Regulation of Epididymal Luminal Acidification

附睾管腔酸化的调节

基本信息

批准号：
7460891
负责人：
NURIA M. PASTOR-SOLER
金额：
$ 12.96万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-07-01 至 2009-12-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7460891
关键词：
Acids Acute Address Adenylate Cyclase Adult Affect Agonist Androgen Antagonists Animals Apical Basic Science Bicarbonates Biological Cadmium Carrier Proteins Cell membrane Cells Chronic Clear Cell Condition Cyclic AMP Cyclic AMP-Dependent Protein Kinases Cyclic GMP Data Development Diethylstilbestrol Duct (organ) structure Electrodes Environment Enzymes Epididymis Epithelial Epithelial Cells Estrogens Ethinyl Estradiol Fertility Fluorescence Flutamide GTP-Binding Proteins General Hospitals Goals Gonadotropin Hormone Releasing Hormone Gonadotropin Releasing Hormone Inhibitor Hormonal Hormone Antagonists Hormones In Situ In Vitro Kidney Knowledge Laboratories Lead Massachusetts Mediating Medicine Membrane Biology Modeling Modification Molecular Mouse Protein Mus Neonatal Numbers Organ Phenotype Physiological Play Poison Production Proteins Proton Pump Proton-Translocating ATPases Protons Rattus Recycling Regulation Research Research Proposals Role Second Messenger Systems Site Source Techniques Thinking Toxin Transgenic Mice Vas deferens structure Vesicle Work adenylyl cyclase 6 apical membrane base career cell motility cell type enhanced green fluorescent protein in vivo interdisciplinary approach male mature animal postnatal programs promoter protein expression reproductive research study response second messenger sperm cell steroid hormone toxicant trafficking urologic

项目摘要

DESCRIPTION (provided by applicant): This applicant proposes a program of research to prepare her for a career in academic medicine and basic science research studying male reproductive/urologic tract function, addressing the regulation of epididymal luminal acidification under physiological and pathophysiological conditions, during adulthood and postnatal development. The research will be conducted in the laboratory of Dr. Sylvie Breton at the Program in Membrane Biology and Renal Unit, Massachusetts General Hospital. The epididymis and vas deferens (VD) are the sites where spermatozoa mature and are stored in an immotile state. The luminal environment of these organs is tightly regulated and maintained at an acidic pH in order to keep spermatozoa from achieving motility. In particular, the applicant will study the mechanism by which an alkaline epididymal/VD luminal pH stimulates H+ATPase trafficking from subapical vesicles to the apical plasma membrane in clear cells. In addition, she will further investigate the role of the bicarbonate-activated "soluble" adenylyl cyclase (sAC), cAMP, and downstream effectors (such as PKA and Epac) on this trafficking phenomenon. Other sources of cAMP, such as G protein-regulated transmembrane adenylyl cyclases, and other second messenger molecules (such as cGMP) in clear cells will also be investigated. These studies will also evaluate the effects of steroid hormone agonists and antagonists (DES, ethinyl estradiol, GnRH antagonist and flutamide) and toxic compounds (Cadmium and Lead) have on the epididymal cellular profile, protein expression, H+ATPase trafficking and function, and on the cellular profile of the epididymis/VD during adulthood. The work proposed in this application will be carried out on rat and/or mouse epididymal/VD epithelial cells during adulthood and postnatal development, in vivo, in situ, and in vitro, using a multidisciplinary approach to evaluate protein expression and vesicle recycling mechanisms. The techniques proposed include microspectrofluorescence, cell biological, molecular biological techniques, and the use of a proton-selective self-referencing electrode. Newly available transgenic mice expressing enhanced green fluorescence protein (EGFP) specifically in clear cells (B1-EGFP mice) will provide a unique model to examine the development and modulation of epithelial cell phenotypes in response to various manipulation.

描述（由申请人提供）：该申请人提出了一项研究计划，为她的学术医学和基础科学研究工作做好准备，研究男性生殖/泌尿外科功能，以探讨在偶然和后期发展期间生理和病理生理条件下的附睾腔腔酸化的调节。该研究将在马萨诸塞州膜生物学和肾单位的计划的Sylvie Breton博士实验室进行。附睾和VAS延期（VD）是精子成熟并存储在不受限制状态的部位。这些器官的腔环境受到严格调节，并保持在酸性pH值，以防止精子达到运动能力。特别是，申请人将研究碱性附睾/VD腔pH刺激H+ATPase运输的机制，从亚皮囊泡到透明细胞中的顶端质膜。此外，她将进一步研究碳酸氢盐激活的“可溶”腺苷酸环化酶（SAC），CAMP和下游效应子（例如PKA和EPAC）对这种运输现象的作用。还将研究其他cAMP的来源，例如G蛋白调节的跨膜腺苷循环酶，以及其他第二个信使分子（例如CGMP）在透明细胞中也将进行研究。这些研究还将评估类固醇激素激动剂和拮抗剂（DES，乙基雌二醇，GNRH拮抗剂和氟丁酰胺）和有毒化合物（镉和铅）对附睾细胞谱，蛋白质表达，H+Atpase运输和功能以及epidiDy的蜂窝状曲线的影响。本应用中提出的工作将在成年期和产后发育期间，体内，原位和体外对大鼠和/或小鼠附睾/VD上皮细胞进行，并使用多学科方法来评估蛋白质表达和囊泡再生机制。提出的技术包括微光谱荧光，细胞生物学，分子生物学技术以及质子选择性自我引用电极的使用。在透明细胞（B1-EGFP小鼠）中特别表达增强绿色荧光蛋白（EGFP）的新的转基因小鼠将提供一个独特的模型，以检查各种操纵的上皮细胞表型的发展和调节。