title: 1-Methyl-4-phenylpyridinium-induced apoptosis in cerebellar granule neurons is mediated by transferrin receptor iron-dependent depletion of tetrahydrobiopterin and neuronal nitric-oxide synthase-derived superoxide.
abstract: In this study, we investigated the molecular mechanisms of toxicity of 1-methyl-4-phenylpyridinium (MPP(+)), an ultimate toxic metabolite of a mitochondrial neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, that causes Parkinson-like symptoms in experimental animals and humans. We used rat cerebellar granule neurons as a model cell system for investigating MPP(+) toxicity. Results show that MPP(+) treatment resulted in the generation of reactive oxygen species from inhibition of complex I of the mitochondrial respiratory chain, and inactivation of aconitase. This, in turn, stimulated transferrin receptor (TfR)-dependent iron signaling via activation of the iron-regulatory protein/iron-responsive element interaction. MPP(+) caused a time-dependent depletion of tetrahydrobiopterin (BH(4)) that was mediated by H(2)O(2) and transferrin iron. Depletion of BH(4) decreased the active, dimeric form of neuronal nitric-oxide synthase (nNOS). MPP(+)-mediated "uncoupling" of nNOS decreased *NO and increased superoxide formation. Pretreatment of cells with sepiapterin to promote BH(4) biosynthesis or cell-permeable iron chelator and TfR antibody to prevent iron-catalyzed BH(4) decomposition inhibited MPP(+) cytotoxicity. Preincubation of cerebellar granule neurons with nNOS inhibitor exacerbated MPP(+)-induced iron uptake, BH(4) depletion, proteasomal inactivation, and apoptosis. We conclude that MPP(+)-dependent aconitase inactivation, Tf-iron uptake, and oxidant generation result in the depletion of intracellular BH(4), leading to the uncoupling of nNOS activity. This further exacerbates reactive oxygen species-mediated oxidative damage and apoptosis. Implications of these results in unraveling the molecular mechanisms of neurodegenerative diseases (Parkinson's and Alzheimer's disease) are discussed.
authors.affiliation: Department of Biophysics and Free Radical Research Center, Medical College of Wisconsin, Milwaukee, 53226, USA.
authors.forename: Tiesong
authors.initials: T
authors.lastname: Shang
authors.forename: Srigiridhar
authors.initials: S
authors.lastname: Kotamraju
authors.forename: Shasi V
authors.initials: SV
authors.lastname: Kalivendi
authors.forename: Cecilia J
authors.initials: CJ
authors.lastname: Hillard
authors.forename: B
authors.initials: B
authors.lastname: Kalyanaraman
mesh_terms: D015655:1-Methyl-4-phenylpyridinium / Q000494:pharmacology / Q000633:toxicity*; D000154:Aconitate Hydratase / Q000037:antagonists & inhibitors; D000818:Animals; D000975:Antioxidants / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects*; D001708:Biopterins / Q000031:analogs & derivatives* / Q000378:metabolism; D002531:Cerebellum / Q000166:cytology; D042967:Electron Transport Complex I / Q000037:antagonists & inhibitors; D006801:Humans; D007395:Interneurons / Q000166:cytology*; D007501:Iron / Q000502:physiology; D024101:Mitochondrial Proteins; D019001:Nitric Oxide Synthase / Q000378:metabolism / Q000502:physiology*; D052248:Nitric Oxide Synthase Type I; D010300:Parkinson Disease; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D011990:Receptors, Transferrin / Q000096:biosynthesis / Q000502:physiology*; D015398:Signal Transduction; D013481:Superoxides / Q000378:metabolism
chemical_list: D000975:Antioxidants; D024101:Mitochondrial Proteins; D017382:Reactive Oxygen Species; D011990:Receptors, Transferrin; D013481:Superoxides; D001708:Biopterins; D007501:Iron; C496320:NOS1 protein, human; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; C496322:Nos1 protein, rat; D000154:Aconitate Hydratase; D042967:Electron Transport Complex I; C003402:sapropterin; D015655:1-Methyl-4-phenylpyridinium