WATER AND ELECTROLYTE MOVEMENT IN RENAL TUBULES

这项建议的长期目标是提供新的和有意义的 与溶质运移相关的结构-功能相关数据 沿着哺乳动物的肾小管。所描述的实验集中在我们的 主要关注集合管，并重点关注 血管紧张素II（All）在闰细胞调节中的作用 结构和功能，特别是在控制跨上皮 H+和HCO 3-运输。该提案包括四个部分， 界定了具体的目标和假设进行测试。 在具体目标I中，我们将研究All对超微结构的影响。 以鉴定特定的闰细胞 对All有反应的亚群，并确定All对 免疫金细胞化学法观察H+ ATP酶亚细胞分布。 待检验的假设：特定的闰细胞亚型是 A1对H ~+-ATP酶活性和酸碱平衡的影响主要是由于A1对H ~+-ATP酶活性和酸碱平衡的影响 在收集管道中运输。 在特异性靶点II中，研究了All对H ~+-ATP酶活性的影响， 参与介导这些效应的信号转导途径将 在显微解剖的集合管段中进行研究。假设是 实验结果表明：A12对H ~+-ATPase活性的抑制作用与A12对H ~+-ATPase活性的抑制作用有关。 集合管通过特异性AT 1受体介导， 将被识别的不同信号系统。 在具体目标III中，我们将确定表达和细胞 所有受体亚型mRNA在集合管中的位置， 定量RT-PCR和原位杂交，并确定效果 酸碱紊乱对All受体mRNA表达的影响。假设 待测：所有受体mRNA表达在不同 闰细胞群并受酸碱影响 扰动 在具体目标IV中，我们将确定所有对质子的影响， 离体灌流兔皮层收集区的碳酸氢根转运 导管和识别响应于 所有.待检验的假设：所有刺激HCO 3分泌的B型 并抑制A型闰细胞分泌H+ 皮层集合管中的一个 本提案中描述的实验旨在继续我们的研究。 长期承诺，以界定结构和功能 肾小管的特性，特别是建立 插入细胞在酸碱调节“微调”中的作用 被肾夹住了。更好地理解控制H+的机制 和HCO 3，在肾小管的这一区域的运输将使 医生处理复杂的酸碱平衡临床问题 往往是危及生命的专业知识和更大的成功。

The long-term objective of this proposal is to provide new and meaningful structural-functional correlative data as it relates to solute transport along the mammalian renal tubule. The experiments described focus our attention primarily on the collecting duct and place a major emphasis on the role of angiotensin II (All) in the regulation of intercalated cell structure and function and specifically in the control of transepithelial H+ and HCO3- transport. The proposal includes four sections with clearly delineated specific aims and hypotheses to be tested. In Specific Aim I we will examine the effect of All on the ultrastructure of intercalated cells to identify specific intercalated cell subpopulations that respond to All, and determine the effect of All on the subcellular distribution of H+ATPase by immunogold cytochemistry. Hypothesis to be tested: Specific intercalated cell subtypes are responsible for the effect of All on H+-ATPase activity and acid-base transport in the collecting duct. In Specific Aim II the effects of All on H+-ATPase activity and the signalling transduction pathways involved in mediating these effects will be studied in microdissected collecting duct segments. Hypothesis to be tested: The inhibitory effect of All on H+-ATPase activity in the collecting duct is mediated via specific AT1 receptors and activation of distinct signalling systems which will be identified. In Specific Aim III we will determine the expression and cellular location of All receptor isoform mRNA in the collecting duct by quantitative RT-PCR and in situ hybridization and determine the effect of acid-base perturbations on All receptor mRNA expression. Hypothesis to be tested: All receptor mRNA expression varies between different populations of intercalated cells and is influenced by acid-base perturbations. In Specific Aim IV we will determine the effect of All on proton and bicarbonate transport in the isolated perfused rabbit cortical collecting duct and identify the subtype(s) of intercalated cells that responds to All. Hypothesis to be tested: All stimulates HCO3 secretion by type B intercalated cells and inhibits H+ secretion by type A intercalated cells in the cortical collecting duct. The experiments described in this proposal are intended to continue our long-term commitment to delineate the structural and functional characteristics of the renal tubule and specifically to establish the role of intercalated cells in the "fine-tuning" of acid-base regulation by the kidney. A better understanding of the mechanisms that control H+ and HCO3, transport in this region of the renal tubule will enable physicians to manage complicated clinical problems of acid-base balance that are often life-threatening with greater expertise and success.