CELLULAR FUNCTION OF THE JUXTAGLOMERULAR APPARATUS

哺乳动物肾脏的肾小球旁结构(JGA)是一种解剖结构 将远端肾单位致密斑细胞与 肾小球血管极。已经证实，氯化钠的变化 致密斑细胞的集中引发了一连串的事件， 改变传入小动脉张力和肾素分泌。为了更好地 了解不同细胞组件之间可能的相互作用 在JGA中，需要知道这些细胞的固有属性。在……里面 其第一部分，这项建议旨在调查秘密和 小动脉传入细胞对血管紧张素II和血管紧张素转换酶的收缩反应 腺苷，两种在水平上具有深刻调节作用的物质 已经注意到它们会影响肾小球前血管。 协同时尚。此应用程序的第二部分将使用 高灵敏度的信使核糖核酸水平评估技术研究 JGA细胞中基因表达的调控。 第1部分的具体目标是描述相互关系的程度 血管紧张素和腺苷对血管收缩作用的依赖性 以了解协同作用的机制。协同效应的程度 将通过检查a)是否以及在多大程度上 血管收缩与肾素分泌抑制 腺苷被血管紧张素II增强，被血管紧张素II抑制 反之，是否以及在多大程度上相同的反应 由血管紧张素II产生的物质被腺苷增强，而被 腺苷拮抗剂，以及b)传入收缩和 致密斑部氯化钠浓度升高对肾素释放的抑制作用 取决于两种激动剂的可用性。其作用机制是： 协同作用将通过检查a)可能的 促进在各自受体水平上的相互作用，b)a 调制肌醇磷脂生成过程中可能的增强作用 代谢物，c)可能对钙的释放有促进作用 细胞内储存或钙进入，d)对跨膜离子电流的影响 增强细胞活性，e)对细胞内循环水平的影响 核苷酸，以及f)对磷脂酶A依赖的生成的影响 产品。 第2部分的一个具体目标是使用聚合酶链式反应(PCR)来 评估单一JGA制剂中的mRNA水平。在这些研究中，我们将a) 优化技术以监控回收或放大效率 每个步骤的聚合酶链式反应方法适用于单个分离的JGA标本，b) 评估一组管家基因和 确定这些信使核糖核酸对肾素调节的反应。 血管紧张素系统，c)研究血管紧张素II、腺苷和 环核苷酸对单个JGA肾素mRNA的影响，以及d)检测 致密黄斑细胞葡萄糖-6-磷酸脱氢酶的研究 JGA中肾素和G6PDH的表达水平呈平行关系。

The juxtaglomerular apparatus (JGA) of the mammalian kidney is an anatomic structure which connects the macula densa cells of the distal nephron with the glomerular vascular pole. It has been established that changes in NaCl concentration at the macula densa cells initiate a chain of events which alters afferent arteriolar tone and renin secretion. In order to better understand possible interactions between the different cellular components of the JGA the intrinsic properties of these cells need to be known. In its first part this proposal aims at investigating the secretory and contractile responses of afferent arteriolar cells to angiotensin II and adenosine, two agents which exert profound modulating effects at the level of the JGA and which have been noted to affect preglomerular vessels in a synergistic fashion. The second part of this application will utilize a highly sensitive technique for assessment of mRNA levels to study regulation of gene expression in cells of the JGA. The specific aims of part 1 are to describe the extent of the mutual dependency of the vasoconstrictor effects of angiotensin and adenosine and to understand the mechanisms for the synergism. The degree of synergism will be studied by examining a) whether and to what extent the vasoconstriction and the inhibition of renin secretion produced by adenosine is potentiated by angiotensin II and inhibited by angiotensin II antagonists, and conversely, whether and to what extent the same responses produced by angiotensin II are potentiated by adenosine and inhibited by adenosine antagonists, and b) whether the afferent constriction and inhibition of renin release elicited by increased NaCl at the macula densa depend upon the availability of both agonists. The mechanisms of the synergistic interaction will be studied by examining a) a possible facilitating interaction at the level of the respective receptors, b) a possible potentiating effect in generating modulating phosphoinositide metabolites, c) a possible potentiating effect on Ca release from intracellular stores or Ca entry, d) effects on transmembrane ion currents which enhance cell activation, e) effects on intracellular levels of cyclic nucleotides, and f) effects on the generation of phospholipase A dependent products. A specific aims of part 2 are to use the polymerase chain reaction (PCR) to assess mRNA levels in single JGA preparations. In these studies we will a) optimize techniques for monitoring recovery or amplification efficiency at each step of the PCR method applied to single isolated JGA specimen, b) assess the response pattern of mRNA for a set of housekeeping genes and define the response of these mRNA species to manipulations of the renin- angiotensin system, c) study the effect of angiotensin II, adenosine, and cyclic nucleotides on renin mRNA in single JGA, and d) examine the role of glucose-6-phosphate dehydrogenase in macula densa cells by assessing in parallel the levels of renin and G6PDH mRNA in the JGA.