Local Ca2+ signaling in sympathetic ganglion neurons

交感神经节神经元中的局部 Ca2 信号传导

• 批准号: 6896137
• 负责人: MARTIN F SCHNEIDER
• 金额: $ 31.41万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-03 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6896137
• 关键词: Rana; action potentials; calcium flux; confocal scanning microscopy; electrophysiology; endoplasmic reticulum; histochemistry /cytochemistry; imaging /visualization /scanning; intracellular; mitochondria; neurons; perfusion; sympathetic ganglion; tissue /cell culture

DESCRIPTION (provided by applicant): The overall goal of this project is to gain further insight into local Ca2+ signaling in neurons. Local spatio-temporal differences in cytosolic (Ca2+), together with localized intracellular Ca2+ receptor molecules, provide one mechanism for selective modulation of diverse cellular functions by the single messenger Ca2+ in the same cell. Our recent confocal imaging of ryanodine receptor (RyR) Ca2+ release channels, endoplasmic reticulum (ER) Ca2+ pumps and mitochondria in cultured dissociated frog sympathetic ganglion neurons has indicated six specialized cellular sub-domains in these neurons: a peripheral ER-rich shell, an underlying mitochondria-rich shell, a central cytoplasm, a peripheral and a central perinuclear ER and the nucleus at the nuclear pole. Our recent video rate confocal fluorescent Ca2+ indicator (fluo-4) imaging in these neurons has revealed discrete sub-plasma membrane sites of preferential initiation of Ca2+ release activated either by caffeine or by a single action potential. The peripheral Ca2+ transients presumably occur via Ca2+ release from peripheral ER since they are eliminated by ER Ca2+ depletion. We now propose to address the following aims. (1) To characterize Ca2+ release, Ca2+ uptake and Ca2+ propagation in and between each of the identified sub-domains in these neurons, and to simulate our observations with a 6 interconnected sub-domain model of the neurons. (2) To study the basis for local peripheral sites of preferential Ca2+ release during single action potentials, and to determine whether these preferential release sites may also generate discrete local Ca2+ release events (Ca2+) "sparks"). (3) To determine the effects of transmitter-induced activation and other physiological perturbations in neurons in culture as well as in ganglia. We will combine high speed "band scan" (4 or 8 ms per band) and line scan (63 us per line) confocal fluorescent Ca2+ imaging of neurons in culture and in partially dissected fresh ganglia, electrophysiology, rapid extracellular perfusion, release of caged compounds, cytosolic injection of cDNA and/or proteins and histochemical study of live and fixed neurons. Our results will provide new insights regarding local Ca2+ signaling mechanisms that could be compromised in neuronal disease.

描述（由申请人提供）：该项目的总体目标是 进一步了解神经元中局部CA2+信号传导。当地的 胞质（Ca2+）的时空差异以及本地化 细胞内Ca2+受体分子，为选择性提供了一种机制 通过单个Messenger Ca2+调制各种细胞函数 相同的单元。我们最近对Ryanodine受体（RYR）CA2+释放的共聚焦成像 培养的通道，内质网（ER）Ca2+泵和线粒体 解散的青蛙交感神经神经元已表明六个专业 这些神经元中的细胞子域：外围ER富含壳，一个 富含线粒体的壳，中央细胞质，外围和一个 中央核能ER和核极的核。我们最近的视频 这些神经元中的速率共聚焦荧光Ca2+指示剂（Fluo-4）成像具有 揭示了Ca2+优先启动的离散亚质子膜位点 释放由咖啡因或单个动作电位激活。这 外围Ca2+瞬态大概是通过Ca2+从外围ER释放而发生的 由于它们被ER Ca2+耗尽消除。我们现在建议解决 以下目标。 （1）表征Ca2+释放，Ca2+摄取和Ca2+ 在这些神经元中每个鉴定的子域之间的传播， 并用6个互连的子域模型模拟我们的观察 神经元。 （2）研究优先的局部外围部位的基础 CA2+在单个动作电位期间释放，并确定是否 优先发布站点也可能会生成离散的本地CA2+发布事件 （Ca2+）“ Sparks”）。 （3）确定发射器诱导的影响 培养神经元中神经元的激活和其他生理扰动 就像在神经节中一样。我们将结合高速“频段扫描”（每带4或8毫秒）和 线扫描（每行63 US）共焦荧光Ca2+神经元的成像 培养和部分解剖的新鲜神经理，电生理学，快速 细胞外灌注，笼子的释放，胞质注射 cDNA和/或蛋白质以及活神经元和固定神经元的组织化学研究。我们的 结果将提供有关本地CA2+信号传导机制的新见解 这可能在神经元疾病中受到损害。