RECEPTOR REGULATED CALCIUM ENTRY IN EXOCRINE SECRETION

受体调节外分泌分泌中的钙进入

基本信息

批准号：
3298000
负责人：
Trevor J. Shuttleworth
金额：
$ 28.35万
依托单位：
UNIVERSITY OF ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
1988
资助国家：
美国
起止时间：
1988-07-01 至 1995-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/3298000
关键词：
calcium transporting ATPase cell membrane ducks exocrine glands flash photolysis fluorimetry hormone receptor hormone regulation /control mechanism inositol phosphates membrane lipids membrane permeability neurotransmitters phosphatidylinositols phospholipids secretion tissue /cell culture

项目摘要

The overall aim of our research is to further understanding of the cellular mechanisms involved in the initiation and control of exocrine ion secretion and, in particular, the nature and origins of the rise in intracellular calcium concentrations ([Ca2+]i) that signal secretion. Such increases in [Ca2+]i arise partly from the mobilization of Ca2+ from intracellular stores, and partly from an enhanced entry of extracellular Ca2+. These events are accompanied by the generation of a series of inositol phosphates certain of which have been proposed as important messenger molecules in this process. Of these, the role of Ins(1,4,5)P3 in the mobilization of Ca2+ from certain intracellular stores is well established. However, the nature and regulation of the receptor-enhanced entry of Ca2+ across the plasma membrane, and the involvement of inositol phosphates in this process, are unknown. Current debate centers on whether the enhanced Ca2+ entry results solely from the emptying of these stores (without any direct requirement for inositol phosphates), or whether Ca2+ entry is regulated by the combined action of Ins(1,4,5)P3 and an additional undefined action of its phosphorylated product Ins(1,3,4,5)P4. However, both ideas emphasize the prior emptying of agonist-sensitive stores before Ca2+ entry can be activated. In direct contradiction to this, we have recently shown that, at least under some circumstances, Ca2+ entry can be activated before any measurable emptying of agonist-sensitive stores. We intend to use the isolated cells of the avian nasal gland as a model system with the specific aims of evaluating the association between the receptor activation of Ca2+ entry and the mobilization of intracellular Ca2+ stores, the possible role of Ins(1,3,4,5)P4 in this process, and to determine the critical features modulating the metabolism of Ins(1,4,5)P3 and the generation of Ins(1,3,4,5)P4 in the intact cell. This involves fluorimetric measurements of [Ca2+]i and electrophysiological measurements of Ca2+-activated whole- cell currents, coupled with flash photolysis, single-cell dialysis and transient electroporation (to introduce impermeant molecules). This will be supported by biochemical studies on the generation and metabolism of inositol phosphates. We also propose to make use of our recent finding of a specific, and rapid increase in the expression of the receptor-activated Ca2+ entry mechanism during adaptive growth and differentiation of these cells in vivo. Our aim is to reproduce this expression system in culture, thereby providing a unique system for studying the specific nature of the receptor-activated Ca2+ entry mechanism, its expression and control.

我们研究的总体目的是进一步了解细胞外分泌离子分泌的启动和控制涉及的机制并且特别是细胞内上升的性质和起源钙浓度（[Ca2+] i）信号分泌。这样的增加 [Ca2+]我部分来自于细胞内Ca2+的动员存储，部分是从增强的细胞外Ca2+进入。这些事件伴随着一系列肌醇磷酸盐的产生其中的某些已被提出为重要的使者分子这个过程。其中，INS（1,4,5）P3在动员中的作用来自某些细胞内存储的Ca2+已建立。但是， Ca2+的受体增强输入的性质和调节质膜和肌醇磷酸盐的参与其中过程，未知。当前的辩论集中于增强的Ca2+是否仅来自这些商店的排空结果（没有任何直接的对肌醇磷酸盐的要求），或CA2+进入是否受到调节 INS（1,4,5）P3的综合作用和其他不确定的动作它的磷酸化产品INS（1,3,4,5）P4。但是，这两个想法都强调在Ca2+进入之前，先前对激动剂敏感商店的清空可以是活性。在与此直接矛盾的情况下，我们最近表明，至少在某些情况下，可以在任何情况下激活Ca2+进入可测量的激动剂敏感商店的排空。我们打算使用禽鼻腺的孤立细胞作为模型系统评估Ca2+受体激活之间关联的目的细胞内Ca2+存储的进入和动员，可能的作用在此过程中的INS（1,3,4,5）P4，并确定关键特征调节INS（1,4,5）P3的代谢和产生完整细胞中的INS（1,3,4,5）P4。这涉及荧光测量 [Ca2+] I和Ca2+活化全-激活的电生理测量细胞电流，再加上闪光光解，单细胞透析和瞬态电穿孔（引入纯净分子）。这会受到关于生成和代谢的生化研究的支持肌醇磷酸盐。我们还建议利用我们最近的发现受体激活的表达的特定且快速增加自适应生长和分化过程中的CA2+进入机制细胞体内。我们的目的是在文化中重现这种表达系统，从而提供了一个独特的系统来研究受体激活的Ca2+进入机制，其表达和控制。