MATURATION OF CATION TRANSPORT IN DISTAL NEPHRON

远端肾单位中阳离子传输的成熟

• 批准号: 2140544
• 负责人: Lisa M. Satlin
• 金额: $ 29.19万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-08-01 至 1998-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2140544
• 关键词: apical membrane; growth /development; infant animal; laboratory rabbit; potassium; potassium channel; renal tubular transport; sodium; sodium potassium exchanging ATPase; voltage /patch clamp

Growing subjects are in a state of positive K balance and have a limited ability to excrete K. This suggests that the immature kidney has a limited capacity for K secretion and/or enhanced ability to reabsorb K. The major regulatory site of K secretion in the kidney is the cortical collecting duct (CCD). In contrast to the high rates of net K secretion observed in CCDs isolated from adult animals and microperfused in vitro, segments from neonatal animals show no significant K transport. Yet, these same neonatal segments absorb Na at a rate half that measured in the adult, suggesting there exists a fundamental difference in cation transport mechanisms between the neonate and adult. K secretion in the CCD, mediated by principal cells, is determined by a two step process: active K uptake into the cell by Na-K-ATPase and passive diffusion down a favorable electrochemical gradient through apical K channels. To determine whether a paucity of apical K secretory channels limits net K secretion early in life, we will use patch clamp analysis to compare the apical K conductances of neonatal and mature principal cells. To examine whether the absence of conducting K secretory channels is due to a low open probability (P-O) of existing channels and/or a low channel number (N), we will next determine whether exposure of the neonatal CCD to factors known to increase P-O or N induces net K secretion. The discrepancy between onset of Na and K transport in the neonatal CCD suggests that qualitative changes in the transepithelial Na absorptive pathway occur during postnatal differentiation. To test this, we will compare the apical Na conductances (patch clamp analysis), membrane transporters active in Na reabsorption (helium glow photometry), and Na-K-ATPase activity (ouabain- sensitive basolateral 86Rb uptake) in neonatal and mature principal cells; the latter findings will be correlated with measurements of basolateral membrane surface area (electron microscopy). Because clearance studies in the neonate indicate significant K retention, we will also test the hypothesis that enhanced K absorption in the neonate reduces net urinary K excretion. To assess the K absorptive capacity of the CCD and outer medullary collecting duct (OMCD), we will determine the contribution of K absorption, measured as unidirectional lumen-to-bath 86Rb fluxes, to net K transport in segments isolated from maturing animals. Should we document significant K absorption early in life, we will test whether K absorption is coupled to H secretion by H-K-ATPase, a transporter immunocytochemically identified in intercalated cells. That population of intercalated cells possessing functional H-K-ATPase will then be identified and the polarity and activity of H-K exchange measured to determine if there is a maturational change in activity of this pump. The studies proposed in this application should help us to understand the physiologic basis for the limited K secretory capacity of the neonatal CCD and provide broad insight into the regulation of the K secretory process and its interrelationship with the mechanism of Na absorption.

生长主体处于正钾平衡状态，且具有有限的 排泄 K 的能力。这表明未成熟的肾脏具有有限的能力 钾分泌能力和/或增强重吸收钾的能力。主要 肾脏钾分泌的调节部位是皮质集合区 导管（CCD）。与观察到的高净 K 分泌率相反 从成年动物中分离并在体外进行微灌注的 CCD，来自 新生动物没有表现出明显的钾转运。然而，这些同样的新生儿 节段吸收钠的速度是成人测量的一半，表明 阳离子传输机制存在根本差异 介于新生儿和成人之间。 CCD 中的 K 分泌，介导 主细胞的活性由两步过程决定：活性 K 吸收到 细胞通过Na-K-ATP酶和被动扩散向下有利 通过顶端 K 通道的电化学梯度。判断是否 顶端钾分泌通道的缺乏限制了早期的净钾分泌 生活中，我们将使用膜片钳分析来比较心尖K 新生儿和成熟主细胞的电导。来检查是否 缺乏传导 K 分泌通道是由于开放度低 现有通道的概率（P-O）和/或低通道数（N），我们 接下来将确定新生儿 CCD 是否暴露于已知因素 增加 P-O 或 N 会诱导净 K 分泌。之间的差异 新生儿 CCD 中 Na 和 K 转运的开始表明定性 跨上皮 Na 吸收途径发生变化 产后分化。为了测试这一点，我们将比较心尖 Na 电导（膜片钳分析），Na 中活跃的膜转运蛋白 重吸收（氦发光光度法）和 Na-K-ATP 酶活性（哇巴因- 新生儿和成熟主细胞中敏感的基底外侧 86Rb 摄取； 后者的发现将与基底外侧的测量相关 膜表面积（电子显微镜）。因为清除研究在 新生儿表现出显着的钾保留，我们还将测试 新生儿钾吸收增强会减少净尿的假设 K排泄。评估 CCD 和外部的 K 吸收能力 髓质集合管（OMCD），我们将确定 K 的贡献 吸收，以单向流明至浴 86Rb 通量测量，以净 从成熟动物中分离出的片段中钾的运输。我们是否应该记录 生命早期有显着的钾吸收，我们将测试是否吸收钾 通过 H-K-ATP 酶（一种转运蛋白）与 H 分泌相结合 通过免疫细胞化学方法在闰细胞中进行鉴定。那人口 具有功能性 H-K-ATP 酶的闰细胞将被 识别并测量 H-K 交换的极性和活性 确定该泵的活动是否发生成熟变化。这 本申请中提出的研究应该有助于我们理解 新生儿 CCD 有限的 K 分泌能力的生理基础 并提供对 K 分泌过程调节的广泛见解 及其与Na吸收机制的相互关系。