Defining the roles and regulation of neuronal autophagy

项目总结 自噬是一种溶酶体降解途径，对维持神经元的动态平衡和存活至关重要。 自噬将受损和老化的细胞成分从细胞内环境和靶点隔离开来 将其转化为溶酶体进行破坏。自噬缺陷与神经发育异常和 哺乳动物的神经退化。此外，激活自噬可以挽救神经退行性变和 与年龄相关的小鼠认知能力下降。然而，人们对自噬的调节和功能知之甚少。 提供神经保护的物质。迄今为止，阐明自噬的分子基础的大部分工作都是 在缺乏神经元形态复杂性的模型系统中进行。此外，我们的研究已经 揭示了几种典型的范例，包括饥饿和mTOR抑制，它们在 非神经细胞不会强烈地诱导神经元的自噬。因此，直接研究自噬是至关重要的。 在神经元中提供洞察力，可能会改进治疗，以减轻神经元功能障碍的疾病。因此， 这项提议的目的是定义神经元自噬的作用和调节，以促进 神经功能与生存。我们证实神经元中的自噬是一种高度区隔的 进程。轴突自噬是一种单向途径，允许货物从远侧的 轴突到胞体进行降解。与轴突自噬的远程途径不同，树突状细胞 自噬是由自噬空泡(AVs)的双向运动定义的，这些空泡可能执行更局部性的 功能。我们的初步数据已经确定了神经元生物学的三个方面(突触连接， 神经营养支持，以及与星形胶质细胞的相互作用)，调节特定神经元的自噬 车厢。我们发现，突触活动选择性地控制树突而不是轴突的房室动力学。 我们发现神经营养因子BDNF诱导轴突逆行自噬通量。最后，我们发现联合- 用星形胶质细胞培养神经元可以降低轴突中的自噬小体密度。这些问题的机制基础 然而，通路及其功能尚不清楚。根据我们的初步数据，我们假设 突触活性、神经营养因子和星形胶质细胞不同地调节神经元的自噬，并且， 自噬在神经元功能和存活中发挥特定的作用。为了检验这一假设， 我们将追求三个目标：(1)确定突触活动如何调节神经元自噬以及自噬如何 影响突触功能；(2)确定神经营养因子如何调节神经元自噬以及如何自噬 影响神经营养因子信号；以及(3)阐明星形胶质细胞如何调节神经元自噬。我们会 使用细胞生物学、生物化学和电生理学中的定量方法来获得机械 了解每条途径。这些研究将全面绘制出 神经元中的自噬对来自其复杂环境的不同相互作用做出反应。获得了新的见解 这项研究将为神经系统疾病的治疗干预提供更好的策略。

PROJECT SUMMARY Autophagy is a lysosomal degradation pathway that is critical to maintain neuronal homeostasis and survival. Autophagy sequesters damaged and aged cellular components from the intracellular environment and targets them to lysosomes for destruction. Defective autophagy is linked to neurodevelopmental abnormalities and neurodegeneration in mammals. Further, activating autophagy can rescue models of neurodegeneration and age-related cognitive decline in mice. Little is known, however, about the regulation and functions of autophagy that provide neuroprotection. Much of the work to date elucidating the molecular basis of autophagy has been performed in model systems that lack the morphological complexity of neurons. Further, our research has revealed that several canonical paradigms, including starvation and mTOR-inhibition, that trigger autophagy in non-neuronal cells do not robustly induce autophagy in neurons. Thus, it is critical to study autophagy directly in neurons to provide insight that may improve therapies to mitigate diseases of neuronal dysfunction. Thus, the objective for this proposal is to define the roles and regulation of neuronal autophagy that facilitate neuronal function and survival. We established that autophagy in neurons is a highly compartmentalized process. Axonal autophagy is a unidirectional pathway that allows cargo delivery from distant regions of the axon to the soma for degradation. In contrast to the long-range pathway for autophagy in axons, dendritic autophagy is defined by bidirectional movement of autophagic vacuoles (AVs) that may execute more localized functions. Our preliminary data have identified three aspects of neuronal biology (synaptic connectivity, neurotrophic support, and interactions with astrocytes) that regulate autophagy in specific neuronal compartments. We find that synaptic activity controls AV dynamics selectively in dendrites and not in the axon. We find that the neurotrophin BDNF induces retrograde autophagic flux along axons. Lastly, we find that co- culturing neurons with astrocytes decreases autophagosome density in axons. The mechanistic basis for these pathways and their functions, however, are unknown. Based on our preliminary data, we hypothesize that synaptic activity, neurotrophins, and astrocytes differentially regulate autophagy in neurons, and, that autophagy plays compartment-specific roles in neuronal function and survival. To test this hypothesis, we will pursue three aims: (1) Define how synaptic activity regulates neuronal autophagy and how autophagy affects synapse function; (2) Determine how neurotrophins regulate neuronal autophagy and how autophagy impacts neurotrophin signaling; and (3) Elucidate how neuronal autophagy is regulated by astrocytes. We will use quantitative approaches in cell biology, biochemistry, and electrophysiology to gain a mechanistic understanding for each pathway. These studies will comprehensively map the pathways and functions for autophagy in neurons in response to diverse interactions from their complex environment. New insights gained from this study will better inform strategies for therapeutic intervention to treat disorders of the nervous system.