Ethanol increases tyrosine hydroxylase gene expression in N1E-115 neuroblastoma cells.
Catecholamines appear to be involved in behavioral responses to acute and chronic ethanol consumption. Since tyrosine hydroxylase (TH) is the rate-limiting enzyme for catecholamine biosynthesis and is regulated by second messenger systems known to be modulated by ethanol, we studied ethanol-induced changes in TH gene expression. In the N1E-115 neural cell line, Northern and Western blot analyses showed that treatment with 25-200 mM ethanol for 3 days caused a dose-dependent increase in TH mRNA and protein levels. N1E-115 cells were also stably transfected with pTH5'CAT, a plasmid containing 773 base pairs of the TH promoter fused to a chloramphenicol acetyltransferase (CAT) reporter gene. Subclones expressing pTH5'CAT showed ethanol-induced increases in CAT activity, suggesting that ethanol modulates TH gene transcription. Furthermore, simultaneous treatment of transfected cells with 100 mM ethanol and 1 nM to 1 microM prostaglandin E1 increased prostaglandin E1-mediated stimulation of TH-promoter activity. Similarly, simultaneous treatment of transfected cells with 100 mM ethanol and either 10 mM (-)-N6-(R-phenylisopropyl)adenosine or 0.5 mM 8-bromo-cAMP also resulted in increased TH-promoter activity compared to treatment with these agents without ethanol. These results suggest that ethanol treatment of N1E-115 cells has a prominent effect on both basal and cAMP-regulated TH expression. Ethanol-induced changes in TH expression may be a critical molecular event in adaptation of the central nervous system to ethanol.
Department of Pharmacology, University of California, San Francisco General Hospital 94110.
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Gayer
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Gordon
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Miles
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000527:Alprostadil / Q000494:pharmacology; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D000431:Ethanol / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D007700:Kinetics; D009447:Neuroblastoma; D010660:Phenylisopropyladenosine / Q000494:pharmacology; D010957:Plasmids; D011401:Promoter Regions, Genetic / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics*
D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D015124:8-Bromo Cyclic Adenosine Monophosphate; D010660:Phenylisopropyladenosine; D000431:Ethanol; D014446:Tyrosine 3-Monooxygenase; D015500:Chloramphenicol O-Acetyltransferase; D000527:Alprostadil