Age-Related Changes in Renal Dopamine Receptor Function

肾多巴胺受体功能与年龄相关的变化

• 批准号: 7097261
• 负责人: Mustafa F. Lokhandwala
• 金额: $ 29万
• 依托单位: UNIVERSITY OF HOUSTON
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2010-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7097261
• 关键词: G protein coupled receptor kinase; aging; ascorbate; colorimetry; dietary supplements; dopamine receptor; enzyme activity; fenoldopam; glomerular filtration rate; hydrogen peroxide; immunofluorescence technique; immunoprecipitation; laboratory rat; nuclear factor kappa beta; oxidative stress; phospholipase D; phosphorylation; protein isoforms; protein kinase C; receptor expression; renal tubule; sodium potassium exchanging ATPase; tissue /cell culture; tocopherols; xanthine oxidase

DESCRIPTION (provided by applicant): Dopamine promotes an increase in renal sodium excretion by activating dopamine D1-like receptors in proximal tubules (PTs) and causing inhibition of Na,H-exchanger and Na,K-ATPase. We have shown that the ability of dopamine to inhibit these sodium transporters is reduced in old rats and the natriuretic response to dopamine is also diminished in older animals. This is due to a defective D1-like receptor-coupled signal transduction pathway, caused by hyper-serine-phosphorylation of D1A receptor in old rats, and an increase in protein kinase C (PKC) activity in the PTs. G-protein coupled receptor kinases (GRKs) are known to phosphorylate and desensitize dopamine D1 receptors. In preliminary studies, we found an increase in oxidative stress in old rats and antioxidant supplementation lowered oxidative stress, decreased basal PKC activity, and restored natriuretic response to D1 receptor activation. This application will test the hypothesis that increase in oxidative stress causes increase in PKC activity, which via activation of GRKs, produces an increase in the basal serine-phosphorylation of D1A receptors, causing it's uncoupling from G-proteins. Experiments are designed to determine the mechanism of oxidative stress-induced increase in basal PKC activity, role of specific PKC isoforms (beta & delta) and GRK isoform (GRK-2) in hyper-serine-phosphorylation of D1A receptor and G-protein uncoupling in proximal tubular cell cultures exposed to oxidants and in old rats. In order to examine the role of oxidative stress in D1A receptor G-protein uncoupling in old rats, animals will be given antioxidants supplementation followed by measurements of oxidant levels, PKC and GRK activities and D1A receptor signaling and natriuretic response to D1-like agonist, fenoldopam. The results will allow us to identify the molecular basis of renal D1 receptor dysfunction in old rats. Our findings will have a far reaching significance as it relates to the use of antioxidants in restoring defective G-protein coupled receptor function and drug responsiveness associated with increased oxidative stress in aging.

描述（申请人提供）：多巴胺通过激活近端小管（PT）中的多巴胺 D1 样受体并抑制 Na,H 交换器和 Na,K-ATP 酶，促进肾钠排泄增加。我们已经证明，老年大鼠中多巴胺抑制这些钠转运蛋白的能力降低，并且老年动物对多巴胺的利尿钠反应也减弱。这是由于老年大鼠 D1A 受体过度丝氨酸磷酸化和 PT 中蛋白激酶 C (PKC) 活性增加引起的 D1 样受体偶联信号转导通路缺陷所致。已知 G 蛋白偶联受体激酶 (GRK) 可使多巴胺 D1 受体磷酸化并使其脱敏。在初步研究中，我们发现老年大鼠的氧化应激增加，抗氧化剂补充剂降低了氧化应激，降低了基础 PKC 活性，并恢复了对 D1 受体激活的利尿钠反应。该应用将测试氧化应激增加导致 PKC 活性增加的假设，PKC 活性通过 GRK 的激活，导致 D1A 受体的基础丝氨酸磷酸化增加，导致其与 G 蛋白解偶联。实验旨在确定氧化应激诱导基础 PKC 活性增加的机制、特定 PKC 异构体​​（β 和 δ）和 GRK 异构体 (GRK-2) 在暴露于氧化剂的近端肾小管细胞培养物和老年大鼠中 D1A 受体丝氨酸过度磷酸化和 G 蛋白解偶联中的作用。为了研究氧化应激在老年大鼠 D1A 受体 G 蛋白解偶联中的作用，给动物补充抗氧化剂，然后测量氧化剂水平、PKC 和 GRK 活性以及 D1A 受体信号传导和对 D1 样激动剂非诺多泮的利尿钠反应。这些结果将使我们能够确定老年大鼠肾 D1 受体功能障碍的分子基础。我们的研究结果将具有深远的意义，因为它涉及使用抗氧化剂来恢复有缺陷的 G 蛋白偶联受体功能以及与衰老过程中氧化应激增加相关的药物反应性。