The role of Parkin in selective dopamine neuronal degeneration

描述(申请人提供)：黑质纹状体(NS)多巴胺(DA)神经元的进行性变性是帕金森病(PD)运动症状的基础，缺乏减缓进展的治疗方法。DA代谢异常被认为是NSDA神经元选择性变性的机制之一。然而，所有的DA神经元在帕金森病中并没有受到同样程度的影响。虽然中脑NSDA神经元严重丢失，但下丘脑结节漏斗(TI)DA神经元保持完好。在体外和体内，当这些DA神经元群体暴露于线粒体复合体I抑制剂时，也可以看到类似的易感性模式。尽管NSDA和TIDA神经元对复合体I的抑制有相似的初始反应，但只有TIDA神经元能够使用依赖于新蛋白质合成的机制恢复。我们的初步数据还表明，Parkin参与了这种差异易感性，因为它在急性毒性损伤后在TIDA神经元中显著上调，而不是NSDA神经元。Parkin是一种多功能蛋白质，支持正常的线粒体功能，具有E3连接酶，标记错误折叠的蛋白质以进行蛋白酶体降解，并似乎保护DA神经元免受各种毒性侮辱。Parkin还具有重要的蛋白小体非依赖功能，可促进DA神经元的存活。这个项目的总体目标是阐明parkin介导的使NSDA神经元对外源性神经毒素敏感而使Tida神经元对外源性神经毒素耐受的分子机制。我们推测，Parkin主要通过非蛋白酶体机制发挥作用，保护TIDA神经元，并可以保护NSDA神经元免受PD相关的病理性应激、线粒体复合体I抑制所致的损伤。为了解决这一中心假设，将在野生型小鼠和parkin零背景小鼠上进行rAAV介导的、空间受限的parkin表达操作。将评估外源性调节parkin表达对TIDA和NSDA神经元对急性和慢性神经毒性应激反应的影响，以确认parkin在允许DA神经元在初始损伤后恢复的中心作用。分子和药理学方法将被用来阐明parkin神经保护作用的初始下游介体。尽管观察到在临床可观察到的帕金森病症状开始时，相当一部分DA神经元已经丢失，但有可能存在处于危险和功能障碍的神经元，但如果提供必要的恢复机制，这些神经元是可以恢复的。由于Tida神经元在PD相关的病理应激源诱导的损伤中的恢复能力是DA神经元中的独特之处，它们为剖析Parkin介导的DA神经元恢复机制提供了一个强大的平台，并可能为神经保护治疗的发展提供新的靶点。

DESCRIPTION (provided by applicant): Progressive degeneration of nigrostriatal (NS) dopamine (DA) neurons underlies the motor symptoms of Parkinson's disease (PD) and therapies that slow progression are lacking. Abnormal DA metabolism has been proposed as a mechanism for the selective degeneration of NSDA neurons. All DA neurons, however, are not affected to the same extent in PD. While there is severe loss of midbrain NSDA neurons, the hypothalamic tuberoinfundibular (TI) DA neurons remain intact. A similar pattern of susceptibility can be seen in these DA neuronal populations when they are exposed to mitochondrial complex I inhibitors in vitro and in vivo. Although NSDA and TIDA neurons have a similar initial response to complex I inhibition, only TIDA neurons are able to recover using a mechanism that is dependent on new protein synthesis. Our preliminary data also implicate that parkin is involved in this differential susceptibility since it is markedly upregulated in TIDA, but not NSDA neurons following an acute toxic injury. Parkin is a multifunctional protein that supports normal mitochondrial function, has an E3 ligase that tags misfolded proteins for proteosomal degradation and appears to protect DA neurons from a variety of toxic insults. Parkin also has important proteosome independent functions that could promote DA neuronal survival. The overall goal of this project is to elucidate the parkin-mediated molecular mechanisms that render NSDA neurons sensitive and TIDA neurons resistant to exogenous neurotoxin exposure. We hypothesize that parkin, acting primarily via proteosome-independent mechanisms, protects TIDA neurons and can rescue NSDA neurons from injury induced by a PD relevant pathological stress, mitochondrial complex I inhibition. To address the central hypothesis, rAAV-mediated, spatially restricted manipulation of parkin expression will be performed in wild-type mice and on mice with a parkin null background. The impact of exogenously modulating parkin expression on the TIDA and NSDA neuronal responses to both acute and chronic neurotoxic stress will be assessed to confirm the central role of parkin in allowing DA neurons to recover following an initial insult. Molecular and pharmacological methods will be used to elucidate the initial downstream mediators of the neuroprotective effects of parkin. Despite the observation that a significant portion of DA neurons have been lost at the onset of clinically observable symptoms of PD, it is plausible that there are neurons that are at risk and dysfunctional, but that could recover if provided the necessary machinery for recovery. Since TIDA neurons are unique amongst DA neurons in their ability to recover from an injury induced by a PD relevant pathological stressor, they provide a powerful platform to dissect the parkin-mediated mechanisms of DA neuronal recovery and could yield novel targets for neuroprotective therapy development.