CALCIUM-SURROGATE ACTIONS OF LEAD IN SECRETION

分泌中铅的钙替代作用

• 批准号: 2153553
• 负责人: Janusz Bogdan Suszkiw
• 金额: $ 21.61万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 1999-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2153553
• 关键词: SDS polyacrylamide gel electrophoresis; annexins; binding proteins; biological signal transduction; calcium binding protein; calcium channel; calcium flux; calcium indicator; calmodulin; chromaffin cells; environmental toxicology; exocytosis; fluorescence spectrometry; gel electrophoresis; high performance liquid chromatography; lead; lead poisoning; membrane permeability; metal metabolism; neurotransmitter metabolism; protein kinase C; secretion; voltage /patch clamp; SDS polyacrylamide gel electrophoresis; annexins; binding proteins; biological signal transduction; calcium binding protein; calcium channel; calcium flux; calcium indicator; calmodulin; chromaffin cells; environmental toxicology; exocytosis; fluorescence spectrometry; gel electrophoresis; high performance liquid chromatography; lead; lead poisoning; membrane permeability; metal metabolism; neurotransmitter metabolism; protein kinase C; secretion; voltage /patch clamp

Disruption of neurotransmitter and hormone release is a likely factor in neurobehavioral, neuroendocrine, and endocrine disturbances associated with low-level lead (Pb2+) toxicity. The overall goal of this research is to assess the role of pb2+ interactions with calcium-signal transduction mechanisms, with focus on the Ca2+-dependent secretion of neurotransmitters and hormones. Previous results from our laboratory indicate the Pb2+ disrupts secretion by high affinity interactions with Ca2+-effector(s) of exocytosis. As an extention of these studies, the aim of the current project is to test a hypothesis that specific Ca2+-modulated lipid-binding proteins, calpactin I, synaptotagmin (p65), protein kinase C, and cytoplasmic form of phospholipase A2 (cPLA2) are important components of the secretory machinery and key targets for Pb2+-trigger action in secretion. The proteins will be isolated and their interaction with pb2+ characterized, using biochemical, biophysical, and immunochemical techniques. Permeabilized bovine adrenal chromaffin cells and cell-free models of exocytosis will be developed to further characterize the role of Pb2+ in the exocytotic mechanism. In addition to the interaction with the exocytotic apparatus, lead ions modify activity of calcium channels in the adrenal chromaffin cells through as yet undefined intracellular mechanism. Whole-cell voltage-clamp and single channel recordings from chromaffin cells in culture will be employed to test a hypothesis that low concentrations of intracellular Pb2+ modulate the l-type Ca2+ currents by slowing-down calcium-dependent channel inactivation. This research is expected to advance our understanding of Pb2+ actions in secretion at the molecular level and at the same time provide important information regarding the nature of exocytotic process itself. In Pb2+ interactions with Ca2+ signal transduction at different levels of cellular organization and may identify the Ca/lipid-binding proteins as major molecular targets in lead toxicity.

神经递质和激素释放的破坏可能是一个因素 神经行为，神经内分泌和内分泌障碍 低级铅（PB2+）毒性。 这项研究的总体目标是 评估PB2+相互作用与钙信号转导的作用 机制，重点是神经递质的Ca2+依赖性分泌 和激素。 我们实验室的先前结果表明PB2+破坏了分泌物 通过与胞吞作用的Ca2+效应器的高亲和力相互作用。 作为 这些研究的扩展，当前项目的目的是测试 假设特定的Ca2+调节的脂质结合蛋白，钙乳蛋白酶 i，突触毒素（p65），蛋白激酶C和细胞质形式的形式 磷脂酶A2（CPLA2）是分泌的重要组成部分 PB2+触发动作的机械和关键目标。 这 蛋白质将被隔离，并且它们与PB2+的相互作用表征， 使用生化，生物物理和免疫化学技术。 透化的牛肾上腺染色体细胞和无细胞模型 将开发胞吐作用以进一步表征PB2+在 胞胞胞菌机制。 除了与胞吐仪的相互作用外，铅离子 通过通过 尚未定义的细胞内机制。 全电池电压钳和 将采用来自培养基中铬蛋白细胞的单信道记录 测试低浓度的细胞内PB2+调节的假设 L型Ca2+电流通过减速钙依赖性通道 失活。 这项研究有望提高我们对PB2+动作的理解 分子水平的分泌，同时提供重要的 有关胞吐过程本身的性质的信息。 在PB2+中 与CA2+信号转导不同水平的细胞的相互作用 组织并可能将CA/脂质结合蛋白识别为主要 铅毒性中的分子靶标。