The neurorestorative and neuroprotective trophic actions of GDNF on midbrain dopamine (DA) neurons provide a promising but controversial therapeutic approach for the treatment of Parkinson's disease. We specifically plan to address the following major issues in the next 5 years of support for the University of Kentucky Udall Center: 1) We hypothesize that GDNF is affecting DA neurons through at least 3 major mechanisms: A) Upregulation of existing DA neurons - direct effects on tyrosine hydroxylase and related proteins (Project 1);B) Repair of DA neurons and connections - improved dendritic and neuronal fiber connections (Projects 2 and 3);and C) Neurogenesis and/or gliogenesis (Project 3). We will investigate these three mechanisms in the proposed experiments in nonhuman primate models of PD. 2) We propose to use a novel indwelling pump that can deliver GDNF chronically in the freely-moving MPTP-lesioned monkey to evaluate multisite delivery of GDNF (putamen vs. putamen + substantia nigra) to improve the functional restoration of DA systems. 3) Recent studies in PD patients involving a Phase 2 clinical trial have yielded controversial data regarding potential GDNF antibody formation and preliminary toxicology studies in rhesus monkeys support a potential toxicity of GDNF to cerebellar Purkinje cells and granule cells, when chronically administered in high doses (100 Lig/day) and subsequently withdrawn. In conjunction with Projects 2 and 3, we will investigate the potential toxic effects of GDNF on dopamine and non-dopamine containing cells in the CNS. In addition, we will investigate potential antibody formation from chronic GDNF administration. 4) Finally, we will use an improved delivery approach, to investigate the effects of GDNF infusion on milder parkinsonism in unilateral MPTP-treated monkeys, to investigate if earlier intervention of GDNF treatment may contribute to better functional recovery of the nigrostriatal system. We will utilize a recently developed stable, unilateral milder parkinsonian monkey model for these studies, which may be an """"""""earlystage"""""""" model of PD. The different highly integrated Projects and Cores will provide key data regarding the functional effects of chronic GDNF treatments to DA neurons, mechanisms of action of GDNF both positive and negative, and will provide data that may be useful to gain a better understanding of the effects of trophic factors on dead or dying neurons. Project 1 Title: Neuropharmacological Studies PI: Greg Gerhardt, PhD DESCRIPTION (provided by applicant): Quantitative Neuropharmacological .Studies of the Effects of Chronic Delivery of GDNF in Rhesus Monkeys. The neurorestorative and neuroprotective trophic actions of GDNF on midbrain DA neurons provide a promising but controversial therapeutic approach for the treatment of Parkinson's disease (PD). Based on past work, we hypothesize that GDNF is affecting DA neurons through at least 3 major mechanisms: 1) Upregulation of existing DA neurons - direct effects on tyrosine hydroxylase and related proteins;2) Repair of DA neurons - improved dendritic and neuronal fiber connections;and 3) Neurogenesis and/or gliogenesis. Project 1 will investigate DA function and repair of DA neurons, in conjunction with Projects 2 and 3. It is our central hypothesis that dual-site chronic intraputamenal/intranigral delivery of GDNF will improve function and restoration of damaged DA neurons with greater efficacy, potency and reduced side effects. In addition, we predict, based on preliminary data, that GDNF treatment in milder parkinsonian animals, a model of early stage PD patients, will lead to optimal restoration of the nigrostriatal DA neuronal system. We propose to use an indwelling pump that can deliver GDNF chronically in the freely-moving unilateral 1-methyl-4-phenyl- 1,2,3,6-tetrahydropyridine (MPTP)-lesioned middle-aged female monkeys. Project 1 of this revised program project grant will perform quantitative neurochemical assessments using in vivo microdialysis, Western immunoblot assays, immunohistochemical studies and high performance liquid chromatography coupled with electrochemical detection (HPLC-EC) to study DA neuronal systems in the striatum (putamen and caudate nucleus) and substantia nigra of Rhesus monkeys that have received chronic GDNF infusions. These neurochemical changes to the monkey striatum during chronic delivery of GDNF will be performed in conjunction with behavioral, immunohistochemical, histological and functional MRI (fMRI) studies of the same monkeys in conjunction with Projects 2 and 3. In addition, Project 1 will begin studies of glutamate regulation in the striatum, substantia nigra and globus pallidus of 6-OHDA lesioned rats and the effects of GDNF infusion on glutamate regulation in the basal ganglia. This is as a new area of scientific growth of the Center. The investigation of the role of glutamate in relation of DA function will improve our understanding of the essential role of this neurotransmitter to basal ganglia function in PD.
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