Social isolation stress: role of neurosteroids in the action of ethanol on GABAAR

描述（由申请人提供）：最近，有人提出乙醇的某些作用可能是通过大脑中神经活性类固醇（如3a，5a-THP）水平的增加来介导的，这些类固醇具有在纳摩尔浓度下增加GABAA受体功能的能力。乙醇诱导的神经活性类固醇浓度的增加似乎是由于刺激下丘脑-垂体-肾上腺（HPA）轴，至少部分是直接刺激脑内eurosteroids的生成。社会隔离是一种长期轻度压力的模式，已被证明与显著的行为改变有关，如运动活动增加、焦虑、抑郁和实验室动物的攻击性。社会隔离导致脑和血浆中神经活性类固醇浓度下降，并伴有对急性应激刺激的异常反应，以及急性给药乙醇引起的神经类固醇生成效应增加。我们打算用C57BL/6J小鼠品系的社会隔离作为长期轻度应激的模型，进一步研究神经类固醇在乙醇对海马和杏仁核GABAA受体功能的急性和慢性作用中的作用及其与应激的相互作用。在社会隔离小鼠中，我们将在以下实验条件下测量乙醇的作用：i)急性给药后；Ii)通过蒸汽吸入慢性暴露后，iii)在自由选择的饮酒范式中，以确定这种情况是否会增加乙醇消耗。用RIA评估脑组织神经类固醇水平。RNase保护实验及免疫组织化学检测GABAA受体基因表达。采用常规全细胞膜片钳在脑切片上记录GABAA受体功能。该项目如果成功，将有助于进一步了解神经类固醇作为乙醇某些药理作用的内源性介质的作用，以及应激诱导神经元可塑性适应性变化的分子机制以及与乙醇消耗的相互作用。鉴于许多压力，以及生理病理条件，如经前症候群，抑郁和焦虑可能与血浆和脑内神经类固醇浓度的显著波动有关，本研究的结果将提供对乙醇滥用的脆弱性的额外见解，这是这些条件的特征。

DESCRIPTION (provided by applicant): Recently, it has been proposed that certain effects of ethanol could be mediated by an increase in the brain levels of neuroactive steroids, such as 3a,5a-THP, which has the ability to increase at nanomolar concentrations the GABAA receptor function. Ethanol-induced increase in neuroactive steroid concentrations appear to result from stimulation of the hypothalamic-pituitary-adrenal (HPA) axis, and at least in part, from a direct stimulation of brain eurosteroidogenesis. Social isolation is a model of prolonged mild stress that has been shown to be associated to marked behavioral alterations, such as increased locomotor activity, anxiety, depression, and aggressiveness in laboratory animals. Social isolation results in a decrease in the brain and plasma concentrations of neuroactive steroids, and is accompanied by an abnormal response to acute stressful stimuli as well as by an increased neurosteroidogenic effect induced by the acute administration of ethanol. We intend to use social isolation in C57BL/6J mouse strain as a model of prolonged mild stress in which to investigate further the role of neurosteroids in the acute and chronic actions that ethanol exerts on the function of GABAA receptors in the hippocampus and amygdala and the interplay with stress. In socially isolated mice, we will measure the effects of ethanol in the following experimental condition: i) after acute administration; ii) after chronic exposure by vapor inhalation, and iii) in a free-choice drinking paradigm in order to establish if this condition can increase ethanol consuption. Neurosteroid levels in brain tissue will be assessed by RIA. GABAA receptor gene expression by RNase protection assay, and by immunohistochemistry. GABAA receptor function will be examined by conventional whole-cell patch clamp recording in brain slices. This project, if successful, by employing the social isolation stress will help to understand further the role of neurosteroids as endogenous mediators of certain pharmacological actions of ethanol as well as the molecular mechanisms underlying the neuronal plastic adaptive changes induced by stress and the interplay with ethanol consumption. Given that many stressful, as well as physiopathological, conditions such as PMS, depression and anxiety could be associated to marked fluctuations in plasma and brain concentrations of neurosteroids, the outcome of this study will provide additional insights about the vulnerability to ethanol abuse, characteristic of such conditions.