Genetically engineered anterograde monosynaptic viral tracers for multi-species neural circuit analysis

用于多物种神经回路分析的基因工程顺行单突触病毒示踪剂

• 批准号: 9795116
• 负责人: Gregory D Horwitz
• 金额: $ 433.27万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-15 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9795116
• 关键词: 3-Dimensional; Achievement; Address; Anatomy; BRAIN initiative; Bacterial Artificial Chromosomes; Brain; Brain region; Cells; Code; Communities; Data; Dependovirus; Detection; Development; Dimensions; Effectiveness; Excision; Exhibits; Gene Expression; Generations; Genes; Genetic; Genetic Engineering; Genome; Glycoproteins; Goals; Herpesvirus 1; Hippocampus (Brain); Hypothalamic structure; Immediate-Early Genes; In Vitro; Label; Learning; Memory; Methodology; Microfluidics; Modification; Molecular; Monkeys; Mus; Neurons; Neurosciences; Pathway interactions; Performance; Physiological; Primates; Process; Property; Proteins; Publishing; Rabies; Rattus; Recombinants; Research; Research Personnel; Resource Sharing; Rodent; Series; Services; Signal Transduction; Simplexvirus; Specificity; Stress; Synapses; System; TK Gene; Testing; Thymidine Kinase; Time; Toxic effect; Tracer; Variant; Viral; Viral Vector; Virulence; Virus; Virus Replication; Visual system structure; Work; base; cell type; cytotoxicity; design; improved; interdisciplinary collaboration; mutant; neural circuit; neural network; nonhuman primate; novel; optical imaging; postsynaptic neurons; presynaptic neurons; recombinase; relating to nervous system; sound; success; tool; tool development; transmission process; vector; virus genetics

Project Summary The development of trans-synaptic viral tracers is an important component of the BRAIN Initiative. At present, the lack of viral-based anterograde monosynaptic tracing tools with high signal strength and low toxicity is a gap in neuroscience. Herpes simplex virus (HSV) type 1 strain 129 (H129) is the most promising viral tool for anterograde neuronal tracing. However, current versions of genetically modified H129 viruses are limited by high virulence and toxicity, weak label signals that require immunostaining for detection, and time-dependent spread across multiple synapses. There is also a concern of the directional specificity of anterograde propagation of H129 recombinants, as they may propagate retrogradely. Investigators in the field have been working actively to develop improved versions of anterograde viral tracers, but progress has been limited. We have formed a strong interdisciplinary collaborative team composed of virologists and systems neuroscientists to develop anterograde monosynaptic recombinant H129 tracers with high signal strength and little or no toxicity for multi-species neural circuit analysis. Our published work and preliminary data establish the feasibility and key methodologies for the proposed research. We will capitalize on our established bacterial artificial chromosome (BAC) based system for rapid generation of recombinant H129 vectors and precise control of the H129 payload. We have a sound plan to reduce viral toxicity, enhance label signals and generate variants carrying different functional payloads. Our overall goal is to create a new set of safe, effective and validated anterograde-directed viral vectors that allow efficient labeling in monosynaptic projection targets of specific neuron types. These new tools will have a broad impact by enabling optical imaging, physiological recording, and activity manipulation of defined anterograde projection networks. For rapid resource sharing, we will create a service platform through the UCI Center for Virus Research to disseminate the new molecular tools to the neuroscience community.

项目概要 跨突触病毒示踪剂的开发是 BRAIN Initiative 的重要组成部分。现在， 缺乏具有高信号强度和低毒性的基于病毒的顺行单突触追踪工具是一个差距 在神经科学中。单纯疱疹病毒 (HSV) 1 型 129 株 (H129) 是最有前途的病毒工具 顺行神经元追踪。然而，当前版本的转基因 H129 病毒受到以下限制： 毒力和毒性高，标记信号弱，需要免疫染色才能检测，并且具有时间依赖性 分布在多个突触上。人们还担心顺行的方向特异性 H129 重组体的繁殖，因为它们可能逆行繁殖。现场调查人员已 积极致力于开发改进版本的顺行病毒示踪剂，但进展有限。我们 组建了由病毒学家和系统神经科学家组成的强大的跨学科协作团队 开发具有高信号强度且毒性很小或无毒性的顺行单突触重组 H129 示踪剂 用于多物种神经回路分析。我们发表的工作和初步数据确立了可行性和 拟议研究的关键方法。我们将利用我们已建立的细菌人工 基于染色体 (BAC) 的系统，用于快速生成重组 H129 载体并精确控制 H129有效载荷。我们有一个健全的计划来降低病毒毒性、增强标签信号并产生变体 携带不同的功能有效载荷。我们的总体目标是创建一套新的安全、有效和经过验证的 顺行定向病毒载体，可有效标记特定的单突触投射目标 神经元类型。这些新工具将通过实现光学成像、生理记录、 和定义的顺行投影网络的活动操纵。为了资源的快速共享，我们将创建 通过 UCI 病毒研究中心建立一个服务平台，向全世界传播新的分子工具 神经科学界。