Molecular Studies of Osmotic Neural Sensing

描述(由申请人提供)： 目前的项目《渗透神经传感的分子研究》是基于 香草素受体相关基因的最新克隆 渗透激活离子通道(VR-OAC)及其可能的识别 中枢神经系统下丘脑的渗透压感受器。在这 提案中，申请人概述了一系列实验，以进一步探索 这一新分子的功能及其在盐调节中的可能作用 体内平衡。渗透压的动态平衡是 所有活细胞。在包括人类在内的脊椎动物中，全身渗透 压力是所有设定值中捍卫最积极的 自我平衡调节的系统。对渗透压的感知以及 适应性反应-饮酒行为和抗利尿激素的分泌- 它们在解剖学上都定位于下丘脑。VR-OAC和Vr-OAC的克隆 它在下丘脑的定位打开了理解 这种分子水平上的动态平衡电路。当前应用程序 描述了一组将进一步表征Vroac基因的实验。 最重要的是，Vroac-/-小鼠将通过基因靶向产生。一位将军 针对中枢神经系统的基因敲除和组织特异性基因敲除 建议。这些基因敲除的小鼠品系将被检查饮酒行为， 渗透调节、肾功能和液体动态平衡 VR-OAC表达神经元的血压调节和轴突投射 和内耳功能。VR-OAC在遗传修复中的潜力 体内有缺陷的渗透感觉将在一种突变株中进行检查 线虫，秀丽线虫，在渗透感觉上有缺陷。在……里面 这些研究表明，具有这种感知缺陷的OSM-9突变蠕虫将被交叉到 过表达大鼠VR-OAC的转基因蠕虫。这一实验设置将 允许快速评估VR-OAC的结构-功能关系 活体内。通过对vroac基因的建议缺失和对其的分析 Vroac-/-小鼠以及突变的蠕虫，申请人打算寻求 提出批判性问题，并牢牢掌握 分子遗传学，特别是小鼠基因打靶。被赋予这些 Vroac基因功能基因组学的技术和经验 应聘者打算发展自己的事业，成为一名独立的大三学生 调查员。

DESCRIPTION (provided by applicant): The current project "Molecular studies of osmotic neural sensing" is based on the recent cloning by the applicant of the vanilloid receptor related osmotically activated ion channel (VR-OAC) and its identification as a possible osmolarity sensor in the hypothalamus of the central nervous system. In this proposal, the applicant outlines a series of experiments to further explore the functions of this novel molecule and its possible role in regulating salt balance in-vivo. Homeostasis of osmotic pressure is a vital prerequisite for all living cells. In vertebrate organisms including humans, systemic osmotic pressure is the most aggressively defended set-point value of all homeostatically regulated systems. Sensing of osmotic pressure as well as the adaptive response - drinking behavior and secretion of antidiuretic hormone - all localize anatomically to the hypothalamus. The recent cloning of VR-OAC and its localization to the hypothalamus opened the door to an understanding of this homeostatic circuit at the molecular level. The current application describes a set of experiments that will characterize the Vroac gene further. Most importantly, Vroac-/- mice will be generated by gene-targeting. A general knockout and a tissue-specific knockout for the central nervous system are proposed. These knockout mouse lines will be examined for drinking behavior, osmotic regulation, kidney function and fluid homeostasis including the regulation of blood pressure, axonal projections of VR-OAC expressing neurons and inner ear function. The potential of VR-OAC to restore genetically defective osmotic sensing in-vivo will be examined in a mutant strain of the roundworm, Caenorhabditis elegans, that has defects in osmotic sensing. In these studies, osm-9 mutant worms with such sensing defects will be crossed to transgenic worms that overexpress rat VR-OAC. This experimental set-up will allow for a rapid evaluation of structure-function relationships of VR-OAC in-vivo. By means of the proposed deletion of the Vroac gene and analysis ot Vroac-/- mice as well as the mutant worms, the applicant intends to pursue a critical question and develop a firm grasp of the immensely powerful tools of molecular genetics and in particular, mouse gene-targeting. Endowed with these techniques and the experience of functional genomics of the Vroac gene, the applicant intends to advance his career and become an independent junior investigator.