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.

项目概要/摘要： 中脑腹侧被盖区（VTA）的多巴胺（DA）产生神经元在脑内多巴胺代谢中起着关键作用。 调节奖赏寻求行为。腹侧被盖区DA神经元在功能和遗传上是异质的， 神经肽和神经肽受体的遗传标记可用于分离VTA亚群。 虽然不同的神经肽能通路已被证明可以有效地调节DA神经元， 作用于神经肽受体下游的细胞内信号传导途径是未知的。腹侧被盖区多巴胺能神经元 表达Gq蛋白偶联受体速激肽受体3（Tacr 3）的细胞，是最低限度的足够的细胞。 DA神经元的亚群，促进奖励强化行为。最近，Tacr 3激活被 显示依赖于瞬时受体电位典型（TRPC）通道信号传导， 下丘脑TRPC 6型（TRPC 6）通道在DA神经元中富集，并通过刺激 Gq偶联受体信号传导。因此，我推测TRPC 6可能是TRPC通道的主要类型。 VTA-DA神经元，其作用于Tacr 3激活的下游。确定TRPC 6在调节 VTA-Tacr 3神经元的生理学和功能，我建议选择性地突变Trpc 6基因，以产生一个新的VTA-Tacr 3神经元。 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（Aim 1），以及 在概率贴现范例和累进比率期间钙动力学的体内记录 动机任务（目标2）。确定TRPC 6在腹侧被盖区-Tacr 3神经元中的功能将提供重要的 深入了解中枢神经系统中这种未充分研究的离子通道的治疗潜力。