Engineered viral tropism for cell-type specific manipulation of neuronal circuits

用于神经元回路的细胞类型特异性操作的工程病毒趋向性

• 批准号: 9034297
• 负责人: Daniel Schmidt
• 金额: $ 35.73万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-24 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9034297
• 关键词: Adopted; Adverse effects; Base of the Brain; Behavior; Benchmarking; Biological Assay; Brain; Capsid Proteins; Categories; Cell surface; Cells; Cloning; Cognition; Color; Communities; Corpus striatum structure; Dependovirus; Development; Disease; Engineering; Generations; Generic Drugs; Glutamates; Goals; Infection; Ion Channel; Light; Maps; Methods; Molecular; Molecular Conformation; Molecular Profiling; Neurons; Neurosciences; Outcome; Pathology; Peptides; Performance; Pharmaceutical Preparations; Protein Engineering; Proteomics; Reagent; Reporter; Resolution; Resources; Rewards; Role; Signal Transduction; Slice; Sorting - Cell Movement; Specificity; Split Genes; Stimulus; Surface; Synapses; Synaptic plasticity; System; Technology; Toxin; Training; Transgenes; Transgenic Animals; Tropism; Viral; Virus; Work; addiction; brain tissue; cell type; combinatorial; design; gene complementation; genetic manipulation; improved; in vivo; in vivo Model; interest; knock-down; neural circuit; neuromechanism; neuronal circuitry; novel; optogenetics; promoter; public health relevance; receptor; receptor expression; relating to nervous system; response; selective expression; sensor; tool; transgene expression; viral gene delivery; voltage

 DESCRIPTION (provided by applicant): It is a longstanding goal in neuroscience to reveal how specific cell types contribute to different neural circuits that underlie cognition, behavior, and disease pathology. Although cell types can be grouped into descriptive categories (excitatory, inhibitory, peptidergic etc.), we know there is a great combinatorial diversity of cels that differ in ion channel and receptor expression levels and fulfill discrete roles within neural circuits. Thus, to improve the resolution of neural circuit maps, to understand how the brain works on a mechanistic level, and to better understand disease pathologies there is a great need for manipulating ever more specific sets of cell in neural circuits. Genetically targeting these different subsets is difficult when delivering transgenes to many neurons (with potentially adverse effects) and relying on cell-type specific promoters for selective expression - the current state of the art. Our agenda is to fundamentally change how cell type specific genetic manipulation is achieved: Since the functional definition of a neuron - its electrophysiological response to a stimulus - is intrinsically a proteomic problem, we propose a novel viral delivery method able to deliver transgenes selectively to neurons that express, on the cell surface, a targeted set of ion channels and receptors. When using this novel method transgene expression can be driven from generic and reliable promoters or other engineered promoter systems (e.g. sensitive to light or drugs). To achieve this transformative goal of a broadly useful tool for in vvo viral gene delivery, we build on Dr. Schmidt's expertise in protein engineering using genetically encoded peptide toxins, and Dr. Thomas' expertise with in vivo models of addiction disorders. In this application we describe the development of a generalizable method for creating engineered viruses with user-selectable tropism that can target specific subsets of neuronal cell types. We furthermore propose to demonstrate utility of these engineered viruses in intact brain tissue, including optogenetically targeting - without relying on transgenic animals or specific promoters - two sets of neurons involved in reward-related synaptic plasticity. The outcome of this work will be a broadly useful and first-in-class viral delivery technology that enables the genetic manipulation of defined sets neuron types in the brain based on what surface receptors they express. This method will enable completely new ways of exploring molecular and cellular mechanism of neural activity.

 描述(申请人提供)：揭示特定细胞类型如何对构成认知、行为和疾病病理基础的不同神经回路做出贡献是神经科学的一个长期目标。虽然细胞类型可以分为兴奋性、抑制性、多肽能性等，但我们知道有很大的组合多样性的CEL，它们在离子通道和受体的表达水平上存在差异，并在神经回路中发挥不同的作用。因此，为了提高神经回路图的分辨率，了解大脑是如何在机械水平上工作的，以及更好地了解疾病病理，非常有必要操纵神经回路中更特定的细胞组。当将转基因传递给许多神经元(具有潜在的不利影响)并依赖细胞类型的特定启动子进行选择性表达时，以这些不同的亚组为基因靶点是困难的--这是目前的技术状态。我们的议程是从根本上改变细胞类型特定的基因操作的实现方式：由于神经元的功能定义-其对刺激的电生理反应-本质上是一个蛋白质组问题，我们提出了一种新的病毒递送方法，能够选择性地将转基因递送到在细胞表面表达一组靶向离子通道和受体的神经元。当使用这种新的方法时，转基因表达可以从通用的和可靠的启动子或其他工程化的启动子系统(例如，对光或药物敏感)驱动。为了实现这一变革性的目标--成为体内病毒基因传递的广泛有用的工具，我们建立了施密特博士在使用基因编码的多肽毒素进行蛋白质工程方面的专业知识，以及托马斯博士在成瘾障碍体内模型方面的专业知识。在这项申请中，我们描述了一种可推广的方法，用于创建具有用户可选取向的工程病毒，该病毒可以针对特定的神经细胞类型子集。此外，我们还建议证明这些工程病毒在完整的脑组织中的实用性，包括光基因靶向--而不依赖转基因动物或特定的启动子--参与奖赏相关突触可塑性的两组神经元。这项工作的结果将是一项广泛有用和一流的病毒传递技术，该技术能够根据大脑中定义的神经元类型表达的表面受体对其进行基因操作。这种方法将为探索神经活动的分子和细胞机制提供全新的途径。