PARACRINE REGULATION OF RENAL FUNCTION BY DOPAMINE IN NORMOTENSIVE AND HYPERT

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。截至2006年2月7日，血压受饮食盐摄入量的影响；事实上，30%血压正常的人和50-70%的高血压患者对盐敏感。多巴胺是一种众所周知的神经递质，通过对肾脏的独立的旁分泌作用，已被确定为血压，钠平衡，肾功能和肾上腺功能的重要调节剂。血管紧张素II也是钠平衡和血压的重要旁分泌调节剂。多巴胺能和肾素-血管紧张素系统已被证明在体外和体内相互作用。在这个方案中，我们将研究抑制肾素-血管紧张素系统对刺激多巴胺能系统的影响，在盐负荷和盐耗尽状态下，在正常血压和高血压的人类受试者中。我们将把这些影响与最近描述的肾近端小管多巴胺受体受体后信号通路中遗传多态性的存在或不存在联系起来，特别是G蛋白偶联受体激酶GRK4。总之，这些信息将帮助我们更好地了解盐和基因型对原发性高血压发病机制的影响，这种疾病影响全世界超过10亿人。假设血管紧张素转换酶（ACE）抑制剂依那普利可增强正常血压和高血压患者在盐负荷和盐耗尽状态下D1受体刺激引起的尿钠。这些影响在具有G蛋白偶联受体激酶（GRK4）多态性的受试者中减少。高血压患者肾素-血管紧张素张力升高，尿钠增加。方法/研究设计：双盲、安慰剂对照、交叉设计。从内科诊所招募了20名血压正常的志愿者和28名高血压患者，年龄在18-55岁之间，男女不限。他们都将有5天的低钠（50毫克/天）和中等钠（300毫克/天）饮食，间隔4周。每位受试者将依次接受四项干预措施，第一项干预措施根据计算机化计划随机进行。四种干预措施分别为：高盐饮食+安慰剂、高盐饮食+依那普利、低盐饮食+安慰剂、低盐饮食+依那普利。所有受试者在输注非诺多巴泮（一种选择性多巴胺（D1）受体激动剂）3小时之前、期间和之后进行肾功能检查。我们将测量尿钠排泄量，通过测量肌酐、多环芳烃和锂离子清除率，我们将分别估计肾小球滤过率、有效肾血浆流量和近端小管钠重吸收。所有受试者将对与原发性高血压相关的常见多态性进行基因分型，特别是GRK4、α内收蛋白、醛固酮合成酶和人类D5受体的遗传变异。我们将比较每位受试者在依那普利/安慰剂治疗期间以及低盐和中盐饮食期间钠排泄的基线变化。我们将分别对正常血压组和高血压组的尿钠排泄量进行研究，然后进行比较。我们将对所有受试者进行基因分型，并使用单位点和多位点分析来确定高血压和正常受试者之间多态性发生的差异。预期结果在血压正常的受试者中，通过ACE抑制血管紧张素II的形成将增加非诺多泮诱导的钠尿，在中等钠饮食中，以及在低钠饮食中，肾素-血管紧张素系统通常具有重要的钠潴留作用。在高血压患者中，非诺多泮的利钠反应已被报道受损，我们预计依那普利将在更大程度上增强肾素-血管紧张素张力升高的患者的利钠反应。我们研究了多巴胺的功能，特别是多巴胺能后受体信号通路在人类冠状动脉平滑肌细胞中的作用，特别是它在大钾通道激活中的可能作用，这种异常可能导致高血压患者肌源性血管张力增加。这项工作的第一作者论文题为“D5受体介导多巴胺对人类冠状动脉平滑肌细胞大KCa通道的影响”，Aruna Natarajan, Guichun Han， Richard White和Pedro Jose将在2006年4月的实验生物学会议上以海报的形式发表。这项工作将包括Aruna Natarajan在乔治城大学生理学和生物物理学博士学位的论文报告，目前正在准备中。3. 最初摘要：多巴胺是一种众所周知的神经递质，对肾脏具有旁分泌作用，最近已被阐明。这些影响导致钠尿反应钠负荷，介导的g蛋白偶联受体，D1受体。由于g蛋白偶联受体激酶GRK4的多态性，盐敏感性高血压患者的受体后信号通路被发现受损。本研究旨在通过研究血管紧张素转换酶（ACE）抑制对D-1受体激动剂非诺多泮（Fenoldopam）对尿盐排泄的影响，阐明盐负荷和缺盐状态下肾多巴胺能系统与肾素血管紧张素系统（RAS）的相互作用。该研究将这些影响与GRK4、醛固酮合成酶、α -内收蛋白和其他多态性的存在与否联系起来，所有这些多态性都在盐敏感性高血压患者中被描述过。这项研究将有助于我们了解盐、多巴胺、RAS和基因组成在原发性高血压发病机制中的相互作用。具体目的：这是一项由研究者发起的研究，由NCRR、NIH、DHSS的指导临床研究学者计划奖（Grant RR 17613）支持。多巴胺能系统对体液和电解质平衡以及血压有重要的调节作用。该项目的目的是确定在不同钠摄入量时，肾多巴胺能和RAS在钠排泄调节中的相互作用。具体的目的是确定：1。钠摄取量对RAS和D1多巴胺受体钠排泄相互作用的影响。2. GRK4基因变异对D1多巴胺受体激动剂的利钠反应的影响及其通过ACE抑制的调节。假设：D1多巴胺受体的利钠作用在钠负荷条件下表现最好，但被AT1血管紧张素受体的抗利钠作用所拮抗。在原发性高血压中，由于激活GRK4变异，D1受体的利钠作用受损，而AT1受体的抗利钠作用增强。总的假设是，在正常血压的受试者中，仅在盐负荷状态下，ACE抑制增加了D1受体刺激引起的尿钠。这些影响在具有GRK4基因变异的受试者和高血压受试者中降低

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. As of 2/07/06 Background Blood pressure is influenced by dietary salt intake; indeed 30% of people with normal blood pressure and 50-70% of people with hypertension are salt-sensitive. Dopamine, a well-known neurotransmitter, has been identified as an important modulator of blood pressure, sodium balance, renal function and adrenal function through an independent, paracrine effect on the kidneys. Angiotensin II is also an important paracrine regulator of sodium balance and blood pressure. The dopaminergic and renin-angiotensin systems have been shown to interact in vitro and in vivo. In this protocol, we will study the effects of inhibition of the renin-angiotensin system on the effects of stimulation of the dopaminergic system, in salt loaded and salt depleted states, in normotensive and hypertensive human subjects. We will correlate these effects with the presence or absence of recently described genetic polymorphisms in the post-receptor