The goal of this research proposal is to elucidate the roles of mitochondrial dysfunction and oxidative stress in a cell model of Parkinson's Disease. Evidence shows that these factors may initiate or at least occur during the process of degeneration characteristic of Parkinson's Disease and MPTP toxicity. A neuroblastoma cell line expressing the dopamine transporter protein (DAT) provides a unique model to study the toxic effects of MPP+, the toxic metabolite of MPTP. Since MPP+ is taken up selectively by cells that express DAT, the effects of MPP+ on this cell line can be investigated to determine the mechanisms involved in producing the damage. Measuring changes in glycolytic metabolism using the Cytosensor microphysiometer, and assaying altered mitochondrial function and oxidative stress utilizing fluorescent indicators in conjunction with a fluorescence plate system, imager and confocal microscopy will allow determination of the sequence of events that lead to MPP+-induced damage. Therapeutics targeted at interrupting this sequence will be utilized to prevent the subsequent damage. In addition, the role of glia in MPP+toxicity will be examined. Based on other lines of evidence that glia may provide protection from such damaging insults, the possibility that glial-mediated circumvention of oxidative stress prevents MPP+-induced damage will be investigated.
