Cellular and Molecular Studies of Renal Transport

肾脏运输的细胞和分子研究

• 批准号: 8135557
• 负责人: Michael J. Caplan
• 金额: $ 111.88万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-12-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8135557
• 关键词: Address; American Heart Association; Animals; Award; Biological Assay; Biophysics; Books; Boron; Carrier Proteins; Cell Culture Techniques; Cell membrane; Cells; Clinical; Collaborations; Commit; Computer software; Confocal Microscopy; Core Facility; Coupled; Development Plans; Disease; Distal; Electrolytes; Energy Metabolism; Epithelial; Equipment; Faculty; Fostering; Funding; Gene Targeting; Generations; Genetic; Goals; Health; Homeostasis; Hypertension; Image; Immune; In Vitro; Individual; Institute of Medicine (U.S.); International; Ions; Joints; Kidney; Laboratories; Leadership; Light; Liquid substance; Mammalian Cell; Medical; Medicine; Methods; Micropuncture; Microscope; Microscopy; Molecular; Molecular Cloning; Mus; Mutagenesis; Operative Surgical Procedures; Organ; Oryctolagus cuniculus; Paraffin; Patch-Clamp Techniques; Pathogenesis; Pathway interactions; Perfusion; Phosphopeptides; Phosphorylation Site; Physiological; Physiology; Play; Positioning Attribute; Preparation; Problem Solving; Process; Productivity; Program Research Project Grants; Proteins; Publishing; Rattus; Recording of previous events; Regulation; Relative (related person); Renal function; Renal tubule structure; Research; Research Personnel; Research Project Grants; Role; Scientist; Secure; Services; Sodium Chloride; System; Techniques; Textbooks; Tissue Fixation; Tissue Stains; Transgenic Mice; United States National Academy of Sciences; Universities; Update; Vesicle; Work; Xenopus oocyte; base; cDNA Expression; cytochemistry; experience; fluorescence imaging; genetic regulatory protein; gratitude; in vivo; insight; interest; kidney cell; medical schools; member; mouse development; patch clamp; professor; programs; protein transport; solute; tissue culture; tool

DESCRIPTION, OVERALL (provided by applicant): The overall goal of this Program Project is to understand the mechanisms underlying renal fluid and electrolyte homeostasis and renal epithelial function in health and the processes that modulate these mechanisms in disease. A broad spectrum of techniques will be used to address a continuum of problems ranging from the molecular characterization of individual transport-related proteins to the contribution of these proteins to integrated renal function at the level of the intact tubule, the organ, and the whole animal. Our strategy to pursue these themes successfully will include close collaboration on interrelated research projects; sharing of expertise, concepts and techniques by Directors of the five individual Projects and the four research Cores; joint use of the research core facilities and single administrative core. The research projects comprise a broad range of experimental preparations including ion/solute transport proteins, transport regulatory proteins, transfected mammalian cells in tissue culture, isolated cell membrane vesicles, Xenopus oocyte expression system, isolated kidney cells and tubules, and whole kidney in vivo. We shall use a wide range of methods including molecular cloning and mutagenesis, functional cDNA expression, generation and use of transgenic mice, immune-cytochemistry, phosphopeptide enrichment coupled to mass spectral identification of phosphorylation sites, confocal microscopy, fluorometric assays of cell ion activities, whole cell clamp and patch- and giant patch-clamp techniques, in vivo and in vitro perfusion of defined tubule segments, and clearance studies. Each of the projects and cores is concerned with the central themes of the Program: to provide important new insights into individual transport proteins that play a role in renal electrolyte homeostasis, to elucidate the regulation of these transport-related proteins, to elucidate the cellular pathways involved in epithelial polarity that is an absolute prerequisite for vectorial solute and fluid transport, and to assess the relative contributions of these proteins to tubule and organ function under conditions of normal and deranged electrolyte and energy metabolism. Our strategy to pursue these themes successfully will include close collaboration on interrelated research projects; sharing of expertise, concepts and techniques by Program investigators; and joint use of core facilities.

总体描述（由申请人提供）： 该计划项目的总体目标是了解肾液和肾液的潜在机制 健康中的电解质稳态和肾上皮功能以及调节这些功能的过程 疾病中的机制。将使用广泛的技术来解决一系列问题，从单个转运相关蛋白的分子表征到这些蛋白对完整肾小管、器官和整个动物水平的肾功能的贡献。我们成功追求这些主题的战略将包括在相关研究项目上的密切合作；五个单独项目和四个研究核心的主任分享专业知识、概念和技术；联合使用研究核心设施和单一行政核心设施。研究项目包括广泛的实验制剂，包括离子/溶质转运蛋白、转运调节蛋白、组织培养中的转染哺乳动物细胞、分离的细胞膜囊泡、非洲爪蟾卵母细胞表达系统、分离的肾细胞和肾小管以及体内的整个肾脏。我们将使用多种方法，包括分子克隆和诱变、功能性cDNA表达、转基因小鼠的产生和使用、免疫细胞化学、磷酸化肽富集与磷酸化位点的质谱鉴定、共聚焦显微镜、细胞离子活性的荧光测定、全细胞钳以及膜片和巨膜片钳技术，在体内和体内 确定的小管段的体外灌注和清除研究。每个项目和核心都涉及该计划的中心主题：为在肾电解质稳态中发挥作用的个体转运蛋白提供重要的新见解，阐明这些转运相关蛋白的调节，阐明参与上皮极性的细胞途径，这是矢量溶质和液体转运的绝对先决条件，并评估这些蛋白质对肾小管和器官的相对贡献 在正常和紊乱的电解质和能量代谢条件下发挥功能。我们成功追求这些主题的战略将包括在相关研究项目上的密切合作；项目研究人员分享专业知识、概念和技术；以及核心设施的联合使用。