PROXIMAL TUBULAR TRANSPORT MECHANISMS

血管紧张素II(AngII)是一种主要的荷尔蒙，负责维持 细胞外容量下降时的电解质平衡和血压 由于钠的耗尽。它的作用部位之一是近端小管。 肾组织中血管紧张素Ⅱ在PM浓度下刺激碳酸氢钠 和液体重吸收，而在最大范围内它降低了NaCI 重吸收。血管紧张素转换酶在近端调节的新的重要方面 由于Angii的发现，小管最近变得明显起来 受体亚型(AT1A；AT1B；AT；非AT1-非AT2)，一种新的信号转导 涉及花生四烯酸的途径以及CFTR和CI的存在 电导。本项目旨在了解信令 与不同基因相关的途径和特定的电解质转运体 血管紧张素Ⅱ受体亚型。具体目标包括：10评估 AT1a、AT1B和AngIII/AngIV激动剂/拮抗剂的作用 AT2)细胞内钙离子水平的受体和穿过灌木丛的质子通量 边缘质膜和基侧质膜。2)重要性的确定 细胞色素P450系统在花生四烯酸依赖信号转导中的作用 近端小管。3)评价腔内CI的重要性 电导与囊性纤维化跨膜调节 蛋白。细胞内钙水平和pH值将用离子敏感法测量 荧光染料和视频显微镜。P450 2CAA的作用将是 通过对Angii的运输反应与 酶水平和花生四烯酸的免疫学/生化分析 非常相似的P450酶基因转导前后的代谢 2C2。CI电导将通过电生理技术和 与cftr信息和蛋白水平相关。其中一个重要的 可用于这些研究的工具是近端小管细胞系，我们 最近成立的。这些细胞系既适合生化的，也适合 分子生物学和细胞生理学研究，因此应该 帮助弥合关于受体/信号的信息之间的差距 途径和电解液的运输。这些研究应该提供 对不同技术的重要性的重要信息和见解 近端肾小管电解质的管腔和基底外侧血管紧张素Ⅱ受体 交通工具。

Angiotensin II (AngII) is a major hormone responsible for maintenance of electrolyte balance and blood pressure when the extracellular volume drops due to sodium depletion. One of its sites of action is the proximal tubule of the kidney where AngII at pM concentrations stimulates Na bicarbonate and fluid reabsorption while in the muM range it decreases NaCI reabsorption. New, important aspects of AngII regulation in the proximal tubule have recently become apparent due to the discoveries of AngII receptor subtypes (AT1A; AT1B; AT; non-AT1-non-AT2), of a novel signaling pathway involving arachidonic acid, and of the presence of CFTR and CI conductance. This project is directed towards understanding the signaling pathways and specific electrolyte transporters associated with different AngII receptor subtypes. The specific aims include:10 An evaluation of the effect of agonists/antagonists of AT1A AT1B, and AngIII/AngIV (non-AT1-non- AT2) receptors on cytosolic Ca levels and proton fluxes across the brush border and basolateral plasma membrane. 2) Determination of the importance of the cytochrome P450 system for arachidonic acid-dependent signaling in the proximal tubule. 3) Evaluation of the importance of luminal CI conductance and the Cystic Fibrosis Transmembrane Regulatory (CFTR) protein. Cytosolic Ca levels and pH will be measured with ion-sensitive fluorescent dyes and video microscopy. The role of P450 2CAA will be assessed by correlation of transport responses to AngII with immunological/biochemical analysis of enzyme levels and arachidonic acid metabolism before and after transfection of the very similar enzyme P450 2C2. CI conductance will be assessed by electrophysiological techniques and correlated with levels of CFTR message and protein. One of the important tools available for these studies are proximal tubule cell lines that we recently established. These cell lines are suitable for both biochemical, molecular biological, and cell-physiological investigations and thus should help to bridge the gap between information about receptors/signaling pathways and electrolyte transport. These studies should provide significant information and insights into the importance of different luminal and basolateral AngII receptors for proximal tubular electrolyte transporT.