Functional Mapping of Dopamine-Dependent Fear Circuitry Through Advanced Genetic

描述(申请人提供)：焦虑症，如创伤后应激障碍(PTSD)，被认为是由于大脑中的恐惧处理中心在创伤事件期间未能形成适当的联想记忆所致。新出现的证据表明，多巴胺神经递质系统对联想恐惧学习很重要，这增加了一种有趣的可能性，即在恐惧经历中，这一系统的失调可能是导致某些焦虑症发展的一个因素。与这一假说一致，我们最近发现，多巴胺神经元相激活的遗传干扰会损害小鼠的巴甫洛夫恐惧条件反射，导致普遍焦虑样行为的表现。到目前为止，对神经回路的调节或由相多巴胺信号调节的情况知之甚少。我们的假设是，在恐惧经历中，对多巴胺神经元的选择性兴奋性输入有助于这些细胞的一个子集的相态激活。随后，多巴胺释放到离散的大脑区域，参与多巴胺D1受体，以促进与恐惧事件相关的记忆的形成。为了验证这一假设，我们将利用多学科方法，涉及小鼠行为、遗传学、分子生物学、病毒介导的基因转移，以及自由行为小鼠的多巴胺神经元活动的体内光纤成像。我们正在创新一项技术，该技术将允许对巴甫洛夫恐惧条件反射过程中投射到特定目标的多巴胺神经元内活动依赖的钙动力学进行纤维荧光显微镜观察，这将使我们能够生成多巴胺神经元激活的相图。此外，我们正在建立一种基于组合病毒载体的方法，用于有条件地灭活神经元中的特定基因，以选择目标，使我们能够将关键的输入映射到多巴胺神经元，以进行恐惧条件反射。最后，我们开发了一种条件性基因重建的方法，它将允许我们生成一张地图，显示需要D1R表达的最小大脑关键区域，以满足恐惧条件反射。综合起来，这些技术将帮助我们建立多巴胺依赖恐惧处理的精确神经回路，并将为解剖整个大脑中与行为相关的回路提供广泛有用的工具。

DESCRIPTION (provided by applicant): Anxiety disorders, such as post-traumatic stress disorder (PTSD) are hypothesized to result from a failure of fear processing centers in the brain to form appropriate associative memories during a traumatic event. Emerging evidence suggests that the dopamine neurotransmitter system is important for associative fear learning, raising the intriguing possibility that disregulation of this system during a fearful experience could be a contributing factor in the development of some anxiety disorders. Consistent with this hypothesis, we recently discovered that genetic disruption of the phasic activation of dopamine neurons impairs Pavlovian fear conditioning in mice, resulting the manifestation of generalized anxiety- like behavior. To date, very little is known about the neural circuitry regulating, or regulated by phasic dopamine signaling. Our hypothesis is that a select excitatory input to dopamine neurons facilitates the phasic activation of a subset of these cells during a fearful experience. Subsequent phasic dopamine release into discrete brain regions engages the dopamine D1 receptor to facilitates the formation of memories related to the fearful event. To test this hypothesis, we will utilize a multidisciplinary approach involving mouse behavior, genetics, molecular biology, viral-mediated gene transfer, and in vivo fiber- optic imaging of dopamine neuron activity in freely behaving mice. We are innovating a technique that will allow for fibered fluorescence microscopy of real-time activity-dependent calcium dynamics within dopamine neurons projecting to specific targets during Pavlovian fear conditioning in mice that will allow us to generate a map of phasic dopamine neuron activation. Additionally, we are establishing a combinatorial viral vector based approach for the conditional inactivation of specific genes in neurons projecting to select targets that will allow us to map the critical inputs to dopamine neurons for fear conditioning. Finally, we have developed a method for conditional gene reconstitution that will allow us to generate a map of the minimal essential brain regions requiring D1R expression for fear conditioning. Together these techniques will help us to establish the precise neural circuitry of dopamine-dependent fear processing and will provide broadly useful tools for the dissection of behaviorally relevant circuits throughout the brain.