Chronic CeA CRF overexpression on gene expression in PVN CRF-expressing cells

描述(由申请人提供)：项目摘要：在许多患有严重抑郁障碍(MOD)的患者中，下丘脑-垂体-肾上腺(HPA)轴过度活跃。随着治疗的成功，这种功能障碍会正常化，这可能是治疗背后的共性。新的治疗方法应该针对HPA轴过度活动的来源：推测是下丘脑室旁(PVN)调节失调的促肾上腺皮质激素释放因子(CRF)。PVN的活性受许多边缘结构的协调调节，其中一个或多个区域的可塑性可能导致PVN CRF输出增加和随后的HPA轴功能障碍。一个可能的罪魁祸首是中央杏仁核(CEA)，这是另一个表达高浓度CRF的区域，被认为是介导应激诱导行为的区域。本研究的目的是确定(1)除行为效应外，CEA慢性增加的CRF驱动是否会产生与抑郁症状相关的内分泌变化；(2)CEA中CRF的慢性过度表达是否会导致PVN CRF神经元中与所观察到的内分泌和行为紊乱相关的基因表达变化。慢病毒载体将在成年动物CEA中CRF表达神经元(LVCRFp3.0CRF)中高表达CRF。通过创造一种表达CRF的细胞也表达绿色荧光蛋白(GFP)的转基因小鼠，可以很容易地识别表达CRF的细胞。这些细胞可以通过荧光激活的细胞分选法分离出来，并进行基因表达分析，以比较在基础条件下或在CEA中慢性过度表达CRF后，PVN和CEA中CRF和非CRF表达的细胞。预计慢性CEA CRF过度表达将导致HPA轴过度活动，这至少部分可以通过PVN CRF细胞中基因表达的变化来解释。与公共健康相关：CRF的慢性升高可能会引发一系列事件，改变基因表达和内分泌信号，最终导致抑郁症状。CRF由许多大脑结构释放，但这些区域CRF之间的关系尚不完全清楚，因为以前的技术无法模拟慢性CRF升高。利用先进的技术，CRF将在一个区域过度表达，然后分离另一个区域的CRF细胞，以确定可能与观察到的内分泌和行为变化相关的基因表达变化。

DESCRIPTION (provided by applicant): Project Summary: The hypothalamic pituitary adrenal (HPA) axis is hyperactive in many patients with major depressive disorder (MOD). This dysfunction normalizes with successful treatment and may be a commonality behind treatments. Novel treatments should target the source of HPA axis hyperactivity: presumably dysregulated corticotropin releasing factor (CRF) from the paraventricular hypothalamus (PVN). PVN activity is regulated by the coordination of numerous limbic structures and plasticity in one or more of these regions may cause enhanced PVN CRF output and subsequent HPA axis dysfunction. One likely culprit is the central amygdala (CeA), another region expressing high concentrations of CRF and thought to mediate stress- induced behavior. The Aims of the present research are to determine (1) if, in addition to behavioral effects, chronic increased CRF drive from the CeA reproduces endocrine changes associated with depressive symptoms and (2) if chronically overexpressing CRF in the CeA results in gene expression changes in PVN CRF neurons that correlate with the observed endocrine and behavioral disruptions. A lentiviral vector will overexpress CRF in CRF-expressing neurons (LVCRFp3.OCRF) in the CeA of adult animals. CRF-expressing cells will be easily identifiable by creating a transgenic mouse in which CRF- expressing cells also express green fluorescent protein (GFP). These cells can be isolated via fluorescence activated cell sorting and subjected to gene expression analysis to compare the CRF and non CRF- expressing cells in the PVN and CeA under basal conditions or after chronically overexpressing CRF in the CeA. It is expected that chronic CeA CRF overexpression will result in HPA axis hyperactivity that can be explained at least partially by gene expression changes in PVN CRF cells. Relevance to Public Health: Chronic elevations in CRF may initiate a chain of events altering gene expression and endocrine signaling and eventually leading to symptoms of depression. CRF is released by numerous brain structures but the relationship between CRF in these regions is incompletely understood because previous technology has been unable to mimic chronic CRF elevations. Using superior techniques, CRF will be overexpressed in one region and then CRF cells in another region will be isolated to identify gene-expression changes that may correlate with observed endocrine and behavioral alterations.