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.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 钙信号和胞吐的调节是所有动物细胞生理学中的中心问题。我们寻求定量的了解，这种信号通过生物物理实验在电兴奋和非兴奋的哺乳动物细胞系：PC 12嗜铬细胞瘤细胞，tsA上皮细胞和胰管上皮细胞。两个长期的假设指导这项工作：（a）Ca 2+的清除和胞吐的调节在不同的细胞中采取不同的形式，并调整到每个细胞的生理作用;（B），几个细胞内的细胞器作出重大贡献的细胞Ca 2+动力学。本研究的主要目的是：（1）验证分泌颗粒对Ca ~（2+）的积累和释放在生理反应中对细胞Ca ~（2+）信号转导的重要作用。(2)测量受体诱发的肌醇1，4，5，三磷酸（IP 3）升高的幅度，并检验通过IP 3的Ca 2+信号传导被IP 3 5-磷酸酶快速代谢IP 3终止，随后Ca 2+快速再摄取到内质网Ca 2+库中的假设。（3）验证细胞骨架的快速重塑和细胞骨架轨迹参与了分泌颗粒从储备池向分泌池的动员。这项工作需要一系列生物物理技术，包括：离子电流的膜片钳;胞吐的电流和电容测量;基因靶向探针，指标和细胞蛋白的转染;比率光度法和荧光共振能量转移（FRET）的指标;视频荧光成像;全内反射显微镜（TIRF）;共聚焦显微镜;和定量动力学建模。 通过虚拟细胞建模将关注Ca 2+的房室动力学和IP 3的产生和分解。