Two-Pore Domain Potassium Channels and Aldosterone Secretion

描述（由申请人提供）：醛固酮是一种早发性致病刺激物，对心血管和肾脏稳态产生不利影响。它在心力衰竭、慢性肾病和某些形式的高血压中升高。虽然体内醛固酮的产生长期以来一直被认为是由Ang I和钾调节的，但K通道调节对Ang II的生理作用的贡献尽管是假设的，但仍未得到证实。最新一代的ASK-1，ASK-3，和ASK-1/ASK-3基因敲除小鼠提供了令人兴奋的可能性，以建立在体内的醛固酮生产的生理控制的ASK通道调节的功能重要性。我们的假设，在体外测试的背景K电流和膜电位的Ang II和酸化的分泌醛固酮的肾上腺髓质肾小球（ZG）细胞的控制，任务亚基。此外，这些通道在体内调节醛固酮产生和盐/水平衡中重要的假设是在基础条件下以及在激素和代谢挑战期间确定的（即，AngII输注，Na负荷和剥夺）。我们追求以下具体目标：目标1：使用qRT-PCR、原位杂交和免疫组织化学生成并验证新型常规和条件性ASK通道亚基敲除小鼠系;目标2：确定对照和ASK通道敲除小鼠中ZG细胞的膜特性，表征背景K通道电流及其受AngII、细胞外K和浴pH的调节，并评价这些通道在设置膜电压和支持ZG细胞兴奋性中的作用;目的3：评价在对照和ASK通道敲除小鼠中，ASK通道表达对体内控制醛固酮产生的重要性，确定：稳态醛固酮分泌对由盐平衡的饮食操纵引起的RAAS抑制的反应，以及醛固酮分泌对体内外源性AngII或酸负荷的递送的反应。这一建议的优势在于结合使用分子和细胞生物学，电生理学和整个动物实验。最低限度，我们的努力将澄清的作用，任务渠道的生产醛固酮。最理想的情况是，我们的工作将确定一个新的目标，用于治疗心力衰竭，慢性肾脏疾病和顽固性高血压患者。 项目叙述：特发性原发性醛固酮增多症（IHA）存在于13%的非选择性高血压患者中，在那些患有顽固性高血压的患者中上升至20%。使用细胞和体内方法的组合，我们的研究确定了TaskK通道作为控制醛固酮（IHA中升高的类固醇激素）产生的关键膜蛋白。该靶标的鉴定可用于为IHA患者以及心力衰竭和慢性肾病患者（其中升高的醛固酮也加速疾病进展）产生有用的治疗。

DESCRIPTION (provided by applicant): Aldosterone is an early-onset pathogenic stimulus that adversely influences cardiovascular and renal homeostasis. It is elevated in heart failure, chronic kidney disease and some forms of hypertension. Although the production of aldosterone in vivo has long been known to be regulated by Ang I and potassium, the contribution of K channel regulation to the physiological actions of Ang II, though hypothesized, has remained unproven. The recent generation of TASK-1, TASK-3, and TASK-1/TASK-3 knockout mice offers the exciting possibility to establish the functional importance of TASK channel regulation to the physiological control of aldosterone production in vivo. Our hypothesis that TASK subunits contribute to background K currents and the control of membrane potential by Ang II and acidification in the aldosterone secreting adrenal zona glomerulosa (ZG) cells is tested in vitro. Moreover, the hypothesis that these channels are important in the regulation of aldosterone production and salt/water balance in vivo is determined under basal conditions and during hormonal and metabolic challenge (i.e., AngII infusion, Na loading and deprivation). We pursue the following specific aims: Aim 1: Generate and validate novel conventional and conditional TASK channel subunit knockout mouse lines using qRT-PCR, in situ hybridization, and immunohistochemistry; Aim 2: Determine membrane properties of ZG cells in control and TASK channel knockout mice, characterizing background K channel currents and their modulation by AngII, extracellular K and bath pH, and evaluating the role of these channels in setting membrane voltage and supporting ZG cell excitability; Aim 3: Evaluate the importance of TASK channel expression to the control of aldosterone production in vivo in control and TASK channel knockout mice determining: the steady-state aldosterone secretory response to RAAS suppression caused by dietary manipulation of salt balance, and the aldosterone secretory response to the delivery of exogenous AngII or acid loading in vivo. The strength of this proposal lies in the combined use of molecular and cell biology, electrophysiology and whole animal experimentation. Minimally, our efforts will clarify the role of TASK channels in the production of aldosterone. Optimally, our work will identify a new target for the therapy of patients with heart failure, chronic kidney disease and resistant hypertension. Project Narrative: Idiopathic primary hyperaldosteronism (IHA) is present in 13% of non-selected hypertensive patients and rises to 20% among those that have resistant hypertension. Using a combination of cellular and in vivo approaches our studies identify TASK K channels as critical membrane proteins that control the production of aldosterone, the steroid hormone that is elevated in IHA. The identification of this target can be used to generate a useful therapy for patients with IHA but also for patients with heart failure and chronic kidney disease where elevated aldosterone also hastens disease progression.