Zebrafish model: molecular basis of acute EtOH tolerance

描述（由申请人提供）：对乙醇的急性功能耐受性（AFT）是一种众所周知的现象，被认为会影响酒精中毒风险责任的个体差异，但AFT的潜在分子底物仍然是一个谜。在本申请中，我们提议测试斑马鱼（Danio rerio）对乙醇产生的行为中断会表现出AFT的假设，其主要目标是表征和发展斑马鱼作为研究AFT分子基础的模式生物。斑马鱼为遗传学研究提供了许多优势，包括世代时间短、后代数量多、成本低和空间需求低。关于斑马鱼基因的信息越来越多，基因操作也越来越容易完成。在AFT研究中，斑马鱼的另一个重要优势是，由于乙醇是通过与保存槽溶液的平衡来施用的，因此可以很容易地维持稳态的血液和脑乙醇浓度；本质上，就是一个非常简单的乙醇夹。在金鱼身上已经证明了AFT对某些行为的影响，但在斑马鱼身上还没有研究过。该项目的具体目标是筛选几种已建立的斑马鱼行为以获得AFT，通过测试几种近交斑马鱼品系来确定AFT是否存在自然发生的遗传变异，并开发自动化、高通量的AFT诱变筛选方法。未来的研究将在大量诱变的斑马鱼上实施高通量筛选，目标是绘制斑马鱼中AFT的分子基础。

DESCRIPTION (provided by applicant): Acute functional tolerance (AFT) to ethanol is a well known phenomenon that is thought to influence individual differences in risk liability for alcoholism, but the underlying molecular substrates of AFT remain a mystery. In this application, we are proposing to test the hypothesis that the zebrafish (Danio rerio) will exhibit AFT to the behavioral disruptions produced by ethanol, with the main goal of the proposal to characterize and develop the zebrafish as a model organism in the study of the molecular basis of AFT. Zebrafish provide numerous advantages for genetic studies including short generation time, large numbers of offspring, low cost and space demands, a growing body of information on the genetics of zebrafish, and the ease with which genetic manipulations can be accomplished. Another important advantage of zebrafish in AFT studies is that a steady-state blood and brain ethanol concentration can easily be maintained since the ethanol is administered via equilibration with the holding tank solution; in essence, an extremely simple ethanol clamp. AFT for certain behaviors has been demonstrated in goldfish, but it has not been examined in zebrafish. The Specific Aims of this project are to screen several established zebrafish behaviors for the acquisition of AFT, determine if there is naturally occurring genetic variation for AFT by testing several inbred zebrafish strains, and to develop methods for automated, high-throughput mutagenesis screens for AFT. Future studies will implement the high-throughput screens on large numbers of mutagenized zebrafish with the goal of mapping the molecular basis of AFT in zebrafish.