Trans-synaptic optogenetics: reversible temporal control of activity at defined synaptic connections

跨突触光遗传学：在定义的突触连接处活动的可逆时间控制

• 批准号: 9975126
• 负责人: Bryan Copits
• 金额: $ 17.72万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9975126
• 关键词: Address; Anatomy; Area; Behavior; Behavior Control; Biological Models; Brain; Brain region; Cell Line; Chemicals; Collaborations; Communities; Complex; Coupled; Disease; Drug Addiction; Electrophysiology (science); Engineering; Exposure to; Future; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTP-Binding Proteins; Goals; Health; In Vitro; Ligands; Light; Location; Masks; Methods; Modification; Molecular; Molecular Conformation; Neurons; Neurosciences; Nucleus Accumbens; Opsin; Optics; Output; Peptides; Physiology; Population; Prefrontal Cortex; Process; Receptor Activation; Research Personnel; Resolution; Resources; Role; Side; Signal Transduction; Slice; Source; Stream; Synapses; Synaptic Cleft; Synaptic Transmission; System; Technology; Testing; Viral; addiction; barrel cortex; base; behavioral response; design and construction; experience; experimental study; high throughput screening; imaging approach; imaging modality; insight; neural circuit; neuroregulation; neurotransmission; novel strategies; optogenetics; postsynaptic; presynaptic; protein activation; screening; synaptic inhibition; tool; tool development; trafficking; transmission process; viral gene delivery

Project Summary/Abstract The introduction of optogenetic methods for controlling activity in defined subsets of neurons has enabled new insights into the functional roles of anatomically distinct brain regions implicated in drug addiction. However, limitations with existing optogenetic tools have made it difficult to address how local connections within a given brain region enable it to integrate and process multiple inputs from other regions. Here we propose to generate a fundamentally unique type of optogenetic tool for silencing defined sets of synapses within local microcircuits, to ultimately understand how they control addiction-related behaviors. Our approach leverages the ability of presynaptic G protein coupled receptors (GPCRs) to inhibit synaptic transmission. In contrast to existing approaches that use direct optical activation of light-sensing GPCRs, our approach involves optical activation of a separate construct on the postsynaptic side of the synaptic cleft, which in turn activates the presynaptic GPCR. In our two aims, we will develop optogenetic tools capable of trans-cellular GPCR activation using high throughput assays in cell lines, and then use experiments in neurons to refine these tools to enable trans-synaptic control of GPCR activation. We will combine optogenetics with electrophysiological readouts in primary neuronal cultures and brain slices to validate their ability to reversibly inhibit synaptic transmission. Successful completion of the proposal will establish new tools with unprecedented synaptic-resolution control over neurotransmission. This technology will enable new experiments to dissect how key brain regions involved in addiction, like the nucleus accumbens, ventral tegmenetal area, and prefrontal cortex, locally integrate information received from diverse brain regions.

项目摘要/摘要 引入用于控制神经元定义子集活动活动的光遗传学方法已使新的 对与药物成瘾有关的解剖学上不同大脑区域的功能作用的见解。然而， 现有的光遗传学工具的限制使得很难解决给定的本地连接 大脑区域使其能够整合并处理来自其他地区的多个输入。在这里我们建议生成 一种从根本上独特的类型的光遗传学工具，用于沉默定义的局部微电路中的突触集， 最终了解他们如何控制与成瘾相关的行为。我们的方法利用了 突触前G蛋白偶联受体（GPCR）抑制突触传播。与现有 使用光感应GPCR的直接光学激活的方法，我们的方法涉及光学激活 突触裂缝的突触后侧的单独构造，进而激活突触前GPCR。 在我们的两个目标中，我们将开发能够使用高的跨细胞GPCR激活的光遗传学工具 细胞系中的吞吐量测定，然后使用神经元中的实验来完善这些工具以启用反式突触 控制GPCR激活。我们将在原发性神经元中将光遗传学与电生理学读数结合 培养和大脑切片以验证其可逆抑制突触传播的能力。成功完成 该提案将建立对神经传递前所未有的突触分辨率控制的新工具。 这项技术将使新的实验能够剖析成瘾中的关键大脑区域如何参与 伏隔核，腹侧tegmenetal区域和前额叶皮层，局部整合从中收到的信息 各种大脑区域。