge reagents. Laboratories that focus on understanding neural circuits, however, typically do not have the resources or expertise to produce high quality rabies vectors or associated helper vectors that are necessary to perform these experiments. Because of this, the Salk Institute's Gene Transfer, Targeting, and Therapeutics (GT3) Core, which currently generates the rabies vectors, is inundated with requests for ready-to- inject viral reagents and demand exceeds production capacity. This R24 application proposes to expand the GT3 Core's capacity for maintaining, propagating, and distributing all G-deleted rabies vector variants and helper vectors (Aim 1). The GT3 Core will also incorporate newly developed tools into the technology platform as they are innovated (Aim 2). Establishing this central rabies production facility will lower reagent costs (through economies of scale) and improve the reproducibility of study findings. Between-lab cost sharing mechanisms will enable the distribution of small aliquots, facilitating pilot experiments and removing the greatest barrier to technology uptake by new laboratories. Newly generated reagents will be immediately distributed to the neuroscience community without publication restrictions, thereby speeding the pace of discovery. These efforts will broaden the impact of this technology and ensure that neuroscientists studying circuits are equipped with the most modern analytic tools.

描述（由申请人提供）：破译哺乳动物大脑内的神经回路如何产生感知、认知和行为对于理解神经系统如何在广泛的空间和计算尺度上运作至关重要。跨多个大脑区域整合信息的级别电路区域，到在专门的大脑结构内执行简单输入/输出转换的微电路，每个级别的分析对于制定对环境条件的反应都很重要。这是极其困难的，因为大多数神经系统结构包含多种类型的神经元，这些神经元具有不可分割地交织在一起的轴突和树突，为了克服这一障碍，开发了糖蛋白（G）删除的狂犬病载体系统，以识别特定神经元群的直接突触输入。将 G 缺失的狂犬病载体与外源包膜蛋白（例如 EnvA）进行分型，该载体会选择性地转导经过基因工程改造的目标神经元，以表达 EnvA 受体。还表达狂犬病糖蛋白，该载体逆行精确一个突触步骤并转导直接突触前神经元。可以改变狂犬病基因组以编码任何感兴趣的基因，包括荧光蛋白以揭示突触前细胞的细胞结构，或神经科学工具（例如钙指示剂或光门控离子通道）可监测或操纵回路活动，因此，G 缺失的狂犬病载体系统允许对回路内的特定细胞类型进行精细操作，从而使研究人员能够测试关联的假设。这项技术彻底改变了神经回路的研究，对这些专注于理解神经回路的尖端试剂产生了很高的需求，但通常没有资源或专业知识来生产。执行这些实验所需的高质量狂犬病载体或相关辅助载体因此，目前生成狂犬病载体的索尔克研究所的基因转移、靶向和治疗 (GT3) 核心充斥着大量准备就绪的请求。 - 注入病毒试剂，需求超出生产能力。此 R24 应用建议扩大 GT3 核心的维护、传播和分发所有 G 删除的能力。 GT3 核心还将在创新时将新开发的工具纳入技术平台中（目标 2），建立这个中央狂犬病生产设施将降低试剂成本（通过规模经济）。提高研究结果的可重复性。实验室之间的成本分摊机制将能够分配小份样品，促进试点实验并消除最大的障碍。新实验室采用的技术将立即分发给神经科学界，不受出版限制，从而加快发现速度，并确保研究电路的神经科学家配备最现代的分析技术。工具。