Localization, Interactions, And Functions of Synaptotagmins in the Pituitary

项目摘要 突触结合蛋白1是一种在突触囊泡上发现的蛋白质，在神经递质释放中充当Ca 2+传感器。 该蛋白质有16个同源物，而这些其他蛋白质被广泛认为具有相关功能 基本上不知道他们在哪里或做什么。初步数据显示， 应用显示几种突触结合蛋白同种型在垂体后叶和/或垂体前叶中丰富， 分泌肽类激素的腺体，这些激素基本上控制哺乳动物生理的所有重要方面。之前 根据这些新的初步结果，垂体已被确定为一种适于定量分析的制剂。 含肽囊泡如何与质膜融合以释放其内容物的生物物理学研究 进入循环。因此，在垂体中发现新的突触结合蛋白亚型提供了一个独特的 有机会详细测试它们的功能，并阐明它们在兴奋-分泌偶联中的确切作用。这 项目将研究垂体synaptotagmin亚型与互补的方法，调查他们的 体外结合特性，以及它们在垂体中的定位和生理功能。功能 研究采用了使用转基因小鼠进行特异性突触结合蛋白同种型的基因消除的策略， 基因靶向以鉴定表达用于电记录的那些同种型的特定细胞群。通过 在识别的细胞和神经末梢中进行电容记录，我们将确定SYT同种型 在Ca 2+敏感性和融合孔方面调节胞吐作用。这里提出的实验将测试 突触结合蛋白4、7、9、10、11和12在控制生长、代谢 压力、体液平衡和繁殖。

PROJECT SUMMARY Synaptotagmin 1, a protein found on synaptic vesicles, serves as a Ca2+ sensor in neurotransmitter release. This protein has 16 homologues, and while these other proteins are widely thought to serve related functions essentially nothing is known about where they are or what they do. Preliminary data presented in this application shows that several synaptotagmin isoforms are abundant in the posterior and/or anterior pituitary, glands that release peptide hormones that control essentially all vital aspects of mammalian physiology. Prior to these new preliminary results, the pituitary had been established as a preparation amenable to quantitative biophysical study of how peptide-containing vesicles fuse with the plasma membrane to release their content into the circulation. Thus, the finding of novel synaptotagmin isoforms in the pituitary offers a unique opportunity to test their function in detail and elucidate their precise roles in excitation-secretion coupling. This project will study the pituitary synaptotagmin isoforms with complementary approaches, investigating their binding properties in vitro, and their localization and physiological function in the pituitary. The functional studies employ a strategy of using transgenic mice for gene ablation of specific synaptotagmin isoforms, and gene targeting to identify specific cell populations that express those isoforms for electrical recording. By performing capacitance recording in identified cells and nerve terminals we will determine how syt isoforms regulate exocytosis in terms of Ca2+ sensitivity and fusion pores. The experiments proposed here will test the functions of synaptotagmins 4, 7, 9, 10, 11, and 12 in the release of hormones that control growth, metabolism, stress, fluid balance, and reproduction.