We propose to investigate the metabolic fate and neurotoxicological properties of a series of compounds that are structurally related to the Parkinsonian inducing neurotoxin 1-methyl-4-- phenyl-1,2,3,6-tetrahydropyridine (MPTP) and which display neurotoxic effects in the monkey (the phenoxytetrahydropyridine 1 and pyrrole substituted 4-piperidinol 4), in man [the 4-pchlorophenyl substituted 4-piperidinol haloperidol (5)] or in the mouse (the 1-methylpyrrol-2-yl analog 3 of MPTP). Based on the documented mechanism of action of MPTP, these studies will focus on the metabolic formation and CNS access of the corresponding pyridinium species, the potential ultimate neurotoxins. All of the test compounds are partially oxidized piperidine derivatives and it is anticipated that the intermediate iminium ion metabolites will undergo spontaneous oxidation to the corresponding pyridinium species. The neurotoxic properties of the test compounds will be evaluated by classical neurotransmitter depletion and histopathology studies in the mouse. In addition to standard staining techniques, we will use immunocytochemical stains to identify specific lesions that may involve a particular neurotransmitter system. These studies will not be limited to the dopaminergic system since pyridinium derivatives appear to be generally neurotoxic with specificity being dependent on transporter based localization. The toxicity of the anticipated pyridinium metabolites will be assessed following direct intracerebral injection or intracerebral perfusion by microdialysis. Studies on the ability of these pyridinium compounds to inhibit mitochondrial respiration, an apparent obligatory step in the mediation of the neurotoxicity of MPTP by its pyridinium metabolite, will be investigated in cell culture with the aid of a supravital dye based assay which employs a spectrofluorometric detector that can estimate mitochondrial function in a single cell or group of cells over time. This multidisciplinary program will benefit from the input of several consultants whose principal research activities rely on the proposed techniques. Preliminary results obtained with haloperidol, its tetrahydropyridine derivative and the corresponding pyridinium derivative support the principal thesis of this proposal, namely that partially oxidized 1,4-disubstituted piperidine derivatives will be biotransformed to neurotoxic pyridinium metabolites that have access to the central nervous system.
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