Trans-synaptic optical control of user-defined synaptic connections

用户定义的突触连接的跨突触光学控制

基本信息

批准号：
10732081
负责人：
Bryan Copits
金额：
$ 167.41万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-19 至 2026-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10732081
关键词：
Acute Axon Behavior Behavioral Binding Biological Assay Biosensor Brain Chemicals Collaborations Communities Complement Coupled Darkness Deposition Disease Dynorphins Endorphins Engineering Enkephalins Esthesia Exposure to G-Protein-Coupled Receptors GTP-Binding Proteins Goals Human Image In Vitro Knowledge Learning Ligands Light Masks Memory Methods Modification Molecular Molecular Conformation Mus Neurons Neurosciences Opioid Receptor Opsin Optics Oxygen Pathologic Peptide Receptor Peptides Perception Pharmacology Physiology Plants Play Population Presynaptic Terminals Process Regional Anatomy Research Research Personnel Resources Role Slice Source Synapses Synaptic Cleft System Technology Testing Validation Work brain cell candidate validation cell type cellular imaging design and construction experimental study imaging approach in vitro testing in vivo in vivo evaluation information processing insight lead candidate mu opioid receptors neural circuit neurotransmission neurotransmitter release novel strategies optogenetics phototropin postsynaptic postsynaptic neurons presynaptic protein protein interaction screening synaptic inhibition tool tool development trafficking viral gene delivery voltage

项目摘要

Abstract The human brain is estimated to contain over 100 billion neurons that are wired together by more than 100 trillion synapses. These synaptic connections increase the computational capabilities of neural circuits and are essential for sensation, perception, learning and memory, and the selection and expression of distinct behavioral states. While many tools now exist to activate, inhibit, or modulate specific cell types in the brain, none are currently capable of manipulating activity between user-defined pre- and postsynaptic cell types. As a result, our knowledge of the roles played by specific synaptic connections and how they contribute to information processing and behavior is still quite limited. Here we propose a fundamentally unique approach to optically control the activity of user-defined synaptic connections between specific cell types. We will develop a two-component system to enable optically reversible, synapse-selective inhibition using trans-synaptic activation of presynaptic inhibitory GPCRs (trans-OptoGi). In Aim 1, we will use high-throughput GPCR screens to engineer light-sensitive ligand: GPCR pairs not expressed in the mammalian brain. We will functionally validate these candidates using trans-cellular imaging assays, optimize their neuronal trafficking, test their activity at both excitatory and inhibitory synaptic connections in acute brain slices, and validate their efficacy in vivo to modulate mouse behavior. In Aim 2, we will build upon this approach to optically control the activity of endogenous presynaptic GPCRs in the brain using similar validation steps. Importantly, these genetically-encoded trans-synaptic tools will use common adeno-associated viral (AAV) gene delivery methods and light sources currently used for standard optogenetic experiments in many neuroscience labs. All constructs generated will be freely shared with the research community. Successful development of these tools will enable reversible, synapse-specific manipulations, which are noticeably lacking in the current neuroscience toolbox.

抽象的据估计，人类大脑包含超过 1000 亿个神经元，这些神经元由超过 100 万亿个突触连接在一起。这些突触连接增加了神经回路的计算能力，对于感觉、感知、学习和记忆以及不同行为状态的选择和表达至关重要。虽然现在存在许多工具来激活、抑制或调节大脑中的特定细胞类型，但目前没有一种工具能够操纵用户定义的突触前细胞类型和突触后细胞类型之间的活动。因此，我们对特定突触连接所发挥的作用以及它们如何促进信息处理和行为的了解仍然相当有限。在这里，我们提出了一种根本上独特的方法来光学控制特定细胞类型之间用户定义的突触连接的活动。我们将开发一种双组分系统，通过突触前抑制性 GPCR (trans-OptoGi) 的跨突触激活来实现光学可逆、突触选择性抑制。在目标 1 中，我们将使用高通量 GPCR 筛选来设计光敏配体：哺乳动物大脑中不表达的 GPCR 对。我们将使用跨细胞成像分析对这些候选药物进行功能验证，优化它们的神经元运输，测试它们在急性脑切片中兴奋性和抑制性突触连接的活性，并验证它们在体内调节小鼠行为的功效。在目标 2 中，我们将基于这种方法，使用类似的验证步骤光学控制大脑中内源性突触前 GPCR 的活动。重要的是，这些基因编码的跨突触工具将使用常见的腺相关病毒（AAV）基因传递方法和目前用于许多神经科学实验室标准光遗传学实验的光源。生成的所有结构都将与研究界免费共享。这些工具的成功开发将实现可逆的、突触特异性的操作，而这在当前的神经科学工具箱中明显缺乏。