Microvascular Transport in the Renal Medulla

项目主任/首席调查员(最后、第一、中间)：Pallone，Thomas L. 项目总结(见说明)： 肾髓质的直血管使NaCI和尿素沉积到肾小管间质和肾间质。 将血液流动分配到外髓和内髓。直肠降支血管(DVR)将所有血流供应到 两个地区都有。构成DVR壁的细胞(平滑肌/周细胞和内皮细胞)调节 通过离子通道和信号分子相互作用的血管活性。DVR内皮提供了这两种功能 屏障(通透性)功能和调节血管活性。内皮细胞是“机械敏感的”，因此 管腔流量增加增加胞浆内钙离子([Ca2+]Cyr)、一氧化氮生成和跨壁 溶质渗透率。对DVR内皮的电生理研究表明，它是一种电信号 合胞体，其中的细胞通过缝隙连接彼此强烈耦合。外源性K+是一种强有力的调节剂 缝隙结电导。强内向整流钾通道亚型调节膜电位和 诱导超极化以响应外界K+离子浓度的小幅升高。因此，K+离子可以 在调节肾髓质的血流灌注中起关键作用。 我们建议从三个方面研究DVR内皮的功能。在目标1中，液压 通透性、血管活性、NO生成和[Ca~(2+)]CVT将被量化为外部K+的函数 集中精神。K(R)和BKCa通道在介导内皮依赖性舒张中的单独作用 将测试渗透率的功能和调节。BKCa通道作为K+可能来源的作用 将通过激活它们并用K|R通道抑制来阻断它们的作用来检测离子。在目标2中， 缝隙连接在内皮细胞和周细胞之间的通讯能力将被研究。病毒的传播 细胞间的电流和荧光探针将用电生理学和荧光进行量化 成像。K+离子、特异性多肽和非特异性化学抑制剂阻断缝隙连接的能力 为了防止电流传导和荧光探针扩散，将对其进行测试。最后，差距的能力 沿着DVR轴传导血管扩张剂和收缩反应的连接点将通过 微血管灌注、视频显微镜和图像分析。在目标3中，DVR的机械敏感性 将探索血管内皮细胞。[Ca~(2+)]CVR升高和PI3激酶信号转导所必需的作用 便利将不会测试任何一代。在不同的系列中，钙离子进入内皮细胞和 本课程将探索通过PISkinase来调节通透性的流动依赖性增加的信号。独立 我们将探讨在一氧化氮合酶阻断过程中，一氧化氮对PI3K通透性的调节作用。 相关性(请参阅说明)： 直肠血管下行为肾脏的髓质提供所有的血液流动，该区域对调节肾脏功能至关重要。 排出尿液中的盐和水。延髓的氧分压很低，很容易受到 当流向它的血液减少时，就会受伤。直肠血管下行收缩和扩张的调节是一种 这是重要的生理过程，也是本提案的研究主题。 项目/

Program Director/Principal Investigator (Last, First, Middle): Pallone, Thomas L. PROJECT SUMMARY (See instructions): The vasa recta of the renal medulla trap NaCI and urea deposited to the interstitium by nephrons and distribute blood flow to the outer and inner medulla. Descending vasa recta (DVR) supply all blood flow to both regions. The cells that comprise the DVR wall (smooth muscle / pericytes and endothelia) regulate vasoactivity through the interactions of ion channels and signaling molecules. DVR endothelia provides both barrier (permeability) function and regulates vasoactivity. The endothelium is "mechanosensitive" such that increases in luminal flow yield increases in cytoplasmic Ca2+ ([Ca2+]cYr), NO generation and transmural solute permeability. Electrophysiological studies of the DVR endothelium have shown that it is an electrical syncytium in which cells strongly couple to one another via gap junctions. External K+ is a strong regulator gap junction conductance. Strong inward rectifier K+ channel isoforms regulate membrane potential and induce hyperpolarization in response to small elevations of external K+ ion concentration. Thus K+ ion may play a key role in the regulation of perfusion of the renal medulla. We propose to study the function of the DVR endothelium with three Aims. In Aim 1, hydraulic permeability, vasoactivity, NO generation and [Ca2+]cvT will be quantified as a function of external K+ concentration. The separate roles of K(R and BKca channels to mediate the endothelium dependent relaxing functions and modulation of permeability will be tested. The role of BKCa channels as a putative source of K+ ion will be examined by activating them and blocking their effects with K|R channel inhibition. In Aim 2, the ability of gap junctions to communicate between endothelia and pericytes will be studied. The spread of current and fluorescent probes between cells will be quantified with electrophysiology and fluorescent imaging. The ability of gap junction blockade with K+ ion, specific peptide and nonspecific chemical inhibitors to prevent conduction of current and fluorescent probe diffusion will be tested. Finally, the ability of gap junctions to conduct both vasodilator and constrictor responses along the DVR axis will be quantified by microvessel perfusion, videomicroscopy and image analysis. In Aim 3 the mechanosensitivity of DVR endothelium will be explored. The requisite roles for [Ca2+]Cvr elevation and signaling via PI3kinase to facilitate NO generation will be tested. In separate series, effects of Ca2+ entry into the endothelium and signaling via PISkinase to mediate flow dependent increases in permeability will be explored. Independence of permeability modulation from NO will be explored for PI3kinase during NOS blockade. RELEVANCE (See instructions): Descending vasa recta provide all blood flow to the medulla of the kidney, a region vital to regulation of the elimination of salt and water in the urine. The medulla has very low oxygen tension and is vulnerable to injury when blood flow to it decreases. Regulation of contraction and dilation of descending vasa recta is a vital physiological process and is the subject of study in this proposal. PROJECT/