INTRACELLULAR DYNAMICS OF CALCIUM SIGNALS AND EXOCYTOSIS

钙信号和胞吐作用的细胞内动力学

• 批准号: 7956413
• 负责人: BERTIL HILLE
• 金额: $ 1.63万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2010-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7956413
• 关键词: Animals; Calcium Signaling; Cell Line; Cells; Computer Retrieval of Information on Scientific Projects Database; Confocal Microscopy; Electric Capacitance; Endoplasmic Reticulum; Epithelial Cells; Exocytosis; Fluorescence Resonance Energy Transfer; Funding; Grant; Inositol; Institution; Ions; Kinetics; Mammalian Cell; Measurement; Measures; Metabolism; Microscopy; Modeling; Organelles; Pancreatic duct; Pheochromocytoma; Photometry; Physiological; Physiology; Production; Proteins; Regulation of Exocytosis; Research; Research Personnel; Resources; Role; Secretory Vesicles; Signal Transduction; Source; Techniques; Testing; Transfection; United States National Institutes of Health; Work; fluorescence imaging; inositol-1,4,5-trisphosphate 5-phosphatase; patch clamp; ratiometric; receptor; research study; response; reuptake; virtual

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Calcium signaling and the regulation of exocytosis are central issues in the physiology of all animal cells. We seek quantitative understanding of such signaling through biophysical experiments in electrically excitable and non-excitable mammalian cell lines: PC12 pheochromocytoma cells, tsA epithelial cells, and pancreatic duct epithelial cells. Two long-term hypotheses guide this work: (a) that Ca2+ clearance and the regulation of exocytosis take different forms in different cells and are tuned to the physiological role of each cell; and (b) that several intracellular organelles make significant contributions to cellular Ca2+ dynamics. The aims in this grant period are: (1) To test the hypothesis that accumulation and release of Ca2+ by secretory granules can make significant contributions to cellular Ca2+ signaling during physiological responses. (2) To measure the amplitude of receptor-evoked inositol 1,4,5, trisphosphate (IP3) elevations and to test the hypothesis that Ca2+ signaling via IP3 is terminated by rapid metabolism of IP3 by IP3 5- phosphatase followed by rapid reuptake of Ca2+ into the endoplasmic reticulum Ca2+ stores. And (3) To test the hypothesis that cytoskeletal tracks and fast cytoskeletal remodeling participate in the mobilization of secretory granules from reserve pools into secretion-competent pools. The work requires a range of biophysical techniques including: patch clamp of ion currents; amperometric and capacitance measurements of exocytosis; transfection of genetically targeted probes, indicators, and cellular proteins; ratiometric photometry and fluorescence resonance energy transfer (FRET) of indicators; video fluorescence imaging; total internal reflection microscopy (TIRF); confocal microscopy; and quantitative kinetic modeling. Modeling by Virtual Cell will concern the compartmental dynamics of Ca2+ and production and breakdown of IP3.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 钙信号传导和胞吐作用的调节是所有动物细胞生理学中的核心问题。我们通过在电动和不可激发的哺乳动物细胞系中的生物物理实验来寻求对这种信号传导的定量理解：PC12嗜铬细胞瘤细胞，TSA上皮细胞和胰管上皮细胞。两个长期假设指导这项工作：（a）Ca2+清除率和胞吞作用的调节在不同的细胞中采用不同的形式，并调整为每个细胞的生理作用； （b）几个细胞内细胞器对细胞Ca2+动力学有重要贡献。在此赠款期间的目的是：（1）检验以下假设：分泌颗粒可以在生理反应过程中对细胞Ca2+信号传导做出重大贡献。 （2）测量受体诱发的肌醇1,4,5的振幅，三磷酸（IP3）升高，并测试以下假设：通过IP3快速代谢IP3的Ca2+信号通过IP3 5-磷酸酶对IP3的快速代谢终止。 （3）检验了一个假说，即细胞骨架轨迹和快速细胞骨架重塑参与了从储备池中分泌颗粒的动员，以分泌能力的池。这项工作需要一系列生物物理技术，包括：离子电流的斑块夹；胞吐作用的安培和电容测量；转染遗传靶向探针，指标和细胞蛋白；指标的比率光度计和荧光共振能量转移（FRET）；视频荧光成像；总内反射显微镜（TIRF）；共聚焦显微镜；和定量动力学建模。 通过虚拟单元进行建模将涉及Ca2+的隔室动态以及IP3的生产和分解。