title: Nitric oxide triggers the toxicity due to glutathione depletion in midbrain cultures through 12-lipoxygenase.
abstract: Glutathione (GSH) depletion is the earliest biochemical alteration shown to date in brains of Parkinson's disease patients. However, data from animal models show that GSH depletion by itself is not sufficient to induce nigral degeneration. We have previously shown that non-toxic inhibition of GSH synthesis with l-buthionine-(S,R)-sulfoximine in primary midbrain cultures transforms a nitric oxide (NO) neurotrophic effect, selective for dopamine neurons, into a toxic effect with participation of guanylate cyclase (GC) and cGMP-dependent protein kinase (PKG) (Canals, S., Casarejos, M. J., de Bernardo, S., Rodríguez-Martín, E., and Mena, M. A. (2001) J. Neurochem. 79, 1183-1195). Here we demonstrate that arachidonic acid (AA) metabolism through the 12-lipoxygenase (12-LOX) pathway is also central for this GSH-NO interaction. LOX inhibitors (nordihydroguaiaretic acid and baicalein), but not cyclooxygenase (indomethacin) or epoxygenase (clotrimazole) ones, prevent cell death in the culture, even when added 10 h after NO treatment. Furthermore, the addition of AA to GSH-depleted cultures precipitates a cell death process that is indistinguishable from that initiated by NO in its morphology, time course, and 12-LOX, GC, and PKG dependence. The first AA metabolite through the 12-LOX enzyme, 12-hydroperoxyeicosatetraenoic acid, induces cell death in the culture, and its toxicity is greatly enhanced by GSH depletion. In addition we show that if GSH synthesis inhibition persists for up to 4 days without any additional treatment, it will induce a cell death process that also depends on 12-LOX, GC, and PKG activation. In this study, therefore, we show that the signaling pathway AA/12-LOX/12-HPETE/GC/PKG may be important in several pathologies in which GSH decrease has been documented, such as Parkinson's disease. The potentiating effect of NO over such a signaling pathway may be of relevance as part of the cascade of events leading to and sustaining nerve cell death.
authors.affiliation: Departamento de Investigación, Servicio de Neurobiología, Hospital Ramón y Cajal, Ctra. de Colmenar, Km. 9, 28034 Madrid, Spain.
authors.forename: Santiago
authors.initials: S
authors.lastname: Canals
authors.forename: Maria José
authors.initials: MJ
authors.lastname: Casarejos
authors.forename: Sonsoles
authors.initials: S
authors.lastname: de Bernardo
authors.forename: Eulalia
authors.initials: E
authors.lastname: Rodríguez-Martín
authors.forename: Maria Angeles
authors.initials: MA
authors.lastname: Mena
mesh_terms: D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid / Q000494:pharmacology; D000818:Animals; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology; D001092:Arachidonate 12-Lipoxygenase / Q000378:metabolism*; D015153:Blotting, Western; D001921:Brain / Q000196:embryology; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured; D003022:Clotrimazole / Q000494:pharmacology; D003470:Culture Media; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D016861:Cyclooxygenase Inhibitors / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D044950:Flavanones; D005419:Flavonoids / Q000378:metabolism; D016166:Free Radical Scavengers / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism*; D006131:Growth Inhibitors / Q000494:pharmacology; D007150:Immunohistochemistry; D007213:Indomethacin / Q000494:pharmacology; D020128:Inhibitory Concentration 50; D015289:Leukotrienes / Q000494:pharmacology; D009637:Masoprocol / Q000378:metabolism; D008636:Mesencephalon / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D009569:Nitric Oxide / Q000378:metabolism*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology; D011485:Protein Binding; D051381:Rats; D015398:Signal Transduction; D013997:Time Factors
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D003470:Culture Media; D016861:Cyclooxygenase Inhibitors; D004791:Enzyme Inhibitors; D044950:Flavanones; D005419:Flavonoids; D016166:Free Radical Scavengers; D006131:Growth Inhibitors; D015289:Leukotrienes; D010975:Platelet Aggregation Inhibitors; D009569:Nitric Oxide; C006680:baicalein; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; C021201:12-HPETE; D009637:Masoprocol; D001092:Arachidonate 12-Lipoxygenase; D017869:Cyclic GMP-Dependent Protein Kinases; D003022:Clotrimazole; D005978:Glutathione; D007213:Indomethacin