TRPC-mediated control of anxiety

 描述(申请人提供)：驱动焦虑样行为的突触、神经元和神经网络机制还不完全清楚。瞬时受体电位通道(TRPC)属于TRPC1/4/5亚家族，广泛分布于杏仁核和终纹床核(BNST)等参与控制先天恐惧反应的脑区。与这种表达模式一致，我们最近发现，属于这个亚家族的特定TRPC亚基可能参与了焦虑机制，可能通过激活Gaq/11蛋白偶联受体(特别是I组mGluR和CCK2受体)来调节大脑特定神经回路中的信号流。然而，TRPC4和TRPC5亚基(TRPC1/4/5亚家族的成员)既可以与TRPC1亚基形成同聚体通道，也可以与TRPC1亚基异构化，从而形成具有不同生物物理和功能特性的异四聚体TRPC通道。这些TRPC可以不同地调节神经元功能，并可能调节焦虑状态，这取决于它们的亚单位组成。结合遗传学、行为学、电生理学和光遗传学工具，我们将探索不同的TRPC亚单位及其组合在焦虑回路的结构组件-杏仁核和BNST-中的功能作用以及它们对控制焦虑样行为反应的贡献。在我们的实验中，我们将专注于缺乏TRPC1、TRPC4和TRPC5(及其组合)亚单位的小鼠的行为和功能分析。我们还将研究TrpC的小鼠，在这些小鼠中，TRPC亚单位将被特定地在杏仁核或BNST的亚区消融。在目标1中，我们将通过检测缺乏不同TRPC亚单位组合的小鼠的焦虑水平，来探讨TRPC通道亚家族TRPC1/4/5在焦虑样行为中的作用。我们还将研究不同TRPC的消融对杏仁核内投射的突触功能、Gaq/11蛋白介导的信号转导和神经元棘波输出的影响。然后，我们将探索TRPC通道在杏仁核局部神经元回路功能中的作用(目标2)。具体地说，我们将检查杏仁核区间投射中兴奋和抑制之间的平衡是否会影响突变小鼠的神经元和中间神经元。最后(目标3)，我们将研究TRPC通道在BNST的突触和神经元功能调节中的作用，重点是它的两个不同的亚区，即椭圆形核和背侧BNST，这两个亚区对焦虑起相反的作用。我们将使用光遗传学工具来实现BLA中产生的并终止于BNST的纤维的投射特异性激活。我们假设，TRPC1/4/5亚基的消融可能改变了杏仁核-BNST回路中的信号流，导致adBNST的激活增强，导致焦虑样反应减弱，因为adBNST的激活被证明是抗焦虑的。这些研究阐明了TRPC通道在焦虑相关行为过程中的作用，可能有助于我们更好地理解焦虑是如何在相应大脑回路的相互作用组件的水平上进行调节的。

 DESCRIPTION (provided by applicant): The synaptic, neuronal and neural network mechanisms driving anxiety-like behaviors are not completely understood. Transient Receptor Potential channels (TRPC), belonging to the TRPC1/4/5 subfamily, are abundant in the brain areas implicated in control of innate fear responses, including the amygdala and the bed nucleus of the stria terminalis (BNST). Consistent with this expression pattern, we found recently that specific TRPC subunits belonging to this subfamily may contribute to anxiety mechanisms, possibly modulating the signal flow within specific neural circuits in the brain through activation of Gaq/11 protein-coupled receptors (specifically, Group I mGluR and CCK2 receptors). However, TRPC4 and TRPC5 subunits (members of the TRPC1/4/5 subfamily) can either form homomeric channels or heteromerize with the TRPC1 subunit, resulting in heterotetrameric TRPC channels with distinct biophysical and functional properties. These TRPCs can differentially regulate neuronal functions and, possibly, anxiety states, depending on their subunit composition. Combining genetic, behavioral, electrophysiological and optogenetic tools, we will explore the functional roles of distinct TRPC subunits and their combinations in structural components of the anxiety circuitry-the amygdala and BNST-and their contribution to control of anxiety-like behavioral responses. In our experiments, we will focus on the behavioral and functional analysis of mice lacking TRPC1, TRPC4 and TRPC5 (and combinations thereof) subunits. We will also study TrpC floxed mice in which TRPC subunits will be ablated specifically in subdivisions of the amygdala or BNST. In Aim 1, we will explore the roles of TRPC1/4/5 subfamily of TRPC channels in anxiety-like behaviors by probing anxiety levels in mice lacking different combinations of TRPC subunits. We will also investigate the effect of different TRPCs' ablation on synaptic functions, Gaq/11 protein-mediated signaling and neuronal spike output in intra-amygdala projections. We will then explore the role of TRPC channels in the function of local neuronal circuits in the amygdala (Aim 2). Specifically, we will examine whether the balance between excitation and inhibition in interdivisional projections in the amygdala is affected in neurons and interneurons in mutant mice. Finally (Aim 3), we will investigate the role of TRPC channels in regulation of synaptic and neuronal functions in the BNST, focusing on its two different subregions, the oval nucleus and anterodorsal BNST, known to exert opposing effects on anxiety. We will use optogenetic tools to achieve projection-specific activation of the fibers arising in the BLA and terminating in the BNST. We hypothesize that the ablation of TRPC1/4/5 subunits may modify the signal flow in the amygdala-BNST circuits, leading to the enhanced activation of adBNST and resulting in diminished anxiety-like responses, as adBNST activation was shown to be anxiolytic. These studies, illuminating the role of TRPC channels in anxiety-related behavioral processes, may contribute to our better understanding of how anxiety is regulated at the level of interacting components of the corresponding brain circuits.