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.

项目概要/摘要 用于控制特定神经元亚群活动的光遗传学方法的引入使得新的方法成为可能。 深入了解与毒瘾有关的解剖学上不同的大脑区域的功能作用。然而， 现有光遗传学工具的局限性使得很难解决给定的局部连接如何 大脑区域使其能够整合和处理来自其他区域的多个输入。这里我们建议生成 一种根本上独特的光遗传学工具，用于沉默局部微电路内定义的突触集， 最终了解他们如何控制与成瘾相关的行为。我们的方法利用了以下能力 突触前 G 蛋白偶联受体 (GPCR) 抑制突触传递。与现有的相比 使用直接光学激活光敏 GPCR 的方法，我们的方法涉及光学激活 突触间隙突触后侧的一个单独的结构，它反过来激活突触前 GPCR。 在我们的两个目标中，我们将开发能够使用高通量跨细胞 GPCR 激活的光遗传学工具。 在细胞系中进行吞吐量测定，然后使用神经元实验来完善这些工具，以实现跨突触 控制 GPCR 的激活。我们将把光遗传学与初级神经元的电生理读数结合起来 培养物和脑切片以验证其可逆抑制突触传递的能力。顺利完成 该提案的一部分将建立新的工具，对神经传递进行前所未有的突触分辨率控制。 这项技术将使新的实验能够剖析与成瘾有关的关键大脑区域，例如 伏隔核、腹侧被盖区和前额皮质，局部整合从 不同的大脑区域。