The role of BNST CGRP in stress-enhanced anxiety behavior and HPA-axis function

描述(由申请人提供)：暴露在引起恐惧的压力源下的个人患上一些精神疾病的风险增加，包括创伤后应激障碍、严重抑郁障碍和广泛性焦虑障碍。这些令人衰弱的疾病是当今美国面临的最严重的公共卫生问题之一。了解中枢神经药理学和分子机制，驱动对恐惧引起的应激源的反应，将使我们能够识别可能导致应激相关障碍发展的调节失调点，并为开发有效的治疗方案确定靶点。终纹床核是最近出现的应激调节回路的重要组成部分，它含有密集的终末，释放神经肽降钙素基因相关肽(CGRP)。最近，行为证据表明BNST内的CGRP信号与焦虑有关，也表明BNST CGRP可能调节对应激的行为和神经内分泌反应。因此，这项研究的第一个目标是检验BNST内的CGRP信号增强焦虑样行为和下丘脑-垂体-肾上腺(HPA)轴对应激反应的假设。我将通过1)将CGRP拮抗剂1CGRP8-37通过颅内小管注入BNST来操纵BNST CGRP信号；2)通过慢病毒介导的RAMP1的过度表达来敏化BNST CGRP受体。RAMP1是CGRP的受体亚单位，赋予CGRP药理学特异性，并影响CGRP受体的糖基化和向细胞表面的运输。在操纵BNST信号之后，大鼠将暴露在一种引起恐惧的应激源(三甲基噻唑啉气味)中。我们将测量恐惧和焦虑样行为(声音惊吓和防御性冻结)，神经内分泌反应(血清皮质酮和促肾上腺皮质激素)，以及CGRP、CGRP受体成分RAMP1和降钙素受体样受体(CRLR)mRNA表达的变化。此外，由于BNST内的CGRP终末在含有应激相关神经肽促肾上腺皮质激素释放因子(CRF)的神经元周围形成体周篮子，本研究的另一个目标是确定CGRP信号是否影响BNST CRF神经元的激活，从而影响焦虑行为和HPA轴活动。为了验证BNST CGRP增强BNST CRF神经元激活从而增加应激相关靶结构中CRF1受体信号的假设，大鼠将被全身CRF1拮抗剂(GSK876008)治疗或感染慢病毒介导的CRF siRNA以下调BNST内CRF的表达，然后它们将被注入BNST内CGRP并进行如上所述的测试。这些研究结果可能为BNST CGRP对行为和神经内分泌应激反应的调节以及对应激相关神经递质信号通路的影响提供第一批证据。这项研究还可能为治疗应激相关精神障碍提供一个新的治疗靶点。 公共卫生相关性：暴露在引起恐惧的压力源下的个人患上一些精神疾病的风险增加，包括创伤后应激障碍、严重抑郁障碍和广泛性焦虑障碍。终纹床核(BNST)含有大量的降钙素基因相关肽(CGRP)免疫阳性终末，最近被认为是皮质信息向神经结构传递的关键中继器，介导对应激的行为和神经内分泌反应。这项拟议的研究将探讨BNST内CGRP信号在恐惧和焦虑样行为和神经内分泌应激反应中的作用，并可能为应激相关精神病理的治疗提供一个新的治疗靶点。

DESCRIPTION (provided by applicant): Individuals exposed to fear-evoking stressors are at increased risk for a number of psychiatric illnesses including post-traumatic stress disorder, major depressive disorder, and generalized anxiety disorder. These debilitating conditions are among the most significant public health problems facing the United States today. Understanding the central neuropharmacological and molecular mechanisms that drive responses to fear- evoking stressors will enable us to identify points of dysregulation that may contribute to development of stress-related disorders and also identify targets for the development of effective treatment options. The bed nucleus of the stria terminalis, which has recently emerged as an important component of stress regulation circuitry, contains dense terminals that release the neuropeptide calcitonin gene-related peptide (CGRP). Very recently, behavioral evidence has implicated CGRP signaling within the BNST in anxiety and also points to the possibility that BNST CGRP may modulate behavioral and neuroendocrine responses to stress. The first goal of this present research, then, is to test the hypothesis that CGRP signaling within the BNST enhances anxiety-like behavioral and hypothalamic-pituitary-adrenal (HPA) axis responses to stress. I will manipulate BNST CGRP signaling using 1) infusion of a CGRP antagonist, 1CGRP8-37 into the BNST via intracranial cannulae and 2) BNST CGRP receptor sensitization by lentiviral-mediated over-expression of RAMP1, the CGRP receptor subunit that confers pharmacological specificity for CGRP and influences glycosylation and trafficking of the CGRP receptor to the cell surface. Following manipulation of BNST signaling, rats will be exposed to a fear-evoking stressor (trimethlythiazoline odor). We will measure fear- and anxiety-like behavior (acoustic startle and defensive freezing), neuroendocrine responses (serum corticosterone and adrenocorticotropin hormone), and changes in mRNA expression of CGRP, and the CGRP receptor components RAMP1 and calcitonin receptor-like receptor (CRLR). Also, because CGRP terminals within the BNST form perisomatic baskets around neurons containing the stress-related neuropeptide corticotropin- releasing factor (CRF), another goal of this research is to determine if CGRP signaling affects activation of BNST CRF-containing neurons to influence anxiety behavior and HPA axis activity. In order to test the hypothesis that BNST CGRP enhances BNST CRF neuron activation to increase CRFr1 receptor signaling in stress-related target structures, rats will be treated with a systemic CRFr1 antagonist (GSK876008) or infected with lentiviral-mediated CRF siRNA to knock down CRF expression within the BNST and then they will be infused with intra-BNST CGRP and tested as described above. The results of these studies could provide the first pieces of evidence to demonstrate BNST CGRP modulation of behavioral and neuroendocrine stress responses and influence on stress-related neurotransmitter signaling pathways. This research may also present a novel therapeutic target in the treatment of stress-related psychiatric disorders. PUBLIC HEALTH RELEVANCE: Individuals exposed to fear-evoking stressors are at increased risk for a number of psychiatric illnesses including post-traumatic stress disorder, major depressive disorder, and generalized anxiety disorder. The bed nucleus of the stria terminalis (BNST), which contains numerous calcitonin gene-related peptide (CGRP) immunopositive terminals, has recently been identified as a pivotal relay of cortical information to neural structures mediating behavioral and neuroendocrine responses to stress. The proposed research will investigate the role of CGRP signaling within the BNST in fear- and anxiety-like behavioral and neuroendocrine stress responses, and may present a novel therapeutic target in the treatment of stress-related psychopathologies.