Role of presynaptic calcium stores

描述（由申请人提供）：细胞内Ca 2+信号是通过神经元回路进行的信息处理的组成部分，所述神经元回路促进突触前和突触后功能，包括突触传递的长时程增强（LTP）和抑制（LTD），长时程突触可塑性的电生理学相关性。由于Ca 2+的微调变化调节各种细胞内功能，信号特异性需要对胞质Ca 2+信号进行严格的空间和时间控制。这种需要对于快速神经递质释放是最明显的，其中电压依赖性Ca 2+信号传导在亚毫秒尺度上触发突触前末梢处的突触囊泡的融合。内质网（ER）可以作为一个Ca 2+库/汇或作为一个Ca 2+源，并可能在突触前末梢平衡这些双重角色，以满足突触释放递质的特定需求。然而，ER对突触前Ca 2+信号传导和神经递质释放的贡献还不清楚。我们假设，ER作为初级突触前钙池在苍蝇NMJ。如果是这样的话，突触前内质网可能不仅暂时缓冲胞浆Ca 2+的SERCA泵，但也永久删除从突触末梢的“隧道”腔Ca 2+进入轴突。终末内有限的ER Ca 2+释放可能调节神经递质的释放，但主要用于线粒体Ca 2+摄取激活线粒体能量产生。我们将测试这一假设，利用遗传操纵的果蝇和图像去极化引起的突触前钙水平的ER腔，线粒体和幼虫NMJ的胞质溶胶的变化。在这个建议的中心是转基因Ca 2+指标的发展，可以忠实地报告Ca 2+的变化，在内腔的ER和线粒体，我们预计也将广泛用于遗传分析的线粒体和/或ER Ca 2+的动态在任何细胞的果蝇。具体而言，我们将确定ER介导的Ca 2+摄取，Ca 2+释放和/或Ca 2+扩散（隧道）后，反复神经刺激的突触前终端的幼虫果蝇NMJ（目标1）的动态和确定的作用，ER-线粒体相互作用突触前Ca 2+稳态在幼虫果蝇NMJ（目标2）。从这个系统的分析，一个基本的框架将出现更好地理解的作用，ER在突触前的Ca 2+信号/稳态突触前末梢扩大我们的理解突触传递的重要调控机制和它们的关系，神经系统的功能可塑性和人类健康。

DESCRIPTION (provided by applicant): Intracellular Ca2+ signals are an integral part of information processing by neuronal circuits facilitating pre- and postsynaptic functions including long-term potentiation (LTP) and depression (LTD) of synaptic transmission, the electrophysiological correlates of long-term synaptic plasticity. Since finely tuned changes in Ca2+ modulate a variety of intracellular functions, signal specificity requires a tight spatial and temporal control of the cytosolic Ca2+ signal. This need is most apparent for fast neurotransmitter release where voltage-dependent Ca2+ signaling triggers the fusion of synaptic vesicles at presynaptic terminals on a sub-millisecond scale. The endoplasmic reticulum (ER) can act as a Ca2+ store/sink or as a Ca2+ source and is likely to balance these dual roles at presynaptic terminals to meet the specific needs of the synapse for transmitter release. However, the contributions of ER to presynaptic Ca2+ signaling and neurotransmitter release are not well understood. We hypothesize that the ER acts as the primary presynaptic Ca2+ sink at fly NMJs. If so, the presynaptic ER may not only temporarily buffer cytosolic Ca2+ by SERCA pumping but also permanently remove Ca from synaptic terminals by "tunneling" luminal Ca2+ into axons. Limited ER Ca2+ release within terminals may modulate neurotransmitter release but mainly serve mitochondrial Ca2+ uptake activating mitochondrial energy production. We will test this hypothesis by exploiting genetically manipulated Drosophila and image depolarization-induced changes in presynaptic Ca levels of the ER lumen, mitochondria and the cytosol of larval NMJs. At the center of this proposal is the development of transgenic Ca2+ indicators that can faithfully report Ca2+ changes in the lumen of the ER and mitochondria, which we expect will also be of wide use for the genetic analysis of mitochondrial and/or ER Ca2+ dynamics in any cell of Drosophila. Specifically, we will determine the dynamics of ER-mediated Ca2+ uptake, Ca2+ release and/or Ca2+ diffusion (tunneling) upon repetitive nerve stimulation at presynaptic terminals of larval Drosophila NMJs (Aim 1) and determine the role of ER-mitochondria interactions for presynaptic Ca2+ homeostasis at larval Drosophila NMJs (Aim 2). From this systematic analysis a basic framework will emerge for better understanding the role of the ER in presynaptic Ca2+ signaling/homeostasis at presynaptic terminals expanding our understanding of important regulatory mechanisms of synaptic transmission and their relation to the functional plasticity of the nervous system and human health.