signaling pathway of the dopamine receptor in the proximal tubule of the kidney, specifically the G protein- coupled receptor kinase, GRK4. Together, this information will help us better understand the effects of salt and genotype on the pathogenesis of essential hypertension, a disease that affects more than a billion people worldwide. Hypothesis The angiotensin converting enzyme (ACE) inhibitor, Enalapril, augments natriuresis caused by D1 receptor stimulation, in normotensive and hypertensive human subjects, in salt loaded and salt depleted states. These effects are reduced in subjects who have polymorphisms of the G protein-coupled receptor kinase, GRK4. The augmentation of natriuresis is increased in hypertensive subjects with increased renin-angiotensin tone. Methods/Study Design This is a double blind, placebo-control, crossover design. 20 normotensive volunteers and 28 hypertensive subjects, recruited from an Internal Medicine clinic, of both sexes, between 18-55 years will participate in the study. They will all have 5 days each of low (50meq/day) sodium, and moderate (300meq/day) sodium diet, 4 weeks apart. Each subject will have four interventions applied in sequence, the first intervention being randomized as per a computerized plan. The four interventions are: high salt diet + placebo, high salt diet + Enalapril, low salt diet +placebo, and low salt diet + Enalapril. All subjects will undergo renal function tests, before, during and after a 3-hour infusion of Fenoldopam, which is a selective dopamine (D1) receptor agonist. We will measure urinary sodium excretion, and, by measuring creatinine , PAH and lithium clearance, we will estimate glomerular filtration rate, effective renal plasma flow, and proximal tubular sodium reabsorption of sodium respectively. All subjects will be genotyped for the common polymorphisms associated with the development of essential hypertension, specifically, GRK4, alpha adducin, aldosterone synthase, and genetic variants of the human D5 receptor. Statistical Analysis We will compare the change in the excretion of sodium from baseline between Enalapril/placebo treatments, and between low and moderate salt diet periods, in each subject. Urinary sodium excretion will be studied separately in normotensive and hypertensive groups, and then compared. We will genotype all subjects and using single locus and multi loci analyses to determine differences in the occurrence of polymorphisms between hypertensive and normotensive subjects. Anticipated Results In normotensive subjects, inhibition of angiotensin II formation by ACE inhibition will augment Fenoldopam -induced natriuresis in the moderate sodium diet, and also in the low sodium diet, where normally the renin-angiotensin system has an important sodium-retaining role. In hypertensive subjects, where the natriuretic response to Fenoldopam has been reported to be impaired, we anticipate that Enalapril will enhance natriuresis to a greater extent in those subjects with increased renin-angiotensin tone. Basic Science Component We study the function of dopamine, specifically dopaminergic post receptor signaling pathways in the human coronary vascular smooth muscle cells in