CYSTOLIC-FREE CALCIUM AND CELL MOTILITY

这项工作的目标是了解（1）的机制， 运动细胞内钙的调节，（2） 钙离子变化与细胞内其他 信号机制，以及（3）这些信号的作用 调节细胞运动的机制。 拟议的工作将 检查两种类型的吞噬细胞：人类中性粒细胞和 小鼠腹腔巨噬细胞。 对于这两种细胞， 是信号产生的广泛的生物化学表征 机制和调节细胞骨架的机制。 还有大量关于动物运动行为的信息 这些细胞对各种刺激的反应。 尽管密集 研究中，控制体内运动的机制仍然存在， 虽然（Ca 2+）i、pHi、激酶C和其他 已经提出并研究了信号。 光学显微镜技术将用于测量 细胞内信号的产生和运动反应， 单细胞 (Ca2+）i将以高时间和 空间分辨率 这些测量将同时进行 通过pHi和细胞形态的测量， 信号的产生和运动之间的相关性 细胞 测量单细胞中激酶C活化的方法将 发展。 三个信号（（Ca 2+）i，pHi， 和激酶C）将被检查。 趋化性和吞噬作用 表现出脱敏/再敏化现象。 机制 为此，将根据对 这三种细胞内信号及其相互关系。 最后，这些信号在形态学和 检查细胞骨架的变化。 趋化性和 吞噬作用是需要在单个细胞上研究的过程， 了解生物化学机制是如何协调的， 产生特定的细胞行为。 所描述的显微镜方法 在该提议中，提供了研究细胞内信号手段 和单细胞水平的细胞反应。

The goal of this work is to understand (1) the mechanisms of intracellular calcium regulation in motile cells, (2) the relationship between calcium changes and other intracellular signalling mechanisms, and (3) the role of these signalling mechanisms in regulating cell motility. The proposed work will examine two types of phagocytic cells: the human neutrophil and the mouse peritoneal macrophage. For both types of cells, there is extensive biochemical characterization of the signal generating mechanisms and the mechanisms for regulation of the cytoskeleton. There is also substantial information on the motile behavior of these cells in response to various stimuli. Despite intensive study, the mechanisms for controlling motility in vivo remain unclear, although roles for (Ca2+)i, pHi, kinase C, and other signals have been proposed and investigated. Optical microscopy techniques will be used to measure both the generation of intracellular signals and the motile response in single cells. (Ca2+)i will be measured with high temporal and spatial resolution. These measurements will be made simultaneous with measurements of pHi and cell morphology to provide detailed correlations between the generation of signals and the motion of cells. Methods to measure kinase C activation in single cells will be developed. The interplay between three signals ((Ca2+)i, pHi, and kinase C) will be examined. Both chemotaxis and phagocytosis exhibit desensitization/resensitization phenomena. The mechanism for this will be studied based on analysis of the properties of these three intracellular signals and their inter-relationships. Finally, the role of these signals in the morphological and cytoskeletal changes will be examined. Both chemotaxis and phagocytosis are processes that require study at the single cell level to understand how biochemical mechanisms are orchestrated to produce specific cell behavior. The microscopic methods described in this proposal provide a means to study intracellular signals and cell responses at the single cell level.