Neuroactive Steroids and Allostasis Induced by Ethanol/Stress

描述(申请人提供)：本项目的目标是系统地描述慢性间歇性乙醇(CIE)暴露对应激挑战后边缘脑区GABA能神经活性类固醇的影响及其调节。神经元和细胞外神经活性类固醇水平将通过免疫组织化学以及微透析液和/或组织标本的气相色谱-质谱仪(GC-MS)分析进行检测，从而首次对大脑边缘区域进行调查。我们建议测试假设，酒精暴露失调的基础或压力诱导的这些类固醇水平，这些变化与酒精消费有关。我们将进一步探索遗传对神经类固醇水平或反应的适应的贡献。第一个目标是确定CIE对C57BL/6J小鼠边缘/奖赏脑区神经元和细胞外3a，5a-THP水平的影响，这些脑区包括C57BL/6J小鼠的前额叶皮质、伏隔核、杏仁核和终纹床核。目的2研究急性应激对C57BL/6J小鼠边缘脑区神经元和细胞外3a，5a-THP的影响，以及CIE 5个周期后应激刺激的影响。目的3将确定在BXD小鼠品系中，边缘/奖赏脑区的基础或CIE相关神经活性类固醇水平是否与饮酒偏好相关。目的4将扩展和比较我们关于神经活性类固醇在小鼠和灵长类动物中对CIE的适应的研究。我们将描述长期饮酒对恒河猴边缘脑区3a，5a-THP水平的影响。我们的结果将与我们的合作者在相同实验条件下进行的神经功能、神经化学、基因表达和饮酒行为研究的变化相结合。这些研究将阐明乙醇对大脑边缘区域GABA能神经活性类固醇的影响，并为基础和应激诱导的神经活性类固醇在慢性间歇酒精暴露的各种不平衡适应中的作用提供新的见解。

DESCRIPTION (provided by applicant): The goal of this project is to systematically delineate the effects of chronic intermittent ethanol (CIE) exposure on GABAergic neuroactive steroids and their regulation in limbic brain areas after stress challenge. Both neuronal and extracellular neuroactive steroid levels will be examined by immunohistochemistry as well as gas chromatography-mass spectroscopy (GC-MS) analysis of microdialysates and/or tissue specimens, allowing investigation of limbic brain regions for the first time. We propose to test the hypotheses that ethanol exposure dysregulates basal or stress-induced levels of these steroids and these changes are related to ethanol consumption. We will further explore genetic contributions to adaptations in neurosteroid levels or responses. The first aim will determine the effects of CIE on neuronal vs. extracellular 3a,5a-THP levels in limbic/reward brain areas, including the prefrontal cortex, nucleus accumbens, amygdala, and bed nucleus of the stria terminalis of C57BL/6J mice. Aim 2 will delineate the effect of acute stress challenge on neuronal vs. extracellular 3a,5a-THP in limbic brain areas and the effect of stress challenge following five cycles of CIE in C57BL/6J mice. Aim 3 will determine if basal or CIE-related neuroactive steroid levels in limbic/reward brain areas are correlated with ethanol preference drinking across BXD mouse strains. Aim 4 will extend and compare our studies on neuroactive steroid adaptations to CIE in mice to primates. We will delineate the effects of prolonged ethanol drinking on 3a,5a-THP levels in limbic brain areas of rhesus monkeys. Our results will be integrated with changes in neuronal function, neurochemistry, gene expression and ethanol drinking behavior studies by our collaborators under the same experimental conditions. These studies will elucidate the effects of ethanol on GABAergic neuroactive steroids in limbic regions of brain and provide novel insights into the role of basal and stress-induced neuroactive steroids in various allostatic adaptations to chronic intermittent ethanol exposure.