Regulation and Function of Spontaneous Mini Release at Synapses

描述(申请人提供)：为了了解突触信号如何调节神经元连通性和促进神经系统疾病的长期目标，我们建议使用果蝇作为一个模型系统来确定自发神经递质释放的分子机制以及它如何调节突触连通性。突触神经元通信的特征主要集中在动作电位触发的突触小泡融合上，自发的微小电位被认为主要代表背景噪声。然而，我们最近发现，突触的自发释放独立于由复杂蛋白(一种突触特异性的SNARE复合体结合蛋白)诱导的释放，并可能调节突触的生长。果蝇复合体蛋白的突变导致突触自发融合的显著增加，这种融合不依赖于细胞外钙或动作电位，并导致突触突触的严重过度生长。络合蛋白也是PKA依赖的磷酸化的靶点，这表明微小频率可以以PKA依赖的方式在突触上独立调节。复合蛋白融合钳对自发融合的调节为神经元突触的信息传递提供了一条不依赖于诱发突触电位的新途径。此外，从精神分裂症到帕金森氏症和亨廷顿病等一系列神经系统疾病都与人类复合蛋白水平的变化有关，这表明复合蛋白功能障碍和自发释放的异常速度可能与人类的几种神经病理有关。在这里，我们建议利用果蝇可用的遗传工具来确定复合素是如何调节自发释放的，以及它的功能是否可以受到神经元活动和PKA途径的调节。这些实验的完成将使我们能够提出未来的研究，以确定Mini的生物学作用及其在大脑中的调节模式，有可能为我们理解由Mini介导的突触和神经元通讯的生物学提供根本性的进步-这种类型的高风险/高回报实验是R21水平支持的理想类型。与公共健康相关：突触交流的特征主要集中在动作电位触发的突触小泡融合上，自发微型电位(MIIS)被认为主要代表背景噪声。我们最近的工作表明，Mini可以在突触中发挥信号作用，并受神经元特异性蛋白复合蛋白的调节。我们将探索复杂蛋白陷阱机制如何调节突触的自发释放，并有助于突触的生长和可塑性。确定Mini在突触生物学中的作用将是我们对大脑功能理解的重要进展，并为进一步理解复杂蛋白失调如何导致神经系统疾病奠定基础。

DESCRIPTION (provided by applicant): With the long-term goal of understanding how synaptic signaling regulates neuronal connectivity and contributes to neurological disease, we propose to use Drosophila as a model system for determining the molecular mechanisms by which spontaneous neurotransmitter release is regulated and how it may regulate synaptic connectivity. Characterization of neuronal communication at synapses has largely focused on action potential-triggered synaptic vesicle fusion, with spontaneous miniature potentials (minis) largely thought to represent background noise. However, we have recently discovered that spontaneous release at synapses is regulated independently of evoked release by complexin, a synapse-specific SNARE complex binding protein, and may modulate synaptic growth. Mutations in Drosophila complexin lead to a dramatic increase in spontaneous fusion at synapses that is independent of extracellular calcium or action potentials, and cause a profound overgrowth of synaptic boutons. Complexin is also a target for PKA-dependent phosphorylation, suggesting that mini frequency can be independently regulated at synapses in a PKA-dependent manner. Regulation of spontaneous fusion by a complexin fusion clamp provides a new avenue for information transfer at neuronal synapses independent of evoked synaptic potentials. In addition, alterations in complexin levels in humans have been implicated in a host of neurological diseases ranging from schizophrenia to Parkinson's and Huntington's Disease, suggesting complexin dysfunction and abnormal rates of spontaneous release may contribute to several human neuropathologies. Here, we propose to take advantage of the genetic tools available in Drosophila to determine how complexin modulates spontaneous release and if its function can be regulated by neuronal activity and the PKA pathway. The completion of these experiments will allow us to propose future studies to define the biological role of minis and their mode of regulation in the brain, potentially providing a fundamental advance in our understanding of the biology of the synapse and neuronal communication mediated by minis - the type of high risk/high reward experiments ideal for R21-level support. PUBLIC HEALTH RELEVANCE: Characterization of synaptic communication has largely focused on action potential-triggered synaptic vesicle fusion, with spontaneous miniature potentials (minis) largely thought to represent background noise. Our recent work suggests that minis can function in a signaling role at the synapse, and are regulated by the neuron specific protein complexin. We will explore how the complexin-SNARE machinery regulates spontaneous release at synapses and contributes to synaptic growth and plasticity. Defining the role of minis in the biology of the synapse would be an important advance in our understanding of brain function and set the stage for further understanding of how complexin dysregulation contributes to neurological disease.