Functional Mapping of Dopamine-Dependent Fear Circuitry Through Advanced Genetic

描述（由申请人提供）：焦虑障碍，如创伤后应激障碍（PTSD）被假设是由于大脑中的恐惧处理中心在创伤事件中无法形成适当的联想记忆而导致的。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，这提出了一种有趣的可能性，即在恐惧经历中，该系统的失调可能是导致某些焦虑症发展的一个因素。与这一假设相一致，我们最近发现，多巴胺神经元的阶段性激活的遗传破坏损害了小鼠的巴甫洛夫恐惧条件反射，导致广泛性焦虑样行为的表现。到目前为止，关于神经回路的调节，或由阶段性多巴胺信号调节的知之甚少。我们的假设是，多巴胺神经元的选择性兴奋性输入促进了这些细胞子集在恐惧经历中的阶段性激活。随后的阶段性多巴胺释放到离散的大脑区域，与多巴胺D1受体结合，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，包括小鼠行为，遗传学，分子生物学，病毒介导的基因转移，以及自由行为小鼠多巴胺神经元活动的体内纤维成像。我们正在创新一种技术，利用纤维荧光显微镜观察小鼠巴甫洛夫恐惧条件反射过程中多巴胺神经元内钙离子动态的实时活动依赖性，从而绘制出多巴胺神经元激活的相位图。此外，我们正在建立一种基于组合病毒载体的方法，用于在神经元中投射特定基因的条件失活，以选择目标，这将使我们能够将关键输入映射到多巴胺神经元以进行恐惧调节。最后，我们已经开发了一种条件基因重组的方法，这将使我们能够生成一个最小的基本大脑区域的地图，这些区域需要D1R表达来调节恐惧。这些技术将帮助我们建立多巴胺依赖性恐惧处理的精确神经回路，并将为解剖整个大脑的行为相关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.