Determining the function of TRPC6 channels in a subpopulation of VTA dopamine neurons

确定 VTA 多巴胺神经元亚群中 TRPC6 通道的功能

• 批准号: 10676673
• 负责人: Mollie Xiaoqi Bernstein
• 金额: $ 4.41万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-16 至 2025-08-15
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10676673
• 关键词: Action Potentials; Adult; Agonist; Antidepressive Agents; Bathing; Behavior; Borates; CRISPR/Cas technology; Calcium; Calcium Signaling; Calcium ion; Cells; Central Nervous System; Control Groups; Coupled; Decision Making; Depressive disorder; Dopamine; Electrophysiology (science); Exhibits; Fluorescence; G alpha q Protein; Genes; Genetic Markers; Goals; Hypericum perforatum; Hypothalamic structure; Image; Ion Channel; KISS1 gene; Learning; Link; Mental Depression; Mental disorders; Midbrain structure; Motivation; Mus; Mutate; Neurons; Neuropeptide Receptor; Neuropeptides; Pathway interactions; Physiology; Play; Population; Probability; Psychological reinforcement; Receptor Signaling; Rewards; Role; Schizophrenia; Serotonin; Signal Pathway; Signal Transduction; Slice; Stimulus; Substance Use Disorder; TACR3 gene; Testing; Therapeutic; Transgenic Mice; Ventral Tegmental Area; cell type; channel blockers; comparison control; discounting; dopaminergic neuron; experimental group; experimental study; genetic technology; hyperforin; in vivo; insight; loss of function; motivated behavior; nervous system disorder; patch clamp; receptor; reuptake

Project Summary/Abstract: Midbrain dopamine (DA)-producing neurons of the ventral tegmental area (VTA) play a critical role in modulating reward-seeking behavior. VTA-DA neurons are functionally and genetically heterogeneous, and genetic markers for neuropeptides and neuropeptide receptors can be used to isolate VTA subpopulations. Though distinct neuropeptidergic pathways have been shown to potently modulate DA neurons, the intracellular signaling pathways that act downstream of neuropeptide receptors are unknown. VTA-DA neurons that express the Gq-protein coupled receptor, tachykinin receptor 3 (Tacr3), are a minimally sufficient subpopulation of DA neurons that promote reward reinforcement behavior. Tacr3 activation has recently been shown to be dependent on transient receptor potential canonical (TRPC) channel signaling in the hypothalamus. TRPC type 6 (TRPC6) channels are enriched in DA neurons and are activated by stimulation of Gq-coupled receptor signaling. Therefore, I hypothesize that TRPC6 is likely the main type of TRPC channel in VTA-DA neurons that acts downstream of Tacr3 activation. To establish the role of TRPC6 in regulating the physiology and function of VTA-Tacr3 neurons, I propose to selectively mutate the Trpc6 gene to generate a loss of function in a cell-type specific manner within the VTA of adult mice using an advanced CRISPR/Cas9 genetic technology. I will perform ex vivo slice electrophysiology and slice calcium imaging (Aim 1), as well as in vivo recordings of calcium dynamics during a probabilistic discounting paradigm and progressive ratio motivational task (Aim 2). Determining the function of TRPC6 in VTA-Tacr3 neurons will provide important insights into the therapeutic potential of this understudied ion channel in the central nervous system.

项目摘要/摘要： 中脑腹侧被盖区 (VTA) 产生多巴胺 (DA) 的神经元在 调节寻求奖励的行为。 VTA-DA 神经元在功能和遗传上具有异质性，并且 神经肽和神经肽受体的遗传标记可用于分离 VTA 亚群。 尽管不同的神经肽能通路已被证明可以有效调节 DA 神经元， 作用于神经肽受体下游的细胞内信号传导途径尚不清楚。腹侧被盖区-多巴胺神经元 表达 Gq 蛋白偶联受体、速激肽受体 3 (Tacr3) 的药物是最低限度足够的 促进奖励强化行为的 DA 神经元亚群。 Tacr3 激活最近已 显示依赖于瞬时受体电位规范 (TRPC) 通道信号传导 下丘脑。 TRPC 6 型 (TRPC6) 通道富含 DA 神经元，并通过刺激而激活 Gq 偶联受体信号传导。因此，我假设 TRPC6 可能是 TRPC 通道的主要类型 VTA-DA 神经元在 Tacr3 激活的下游发挥作用。确定 TRPC6 在调节中的作用 由于VTA-Tacr3神经元的生理学和功能，我建议选择性地突变Trpc6基因以产生 使用先进的 CRISPR/Cas9 在成年小鼠的 VTA 内以细胞类型特异性方式丧失功能 基因技术。我将进行离体切片电生理学和切片钙成像（目标 1），以及 概率贴现范式和渐进比率期间钙动态的体内记录 激励任务（目标 2）。确定 TRPC6 在 VTA-Tacr3 神经元中的功能将提供重要的信息 深入了解这种正在研究的离子通道在中枢神经系统中的治疗潜力。