Neuronal apoptosis induced by pharmacological concentrations of 3-hydroxykynurenine: characterization and protection by dantrolene and Bcl-2 overexpression.
We have studied neurotoxicity induced by pharmacological concentrations of 3-hydroxykynurenine (3-HK), an endogenous toxin implicated in certain neurodegenerative diseases, in cerebellar granule cells, PC12 pheochromocytoma cells, and GT1-7 hypothalamic neurosecretory cells. In all three cell types, the toxicity was induced in a dose-dependent manner by 3-HK at high micromolar concentrations and had features characteristic of apoptosis, including chromatin condensation and internucleosomal DNA cleavage. In cerebellar granule cells, the 3-HK neurotoxicity was unaffected by xanthine oxidase inhibitors but markedly potentiated by superoxide dismutase and its hemelike mimetic, MnTBAP [manganese(III) tetrakis(benzoic acid)porphyrin chloride]. Catalase blocked 3-HK neurotoxicity in the absence and presence of superoxide dismutase or MnTBAP. The formation of H(2)O(2) was demonstrated in PC12 and GT1-7 cells treated with 3-HK, by measuring the increase in the fluorescent product, 2',7'-dichlorofluorescein. In both PC12 and cerebellar granule cells, inhibitors of the neutral amino acid transporter that mediates the uptake of 3-HK failed to block 3-HK toxicity. However, their toxicity was slightly potentiated by the iron chelator, deferoxamine. Taken together, our results suggest that neurotoxicity induced by pharmacological concentrations of 3-HK in these cell types is mediated primarily by H(2)O(2), which is formed most likely by auto-oxidation of 3-HK in extracellular compartments. 3-HK-induced death of PC12 and GT1-7 cells was protected by dantrolene, an inhibitor of calcium release from the endoplasmic reticulum. The protection by dantrolene was associated with a marked increase in the protein level of Bcl-2, a prominent antiapoptotic gene product. Moreover, overexpression of Bcl-2 in GT1-7 cells elicited by gene transfection suppressed 3-HK toxicity. Thus, dantrolene may elicit its neuroprotective effects by mechanisms involving up-regulation of the level and function of Bcl-2 protein.
Wei
H
H
Section on Molecular Neurobiology, Biological Psychiatry Branch, National Institute of Mental Health, National Institutes of Health, Bethesda, Maryland, USA.
Leeds
P
P
Chen
R W
RW
Wei
W
W
Leng
Y
Y
Bredesen
D E
DE
Chuang
D M
DM
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002531:Cerebellum / Q000166:cytology / Q000378:metabolism; D002843:Chromatin / Q000648:ultrastructure; D053938:DNA Fragmentation; D003620:Dantrolene / Q000494:pharmacology*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016166:Free Radical Scavengers; D015870:Gene Expression; D007502:Iron Chelating Agents / Q000494:pharmacology; D007737:Kynurenine / Q000031:analogs & derivatives* / Q000494:pharmacology; D008665:Metalloporphyrins / Q000494:pharmacology; D009474:Neurons / Q000502:physiology*; D016716:PC12 Cells; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013482:Superoxide Dismutase / Q000494:pharmacology; D014969:Xanthine Oxidase / Q000037:antagonists & inhibitors
D002843:Chromatin; D004791:Enzyme Inhibitors; D016166:Free Radical Scavengers; D007502:Iron Chelating Agents; D008665:Metalloporphyrins; D019253:Proto-Oncogene Proteins c-bcl-2; C097284:manganese(III)-tetrakis(4-benzoic acid)porphyrin; C005045:3-hydroxykynurenine; D007737:Kynurenine; D013482:Superoxide Dismutase; D014969:Xanthine Oxidase; D003620:Dantrolene