humans, specifically its possible role in activation of big potassium channels, abnormalities in which may contribute to the increase in myogenic vascular tone seen in hypertension. This work has led to a first author abstract entitled 'D5 receptor mediates Dopamine effects on Big KCa channels in human coronary artery smooth muscle cells' Aruna Natarajan, Guichun Han, Richard White and Pedro Jose, which will be presented as a poster at the Experimental Biology meeting in April 2006. This work will comprise Aruna Natarajan's thesis report for a PhD in Physiology and Biophysics at Georgetown University, currently in preparation. 3. Please provide scientific progress achieved to date (since last year's report if applicable): Initial abstract Dopamine is a well-known neurotransmitter, with paracrine effects on the kidney that have recently been elucidated. These effects result in natriuresis in response to a sodium load, mediated by a G-protein coupled receptor, the D1 receptor. Post-receptor signaling pathways have been found to be impaired in people with salt sensitive hypertension, attributed to a polymorphism of a G-protein coupled receptor kinase, GRK4. This study aims to elucidate the interaction between the renal dopaminergic system and the renin-angiotensin system (RAS) in salt-loaded and depleted states, by studying the effect of angiotensin converting enzyme (ACE) inhibition on the effect of Fenoldopam, a D-1 receptor agonist on salt excretion in the urine. The study will correlate these effects with the presence or absence of genetic polymorphisms of GRK4, aldosterone synthase, alpha-adducin and other polymorphisms, all of which have been described in people with salt-sensitive hypertension. This study will help us understand the interactions of salt, dopamine, RAS and genetic makeup in the pathogenesis of essential hypertension. SPECIFIC AIMS: This is an investigator-initiated study supported by the mentored Clinical Research Scholar Program Award, Grant RR 17613, from the NCRR, NIH, DHSS. The dopaminergic system has a major regulatory effect on fluid and electrolyte balance and blood pressure. The objective of this project is to determine the interaction of the renal dopaminergic and RAS in the regulation of sodium excretion during varying amounts of sodium intake. The specific aims are to determine: 1. The effect of sodium intake on the interaction between the RAS and D1 dopamine receptors on sodium excretion. 2. The effect of GRK4 gene variants on the natriuretic response to D1 dopamine receptor agonist and its modulation by ACE inhibition. Hypothesis: The natriuretic effect of D1 dopamine receptors, which is best manifest under conditions of sodium loading is antagonized by the antinatriuretic effect of AT1 angiotensin receptors. In essential hypertension the natriuretic effect of D1 receptors is impaired because of activating variants of GRK4 while the antinatriuretic effect of AT1 receptors is enhanced. The overall hypothesis is that ACE inhibition augments the natriuresis caused by D1 receptor stimulation in normotensive subjects, only in the salt loaded state. These effects are reduced in those subjects with GRK4 gene variants, in hypertensive subject