Regulation of Renal Response to Vasopressin by Glycogen Synthase

描述（由申请人提供）：糖原合成酶激酶3（GSK 3）是丝氨酸/苏氨酸蛋白激酶家族，由两种亚型GSK 3 α和GSK 3 β组成。在肾脏中，已知GSK 3 β调节细胞分化和正常上皮功能。尽管体外和体内证据表明锂，一种双相情感障碍的常用疗法和一种有效的GSK 3 β抑制剂可以减少肾脏对加压素的反应，但GSK 3 β在肾脏水转运中的作用尚不清楚。我们在锂诱导的多尿小鼠模型以及集合管特异性GSK 3基因敲除小鼠中的观察表明，GSK 3通过调节水通道蛋白2的表达在肾脏水重吸收中起重要作用。在GSK 3基因敲除小鼠中，腺苷酸环化酶活性和cAMP水平通过一种未确定的机制降低，这可能导致水通道蛋白2的低表达和对加压素的反应。基于这些证据，我们推测GSK 3 β在肾脏水平衡中起着关键作用。该激酶在正常肾脏水重吸收中的作用将在以下3个目标中确定。1)第一个目标将确定GSK 3如何调节腺苷酸环化酶活性，通过检查GSK 3是否结合或磷酸化腺苷酸环化酶使用定点诱变。2)AVP信号传导激活GSK 3的机制将通过检验AVP信号传导通过抑制经典Wnt信号传导激活GSK 3的假设来确定。一个负反馈回路，蛋白激酶A可能调节GSK 3也将被检查。3)第三个目的是检验AVP抗性-锂诱导的NDI是肾GSK 3抑制的病理生理学结果的假设。由于增加的AVP信号传导和高cAMP水平有助于多囊肾疾病的进展，因此该目的还将测试GSK 3的抑制或基因缺失可以减少囊肿形成的假设。这些研究将利用野生型和集合管特异性GSK 3基因敲除小鼠，内髓集合管细胞和小鼠皮质集合管细胞的原代培养物。通过这些研究，我们期望确定GSK 3调节肾集合管AVP信号的机制及其生理和病理生理意义。

DESCRIPTION (provided by applicant): Glycogen synthase kinase 3 (GSK3) is a family of serine/threonine protein kinases that consists of two isoforms, GSK3a and GSK3�. In the kidney, GSK3� is known to regulate cell differentiation and normal epithelial function. Despite in vitro and in vivo evidence that lithium, a common therapy for bipolar disorders and a potent inhibitor of GSK3� can reduce renal response to vasopressin, the role of GSK3� in renal water transport is not clear. Our observations in the lithium induced polyuric mouse model as well as collecting duct specific GSK3� knockout mice showed that GSK3� plays a significant role in renal water reabsorption by regulating aquaporin 2 expression in response to vasopressin. In the GSK3� knockout mice, adenylate cyclase activity and cAMP levels were reduced by an undetermined mechanism, which could have led to low aquaporin 2 expression and trafficking in response to vasopressin. Based on these evidences we hypothesize that GSK3� plays a critical role in renal water homeostasis. The role of this kinase in normal renal water reabsorption will be determined in the following 3 aims. 1) The first aim will determine how GSK3 regulates adenylate cyclase activity by examining if GSK3 binds to or phosphorylates adenylate cyclase using site directed mutagenesis. 2) The mechanism by which AVP signaling activates GSK3 will be determined by testing the hypothesis that AVP signaling activates GSK3 by inhibiting the canonical Wnt signaling. A negative feedback loop by which protein kinase A might regulate GSK3 will also be examined. 3) The third aim will test the hypothesis that AVP resistant- lithium induced NDI is a pathophysiological consequence of inhibition of renal GSK3. Since increased AVP signaling and high cAMP levels contribute to the progression of polycystic kidney disease, this aim will also test the hypothesis that inhibition or gene deletion of GSK3 can reduce cystogenesis. These studies will utilize wild type and collecting duct specific GSK3� knockout mice, primary cultures of inner medullary collecting duct cells and mouse cortical collecting duct cells. Through these studies we exoect to identify the mechanism by which GSK3 regulates AVP signaling in the renal collecting duct and its physiological and pathophysiological significance.