MECHANISMS OF NEURONAL DEATH IN PARKINSON'S DISEASE

描述（由申请人提供）：轴突运输受损可能在包括帕金森病（PD）在内的多种神经退行性疾病中发挥早期关键作用。当轴突运输中断时，营养支持丧失，突触囊泡和递质耗尽，生物物质积累，发生变性，最终神经元死亡。作为一种病理前兆，受损的轴突运输在PD中特别引人注目，因为突触功能的破坏和轴突病理的存在是早期的共同特征。然而，由于缺乏选择性成像和靶向CNS轴突的工具，这一领域的研究受到限制。为此，我们已经修改了一个分隔系统设计的外周文化，使中枢神经系统神经元可以生长与轴突隔离到一侧的障碍。当与源自基因工程小鼠的GFP标记的多巴胺能神经元一起使用时，可以使用活细胞、真实的时间成像来检查多巴胺能轴突。本申请的目标是利用我们监测CNS多巴胺能神经元中轴突运输的独特能力来测试一组嵌套假设，即PD相关神经毒素和基因突变触发轴突运输的早期变化，这些变化导致突触功能丧失和细胞死亡。具体而言，我们假设PD模拟MPP+影响线粒体和囊泡轴突运输，导致突触功能丧失，这是通过神经元依赖性和独立的过程发生的，并且PD连锁基因LRRK2的突变将影响与MPP+相似的细胞器和囊泡的运输过程。光学，分子和细胞技术将被用来确定轴突运输和信号通路与多巴胺能轴突的线粒体和囊泡运动。这些实验将提供不可能从标准培养或动物模型中获得的见解。两者合计，拟议的研究将确定环境或遗传诱导的轴突损伤是否在多巴胺能神经元的死亡中起核心作用。如果是这样的话，新的治疗方法可以靶向阐明的控制点，以阻止或减缓疾病进展。与帕金森氏病相关的环境或遗传因素可能会影响神经元将生物物质沿轴突传递的基本机制。这可能会损害神经元之间相互交流的能力，导致蛋白质和细胞器的积累，最终导致轴突和神经元退化。识别和表征帕金森病中轴突损伤如何发生，对于发现新的疗法具有巨大的潜力，这些疗法可以靶向阐明的控制点，以阻止或减缓疾病进展。

DESCRIPTION (provided by applicant): Impaired axonal transport may play an early, pivotal role in a variety of neurodegenerative disorders including Parkinson's disease (PD). When axon transport is disrupted, trophic support is lost, synaptic vesicles and transmitters are depleted, biological materials accumulate, degeneration occurs and ultimately the neuron dies. As a pathological precursor, impaired axonal transport is particularly compelling in PD because disruption of synaptic function and the presence of axon pathology is an early, common feature. Research in this area is limited, however, by the lack of tools to selectively image and target CNS axons. Towards this end, we have modified a compartmented system designed for peripheral cultures such that CNS neurons can be grown with axons segregated to one side of a barrier. When used with GFP-labeled dopaminergic neurons derived from genetically engineered mice, dopaminergic axons can be examined using live cell, real time imaging. The goal of the current application is to utilize our unique ability to monitor axonal transport in CNS dopaminergic neurons to test a nested set of hypotheses that PD-associated neurotoxins and genetic mutations trigger early changes in axon transport that contribute to the loss of synaptic function and cell death. Specifically, we hypothesize that the PD-mimetic MPP+ affects mitochondrial and vesicular axon trafficking leading to the loss of synaptic function, that this occurs via mitochondrial-dependent and independent processes, and that mutations in the PD-linked gene LRRK2 will affect similar transport processes of organelles and vesicles as MPP+. Optical, molecular and cellular techniques will be used to determine axonal transport and signaling pathways associated with mitochondrial and vesicular movement in dopaminergic axons. These experiments will provide insights impossible to obtain from standard culture or animal models. Taken together, the proposed studies will determine whether environmentally or genetically induced axonal injury plays a central role in the death of dopaminergic neurons. If so, novel therapies can be targeted to the elucidated control points in order to stop or slow disease progression. PUBLIC HEALTH RELEVANCE Environmental or genetic factors associated with Parkinson's disease may affect fundamental mechanisms underlying the way in which a neuron sends biological materials down an axon. This may compromise the ability of neurons to communicate with each other, lead to the accumulation of proteins and organelles, and ultimately cause the axon and subsequently the neuron to degenerate. Identifying and characterizing how axonal injury occurs in Parkinson's disease has enormous potential for the discovery of novel therapies that can be targeted to the elucidated control points in order to stop or slow disease progression.