ROMK-CFTR INTERACTIONS IN THE DISTAL NEPHRON

远端肾单位中的 ROMK-CFTR 相互作用

• 批准号: 6725891
• 负责人: STEVEN C HEBERT
• 金额: $ 30.14万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-30 至 2008-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6725891
• 关键词: Xenopus oocyte; chloride channels; chloride ion; cyclic AMP; electrophysiology; fluorescence; genetically modified animals; glyburide; ion transport; laboratory mouse; laboratory rat; potassium channel; potassium ion; protein protein interaction; renal tubular transport; renal tubule; voltage /patch clamp

Kir1.1(ROMK) forms the small conductance (SK) ATP-sensitive K channel (KATP) mediating apical K recycling in thick ascending limb (TAL) and K secretion in principal cells of the mammalian kidney. SK channel activity is absent in mice with deletion of the Kcnj 1 (Kir1.1) gene and these ROMK-deficient mice have a salt wasting phenotype consistent with human ROMK Bartter's syndrome. Thus, ROMK is crucial to renal Na and K handling. All KATP channels are thought to be formed by four pore-forming subunits (Kir6.x or Kir1.1) in association four ATP-Binding Cassette (ABC) proteins. The associated ABC proteins mediate sensitivity to sulfonylureas (e.g., glibenclamide) and are believed play crucial roles in metabolic sensing of nucleotides. CFTR, which is expressed at apical borders of TAL and principal cells, has been suggested to be the ABC partner of ROMK based on co-expression studies in X. laevis oocytes. Yet, virtually nothing is known in renal TAL and principal cells about the roles of CFTR in SK channel activity and metabolic regulation, the circumstances where CFTR may function as an apical C1 channel and the functional consequences of this anion channel activity on cation transport, or the effects that ROMK has on CFTR CI channel activity. We hypothesize that: (1) CFTR mediates glibenclamide sensitivity and is involved in nucleotide sensing (ATP inhibition and ADP activation) by SK channels when cell cyclic AMP (cAMP) is low; (2) the functional interaction between CFTR and ROMK (or ROMK on CFTR) is lost when cyclic AMP is high with consequent loss of glibenclamide sensitivity and reduced ability of ATP to inhibit SK; (3) apical CFTR Cl channels are silent during low, but activated during high, cAMP conditions; and (4) CFTR Cl current partially shunts the apical K secretory current allowing for electrical dissociation of K secretion from Na absorption in principal cells. These cAMP-mediated changes in CFTR-ROMK interactions and function with cell high cAMP could enhance K secretion in principal and thick ascending limb cells when distal tubular flow is low. Biochemical and patch clamp studies in X. laevis oocytes expressing CFTR-ROMK will be used to assess the nature of channel interactions and the effects of cAMP. Patch clamping of apical membranes of rat TAL and principal cells will be used to assess SK and CFTR Cl channel activity, regulation and interactions. In vivo and in vitro microperfusion will be used to study the functional consequences of ROMK-CFTR interactions in rat distal nephron segments. Similar studies in ROMK and/or CFTR deficient mice will be used to dissect out the effects of CFTR on ROMK function and of ROMK on CFTR function.

Kir1.1（ROMK）形成小电导（SK） atp敏感钾通道（KATP），介导厚升肢（TAL）的顶端钾循环和哺乳动物肾脏主要细胞的钾分泌。在Kcnj 1 （Kir1.1）基因缺失的小鼠中，SK通道活性缺失，这些ROMK缺陷小鼠具有与人类ROMK Bartter综合征一致的盐消耗表型。因此，ROMK对肾脏Na和K的处理至关重要。所有的KATP通道都是