Trans-synaptic control of presynaptic neurotransmitter release

摘要： 许多神经精神和神经系统疾病造成的严重健康负担可能是 与指导发育的分子途径的限制或破坏有关， 突触连接的维持或可塑性。重要的是，许多神经回路 持续活跃的单个突触连接必须经历连续的， 协调的稳态变化，以抵消持续的突触增强/减弱 以及网络不稳定性的发展。因此，绘制活动地点的功能图- 分子和信号通路的依赖动态协调， 这一过程对于理解许多CNS疾病至关重要。拟议的工作是 它具有独特的重要性，因为它将阐明新的，但有效的分子信号传导途径， 介导突触功效中准确且可再现的活性依赖性适应。 具体地说，研究集中在了解如何通过突触后的活动感知， mTORC 1/BDNF信号通路介导突触前神经递质释放增加 在减少对突触后元件的兴奋性输入期间。调查将测试 tomosyn-1是mTORC 1/BDNF/TrkB受体中枢突触前靶点假说 信号和突触前神经递质释放的跨突触调节， 信号通路通过UPS调节Tomosyn-1蛋白水平而发生。我们还将定义 分泌途径中的特定位点，tomosyn通过该位点控制突触前 神经递质释放这种跨突触机制的共同性也将涉及 海马苔藓纤维/CA 3和CA 3/CA 1突触的综合评价 切片，因为这些电路被称为表现出惊人的分歧突触可塑性。此外，本发明还提供了一种方法， 我们将表征负责tomosyn的UPS调节的E3连接酶，确定 E3连接酶活性对mTORC 1/BDNF/TrkB信号传导敏感，如果tomosyn的泛素化， 通过E3连接酶调节tomosyn蛋白水平和神经递质释放。的 研究采用囊泡循环的最新光学成像，遗传模型靶向 蛋白质/信号传导功能、神经元兴奋性的光遗传学控制和突触的分析 海马神经元和海马切片培养物中的功能。生化分析将 通过UPS定量并建立对tomosyn蛋白的活性依赖性控制。这 信息将大大促进对活动机制的理解， 突触后mTORC 1活性的依赖性变化协调空间和时间 调节突触前神经递质的释放

Abstract: A severe health burden imposed by many neuropsychiatric and neurological diseases can be linked to limitations in, or disruption of, molecular pathways which guide development, maintenance or plasticity of synaptic connections. Importantly, as many neural circuits are persistently active the individual synaptic connections must undergo continuous and coordinated homeostatic changes to counteract continual synaptic strengthening/weakening and development of network instability. Thus, developing a functional map of sites of activity- dependent dynamic coordination of the molecules and signaling pathways that define the process is central to enabling an understanding of many CNS diseases. The work proposed is uniquely important as it will elucidate novel and yet potent molecular signaling pathways that mediate accurate and reproducible activity-dependent adaptations in synaptic efficacy. Specifically, investigations focus on understanding how post-synaptic sensing of activity via the mTORC1/BDNF signaling pathway mediates increased presynaptic neurotransmitter release during reduced excitatory input to the post-synaptic element. Investigations will test the hypothesis that tomosyn-1 is a central presynaptic target of mTORC1/BDNF/TrkB receptor signaling and that trans-synaptic adjustments in presynaptic neurotransmitter release via this signaling pathway occur via UPS regulation of tomosyn-1 protein levels. We will also define specific sites within the secretory pathway by which tomosyn exerts control over presynaptic neurotransmitter release. Commonality of this trans-synaptic mechanism will also involve comprehensive evaluation at Mossy fiber/CA3 and at CA3/CA1 synapses of hippocampal brain slices, as these circuits are known to exhibit strikingly divergent synaptic plasticity. In addition, we will characterize the E3 ligase responsible for UPS regulation of tomosyn, determine if the E3 ligase activity is sensitive to mTORC1/BDNF/TrkB signaling and if ubiqutination of tomosyn by the E3 ligase regulates tomosyn protein levels and neurotransmitter release. The investigations employ state of the art optical imaging of vesicle cycling, genetic models targeting protein/signaling function, optogenetic control of neuronal excitability and analysis of synaptic function in cultures of hippocampal neurons and hippocampal slices. Biochemical assays will quantify and establish activity dependent control on tomosyn protein via the UPS. This information will significantly advance understanding on mechanisms by which activity- dependent changes in post-synaptic mTORC1 activity coordinate spatial and temporal adjustments in presynaptic neurotransmitter release.