Neurobiology of loss: role of microglia

项目摘要 当一个人被剥夺了一些有价值的东西，如社会关系、身体上的东西时，就会发生心理损失 健康，或金融稳定。这些经历通常会导致类似抑郁的症状，如运动障碍， 社交退缩、体重增加和不活跃的压力反应。虽然损失的经历很常见，但很少有 了解导致丢失症状的分子机制。我们实验室使用的是浓缩法。 (呃)模拟大鼠的失落体验。在这个方案中，老鼠首先被安置在积极的，高度 刺激环境浓缩(EE)一个月，然后搬到单独的住房，不再进一步 刺激物。这种移除改变了老鼠对环境的反应方式，产生了 特别类似于人类的减重，包括被动应对增加、体重增加和缺乏活力的压力 回应。我们使用了一种创新的基于发现的多组学方法来研究ER分子 杏仁基底外侧核(BLA)的机制，这是一个传统上不同的压力促进区域 在EE和ER中激活。利用RNA测序、蛋白质组学和蛋白质组学的组合，我们发现了一种 内质网独特的小胶质细胞表型。我们假设内质网减少了BLA小胶质细胞的信号，损害了 它们能够调节附近的神经元并抑制促进应激的血乳酸信号。这种失调的小胶质细胞 信号在所有三个组学平台上都受支持，预计将有助于ER行为和 生理表型。进一步的分析表明，MAP3K8，一种小胶质细胞特异的激酶，是一种 信号中枢在这一表型，并可能是一个潜在的靶点，以改善ER的表型。的目标是 这项建议是为了进一步确定内质网中BLA小胶质细胞的表型，并研究信号中枢的作用 MAP3K8在ER的行为和生理表型中的表达。特定目标1将使用荧光激活细胞 流式细胞仪分离BLA小胶质细胞，定量聚合酶链式反应检测KEY的特异性表达 在我们的初步分析中发现的基因。AIM 1还将使用共聚焦显微镜和定量 形态计量学方法测定ER相关小胶质细胞的形态及MAPK活性标志物的表达。这些 为了清楚地确定小胶质细胞的特定作用，有必要进行完善的细胞类型特异性研究，如 与其他细胞类型相反，在内质网表型上。特定目标2将使用行为药理学来检查 MAP3K8在内质网行为和生理表型中的作用我们将植入血乳酸靶向插管 附着于含有MAP3K8拮抗剂的渗透性微泵，在ER范例和行为之前 测试。不同的给药时间将使我们能够调查 MAP3K8信号转导。总体而言，这些研究的结果将确定BLA小胶质细胞和MAP3K8的作用 ER表型中的信号转导，并为人类丢失的负面后果提供可能的洞察力。

Project Summary Psychological loss occurs when one is deprived of something valuable, such as social relationships, physical health, or financial stability. These experiences often lead to depression-like symptoms such as amotivation, social withdrawal, weight gain and hypoactive stress responses. While loss experiences are common, little is known about the molecular mechanisms driving loss symptomology. Our laboratory uses enrichment removal (ER) to simulate the experience of loss in rats. In this protocol, rats are first housed in positive, highly stimulating environmental enrichment (EE) for a month, and then moved to single housing with no further stimuli. This removal alters how the rats respond to their environment, producing behaviors and physiology that specifically resemble loss in humans, including increased passive coping, weight gain, and hypoactive stress responses. We used an innovative discovery-based multi-omics approach to investigate ER molecular mechanisms in the basolateral amygdala (BLA), a traditionally stress promoting region that is differentially activated in EE and ER. Using a combination of RNA sequencing, proteomics, and kinomics, we identified a unique microglia phenotype in ER. We hypothesize that ER decreases signaling in BLA microglia, impairing their ability to regulate nearby neurons and dampen stress-promoting BLA signals. This dysregulated microglia signaling is supported on all three omics platforms and is expected to contribute to ER behavioral and physiological phenotypes. Additional analyses revealed that MAP3K8, a microglia-specific kinase, is a signaling hub in this phenotype and could be a potential target to ameliorate ER phenotypes. The objective of this proposal is to further define BLA microglia phenotypes in ER and investigate the role of the signaling hub MAP3K8 in ER behavioral and physiological phenotypes. Specific Aim 1 will use fluorescence activated cell sorting (FACS) to isolate BLA microglia, followed by qPCR to determine microglia-specific expression of key genes identified in our preliminary analyses. Aim 1 will also use confocal microscopy and quantitative morphometry to determine ER-related microglia morphology and expression of MAPK activity markers. These well-established, cell type specific studies are necessary to clearly define the specific role of microglia, as opposed to other cell types, in ER phenotypes. Specific Aim 2 will use behavioral pharmacology to examine the role of MAP3K8 in ER behavioral and physiological phenotypes. We will implant BLA-targeted cannula attached to osmotic minipumps containing a MAP3K8 antagonist prior to the ER paradigm and behavioral testing. Differential timing of drug delivery will allow us to investigate both the necessity and sufficiency of MAP3K8 signaling. Overall, the findings of these studies will define the role of BLA microglia and MAP3K8 signaling in ER phenotypes, and provide possible insight into the negative consequences of loss in humans.