Genomics analysis of social stress and individual variation in ethanol drinking

描述(由申请人提供)：我们实验室最近对C57BL/6近交系小鼠选择饮用2瓶酒精的研究证明，日常饮酒行为非常一致，存在显著的个体差异。这种饮酒的个体差异受到环境因素的影响，包括通过与非兄弟姐妹的集体住房形成社会等级。这种饮酒量的变化在糖精或总液体摄入量中看不到。在单个动物身上进行的初步微阵列研究已经确定了与这种饮酒行为高度相关的表达模式。我们的假设是，压力(隔离、社会等级)已经对这些基因相同的动物的饮酒行为产生了长期的影响。这提供了一个独特的机会，可以细致地剖析环境影响导致饮酒行为差异的分子机制，特别是在社会互动方面。我们预测，我们的结果将与“传统”遗传方法确定的与饮酒行为相关的基因网络部分重叠，并识别出在人群研究中无法辨别的饮酒行为的新调节因素。我们将实施五个具体目标。目的1将进一步表征我们的行为模型，以确定由集体住房引起的个人饮酒行为变化的稳定性。目标2将描述饮酒行为的个体差异是否与焦虑和压力的行为或神经化学措施相关。这些研究将包括评估血浆皮质酮水平和大脑区域促肾上腺皮质激素释放因子(CRF)和神经类固醇别孕酮的水平。AIM 3将使用全基因组微阵列研究来识别与饮酒行为或社会压力因素(如集体住房或隔离住房)相关的基因表达模式。AIM 4将通过INIA-Stress生物信息学核心以及与切斯特、兰斯顿和威廉姆斯博士的合作，使用生物信息资源来提炼微阵列数据。候选基因将通过多变量排序方案进行选择，并通过Q-RTPCR、Western blotting和免疫细胞化学验证其表达。从我们的微阵列研究中挑选的候选者也将被用来确定相同的基因是否在来自比吉奥博士(项目11)和灵长类动物(格兰特博士，项目3)的老鼠样本中显示出与压力或饮酒行为的相关性。在目标5中，这些基因将用于预测来自BXD重组近交系或来自该INIA的其他小鼠资源的小鼠品系的饮酒行为或饮酒对社会压力的反应。最后，这些候选基因将通过从Genetrap ES细胞中产生空小鼠系(通过Knokout Core，德尔皮尔博士)或使用病毒载体将基因构建传递到靶向大脑反应来进行行为学验证，然后进行行为研究。因此，这些研究可能会识别导致人类饮酒量因人而异的基因或基因网络，导致一些人过度饮酒并有酗酒的风险。

DESCRIPTION (provided by applicant): Recent studies in our laboratory on ethanol 2-bottle choice drinking with C57BL/6 inbred mice have documented exceedingly consistent, substantial individual variation in daily drinking behavior. This individual variation in ethanol drinking is modified by environmental factors, including formation of social hierarchy through group housing with non-siblings. This variation in drinking is not seen for saccharin or total fluid intake. Initial microarray studies done on individual animals have identified expression patterns highly correlated with this drinking behavior. It is our hypothesis that stress (isolation, social hierarchy) has created long-lasting influences altering drinking behavior in these genetically identical animals. This offers a unique opportunity to finely dissect molecular mechanisms underlying difference in drinking behavior generated by environmental influences, particularly in regard to social interactions. We predict our results will partially overlap with drinking behavior-related gene networks identified by "traditional" genetic approaches, as well as identify novel modulators of drinking behavior not discernable in studies on populations. We will conduct five specific aims. Aim 1 will further characterize our behavioral model to determine the stability of the variation in individual drinking behavior induced by group housing. Aim 2 will characterize whether individual variation in drinking behavior correlates with behavioral or neurochemical measures of anxiety and stress. These studies will include assessment of plasma corticosterone levels and brain regional levels of corticotropin releasing factor (CRF) and the neurosteroid, allopregnanaolone. Aim 3 will use whole genome microarray studies to identify gene expression patterns correlating with drinking behavior or social stressors such as group housing or isolation housing. Aim 4 will refine microarray data using bioinformatic resources through the INIA-Stress bioinformatics core and collaborations with Drs. Chester, Langston and Williams. Candidate genes will be selected through a multivariate ranking scheme and expression verified by Q-rtPCR, Western blotting and immunocytochemistry. Candidates selected from our microarray studies will also be used to determine if the same genes show correlations with stress or drinking behavior in mouse samples from Drs. Biggio (Project 11) and primates (Dr. Grant, Project 3). In Aim 5, these genes will be used to predict drinking behavior or drinking responses to social stress in mouse lines derived from the BXD recombinant inbred panel or othe mouse resources from this INIA. Finally, these candidate genes will be behaviorally verified by generating null mouse lines from genetrap ES cells (via the Knockout Core, Dr. Delpire) or using viral vectors to deliver gene constructs to target brain resions followed by behavioral studies. These studies might thus identify genes or networks of genes that cause human alcohol consumption to vary across individuals, leading some to excessive alcohol intake and risk for alcoholism.