Cell Type Specific Genomic and Functional Dissection of Fear Off Amygdala Pathways

与恐惧有关的疾病，如创伤后应激障碍（PTSD），通常以 无法抑制和消除恐惧记忆，导致恐惧相关行为的病理表达。 为了使合理设计的靶向治疗方法取得进展，需要更好地了解 需要调节恐惧抑制和消退的神经回路。这项提案利用了尖端的细胞- 针对杏仁核的特定类型的方法，通过跨越许多领域，与NIMH的研究重点保持一致 RDoC在NIH战略计划中用于识别恐惧相关疾病的病理生理学。 我们必须了解杏仁核内支持恐惧抑制的特定细胞类型的作用 害怕灭绝学习。已知基底外侧杏仁核（BLA）通过以下方式调节恐惧表达： 投射到中央杏仁核的内侧（CeM）和外侧（CeL），部分通过调制 通过杏仁核的嵌入细胞核（ITC）接收来自内侧前额叶皮层的输入 （mPFC），并充当细胞L的抑制门。我们将靶向Thy 1、FoxP 2和PKC β亚群 分别在BLA、ITC、CeL中，被认为与恐惧抑制和消退相关 在杏仁核内-这里称为“恐惧”人群。 通过多种细胞类型特异性的方法，我们将确定潜在的抑制事件， 杏仁核中的三种“恐惧消除”细胞类型。实验将从功能上和分子上鉴定 参与这些过程的BLA、ITC和CeL内的神经元群体。我们的中央 假设是BLA-ITC-CeL回路中的Fear-Off途径作为mPFC调节的 抑制门，在消除训练和保持期间促进恐惧抑制，部分通过 miRNA对可塑性的调控。针对恐惧途径的多个方面将扩大我们的 理解恐惧抑制。 这些假设将通过以下具体目标进行检验：目标1） 杏仁核恐惧细胞：为了确定从mPFC投射到BLA的差异传入调节， ITC和CeA 'Fear-Off'神经元通过直接操纵传入神经元。目的2）杏仁核内 恐惧细胞的调节：为了确定BLA，ITC和CeA恐惧神经元的确切作用， 调节杏仁核的恐惧输出回路;和目的3）恐惧细胞的miRNA调节：为了确定 miRNAs在BLA、ITC和CeA恐惧神经元中对行为、神经元功能和可塑性的作用 通过直接操纵miRNA的功能。我们使用一种组合的方法， 分子表征杏仁核神经元的特定目标群体，这将阐明重要的 杏仁核中控制恐惧消退的微电路新目标的识别将进一步 我们对恐惧行为背后的电路的理解将为治疗提供独特的途径。

Fear-related disorders such as Post-Traumatic Stress Disorder (PTSD) are often characterized by an inability to inhibit and extinguish fear memories leading to pathological expression of fear-related behaviors. For progress to occur with targeted rationally-designed therapeutic approaches, a greater understanding of the neural circuitry mediating fear inhibition and extinction is needed. This proposal utilizes cutting-edge, cell- type specific approaches targeting the amygdala to align with NIMH research priorities by cutting across many of the RDoC in the NIH strategic plan for identifying the pathophysiology of fear-related disorders. It is critical that we understand the role of specific cell types within the amygdala supporting fear inhibition and fear extinction learning. It is known that the Basolateral Amygdala (BLA) modulates fear expression via projections to the medial (CeM) and lateral division (CeL) of the central amygdala, in part through modulation via the Intercalated Cell Nuclei of the Amygdala (ITC), which receive inputs from medial prefrontal cortex (mPFC) and act as an inhibitory gate to the CeL. We will target the Thy1, FoxP2, and PKCsubpopulations within the BLA, ITC, CeL, respectively, that are proposed to be associated with fear inhibition and extinction within the amygdala – here termed the ‘Fear-Off’ populations. Through a variety of cell-type specific approaches, we will determine the events underlying the inhibition of fear within three ‘Fear-Off’ cell types in the amygdala. Experiments will functionally and molecularly identify populations of neurons within the BLA, ITC, and CeL that participate in these processes. Our central hypothesis is that the Fear-Off pathway within the BLA-ITC-CeL circuit acts as an mPFC-regulated inhibitory gate facilitating inhibition of fear during extinction training and retention, in part through miRNA regulation of plasticity. Targeting multiple aspects of the Fear-Off pathway will expand our understanding of fear inhibition. These hypotheses will be tested through the following Specific Aims: Aim 1) mPFC Regulation of Amygdala Fear-Off Cells: To determine the differential afferent regulation from mPFC projections to BLA, ITC, and CeA ‘Fear-Off’ neurons through direct manipulation of afferent neurons. Aim 2) Intra-Amygdala Regulation of Fear-Off Cells: To determine the precise role of BLA, ITC, and CeA Fear-Off neurons in regulating fear output circuitry of the amygdala; and Aim 3) miRNA Regulation of Fear-Off Cells: To determine the role of miRNAs on behavior, neuronal function and plasticity within BLA, ITC, and CeA Fear-Off neurons through direct manipulation of miRNA function. We use a combinatorial approach to functionally and molecularly characterize specific target populations of amygdala neurons, which will elucidate important microcircuitry within the amygdala governing fear extinction. The identification of novel targets will advance our understanding of circuitry underlying fear behaviors and will provide unique avenues for therapeutics.