Glutathione depletion switches nitric oxide neurotrophic effects to cell death in midbrain cultures: implications for Parkinson's disease.
Nitric oxide (NO) exerts neurotrophic and neurotoxic effects on dopamine (DA) function in primary midbrain cultures. We investigate herein the role of glutathione (GSH) homeostasis in the neurotrophic effects of NO. Fetal midbrain cultures were pretreated with GSH synthesis inhibitor, L-buthionine-(S,R)-sulfoximine (BSO), 24 h before the addition of NO donors (diethylamine/nitric oxide-complexed sodium and S-nitroso-N-acetylpenicillamine) at doses tested previously as neurotrophic. Under these conditions, the neurotrophic effects of NO disappeared and turned on highly toxic. Reduction of GSH levels to 50% of baseline induced cell death in response to neurotrophic doses of NO. Soluble guanylate cyclase (sGC) and cyclic GMP-dependent protein kinase (PKG) inhibitors protected from cell death for up to 10 h after NO addition; the antioxidant ascorbic acid also protected from cell death but its efficacy decreased when it was added after NO treatment (40% protection 2 h after NO addition). The pattern of cell death was characterized by an increase in chromatin condensed cells with no DNA fragmentation and with breakdown of plasmatic membrane. The inhibition of RNA and protein synthesis and of caspase activity also protected from cell death. This study shows that alterations in GSH levels change the neurotrophic effects of NO in midbrain cultures into neurotoxic. Under these conditions, NO triggers a programmed cell death with markers of both apoptosis and necrosis characterized by an early step of free radicals production followed by a late requirement for signalling on the sGC/cGMP/PKG pathway.
Canals
S
S
Departamento de Investigación, Servicio de Neurobiología, Hospital Ramón y Cajal, Madrid, Spain.
Casarejos
M J
MJ
de Bernardo
S
S
Rodríguez-Martín
E
E
Mena
M A
MA
D000470:Alkaloids / Q000494:pharmacology; D000634:Aminoquinolines / Q000494:pharmacology; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D001205:Ascorbic Acid / Q000494:pharmacology; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D002227:Carbazoles; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured / Q000187:drug effects / Q000378:metabolism; D017869:Cyclic GMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000502:physiology; D004298:Dopamine / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005609:Free Radicals; D005978:Glutathione / Q000172:deficiency / Q000502:physiology*; D005981:Glutathione Synthase / Q000037:antagonists & inhibitors; D006162:Guanylate Cyclase / Q000037:antagonists & inhibitors / Q000502:physiology; D006706:Homeostasis; D006834:Hydrazines; D007211:Indoles; D008636:Mesencephalon / Q000166:cytology* / Q000196:embryology; D008751:Methylene Blue / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000032:analysis / Q000096:biosynthesis / Q000502:physiology; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009569:Nitric Oxide / Q000502:physiology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D009589:Nitrogen Oxides; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D010300:Parkinson Disease / Q000378:metabolism*; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014446:Tyrosine 3-Monooxygenase / Q000032:analysis / Q000096:biosynthesis
D000470:Alkaloids; D000634:Aminoquinolines; D000975:Antioxidants; D002227:Carbazoles; D004791:Enzyme Inhibitors; D005609:Free Radicals; D006834:Hydrazines; D007211:Indoles; D009419:Nerve Tissue Proteins; D020030:Nitric Oxide Donors; D009589:Nitrogen Oxides; D019384:Nucleic Acid Synthesis Inhibitors; D011500:Protein Synthesis Inhibitors; C110881:S-nitro-N-acetylpenicillamine; C073601:KT 5823; D009569:Nitric Oxide; D019328:Buthionine Sulfoximine; C084012:1,1-diethyl-2-hydroxy-2-nitrosohydrazine; C041715:6-anilino-5,8-quinolinedione; D014446:Tyrosine 3-Monooxygenase; D017869:Cyclic GMP-Dependent Protein Kinases; D006162:Guanylate Cyclase; D005981:Glutathione Synthase; D005978:Glutathione; D010396:Penicillamine; D001205:Ascorbic Acid; D008751:Methylene Blue; D004298:Dopamine