Microvascular Transport in the Renal Medulla

项目负责人/主要研究者（最后一名、第一名、中间名）：Pallone，托马斯L. 项目总结（见说明）： 肾髓质的直小血管被肾单位沉积于肾小管， 将血流分配到外髓和内髓。直降血管（DVR）提供所有血液流量， 两个区域。组成DVR壁的细胞（平滑肌/周细胞和内皮细胞）调节 血管活性通过离子通道和信号分子的相互作用。DVR内皮提供了 屏障（渗透性）功能和调节血管活性。内皮是“机械敏感的”，使得 增加管腔流量增加细胞质Ca 2+（[Ca 2 +]cYr），NO生成和透壁 溶质渗透性DVR内皮的电生理学研究表明，它是一种电刺激， 合胞体，细胞通过缝隙连接彼此紧密结合。外部K+是一种强调节剂 间隙结电导强内向整流钾通道亚型调节膜电位， 诱导对外部K+离子浓度小幅升高反应超极化。因此，K+离子可能 在肾髓质灌注的调节中起关键作用。 我们建议从三个方面来研究DVR内皮的功能。在目标1中，液压 渗透性、血管活性、NO生成和[Ca 2 +]cvT将被定量为外部K+的函数 浓度. K（R）和BKca通道介导内皮依赖性舒张的不同作用 将测试渗透性的功能和调节。BKCa通道作为K+来源的作用 离子将通过激活它们并用K阻断它们的作用来检查|R通道抑制。在目标2中， 将研究间隙连接在内皮细胞和周细胞之间通讯的能力。的传播 细胞之间的电流和荧光探针将用电生理学和荧光探针定量。 显像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 以防止电流的传导，并且将测试荧光探针扩散。最后，差距的能力 沿着DVR轴传导血管舒张和收缩反应的连接将通过以下方式量化： 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 将探索内皮。[Ca 2 +]Cvr升高和通过PI 3激酶信号传导对细胞凋亡的必要作用 将测试促进NO生成。在单独的系列中，Ca 2+进入内皮和 将探索通过PISkinase介导渗透性的流动依赖性增加的信号传导。独立 在NOS阻断过程中，将探讨NO对PI 3激酶的通透性调节作用。 相关性（参见说明）： 下行的直小血管提供所有的血液流向肾脏的髓质，这是一个对调节肾功能至关重要的区域。 排除尿液中的盐和水。髓质的氧分压很低， 当血液流向它时，它会减少。调节下行直血管的收缩和扩张是一种 这是一个重要的生理过程，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/