Functional Mapping of Dopamine-Dependent Fear Circuitry Through Advanced Genetic

描述（由申请人提供）：焦虑症，例如创伤后应激障碍（PTSD）被假设是由于大脑中的恐惧处理中心在创伤事件期间未能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是重要的联想恐惧学习，提高了有趣的可能性，在一个可怕的经验，该系统的失调可能是一个促成因素，在一些焦虑症的发展。与这一假设一致，我们最近发现多巴胺神经元阶段性激活的遗传破坏会损害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，很少有人知道神经回路调节，或调节相位多巴胺信号。我们的假设是，选择兴奋性输入多巴胺神经元促进阶段性激活的一个子集，这些细胞在一个可怕的经验。随后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，我们将利用涉及小鼠行为、遗传学、分子生物学、病毒介导的基因转移和自由行为小鼠中多巴胺神经元活性的体内光纤成像的多学科方法。我们正在创新一种技术，该技术将允许在小鼠巴甫洛夫恐惧条件反射期间投射到特定目标的多巴胺神经元内的实时活性依赖性钙动力学的纤维荧光显微镜，这将使我们能够生成阶段性多巴胺神经元激活的地图。此外，我们正在建立一种基于组合病毒载体的方法，用于神经元中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条件反射。最后，我们开发了一种条件基因重建的方法，这将使我们能够生成一个地图的最小必要的大脑区域需要D1R表达的恐惧条件。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。 公共卫生相关性：焦虑症，如创伤后应激障碍（PTSD），在创伤性生活经历后表现出来，是由于大脑中恐惧处理中心的失调。对于大脑中对正常恐惧处理和预防广泛性焦虑至关重要的特定连接，我们知之甚少。我们将使用病毒介导的基因转移和小鼠深部脑组织光纤成像的最新技术，精确绘制大脑中对恐惧处理至关重要的多巴胺神经递质系统的连接。通过了解多巴胺如何与大脑中的其他递质系统相互作用，我们可以开发更好的疗法来治疗这些疾病。

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. PUBLIC HEALTH RELEVANCE: Anxiety disorders, such as post-traumatic stress disorder (PTSD) that manifest following a traumatic life experience result from the disregulation of fear processing centers in the brain. Little is known about the specific connections in the brain that are critical for normal fear processing and the prevention of generalized anxiety. We will use state of the art techniques in viral-mediated gene transfer and fiber-optic imaging of deep brain tissue in mice to precisely map connections of the dopamine neurotransmitter system in the brain that are critical for fear processing. By understanding how dopamine interfaces with other transmitter systems in the brain, we can develop better therapies to treat these disorders.