Abstract
Ethanol stimulates hypothalamic-pituitary-adrenal axis activity in vivo. To determine the cellular and molecular mechanisms through which ethanol regulates corticotropin-releasing factor (CRF) gene expression, we compared the effect of ethanol and forskolin on CRF peptide secretion and messenger RNA levels in hypothalamic primary cell cultures, and on CRF promoter activity in the NG108-15 cell line. CRF secretion, mRNA levels, and gene transcription significantly increased in response to ethanol or forskolin. Mutation of the cAMP-response element (CRE) reduced luciferase activity under basal conditions as well as in response to forskolin or ethanol. On the other hand, plasmid with five CRE repeats yielded dramatically elevated basal luciferase activity and significantly increased upregulation by ethanol. Inclusion of adenosine deaminase reduced the promoter response to ethanol. Finally a PKA inhibitor and a cAMP antagonist both decreased ethanol-induced CRF peptide secretion, gene expression, and transcription. These results suggest that ethanol upregulates CRF expression through cAMP/PKA-dependent pathways.
Publication types
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Comparative Study
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Research Support, N.I.H., Extramural
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Animals
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Animals, Newborn
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Cells, Cultured
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Central Nervous System Depressants / pharmacology
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Colforsin / pharmacology
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Corticotropin-Releasing Hormone / genetics*
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Corticotropin-Releasing Hormone / metabolism
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Cyclic AMP / antagonists & inhibitors
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Cyclic AMP / metabolism
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Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
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Cyclic AMP-Dependent Protein Kinases / metabolism
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Enzyme Inhibitors / pharmacology
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Ethanol / pharmacology*
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Gene Expression / drug effects
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Gene Expression / physiology
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Hypothalamo-Hypophyseal System / cytology
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Hypothalamo-Hypophyseal System / drug effects*
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Hypothalamo-Hypophyseal System / metabolism
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Hypothalamus / cytology
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Hypothalamus / drug effects*
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Hypothalamus / metabolism
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Luciferases / genetics
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Plasmids / genetics
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Promoter Regions, Genetic / drug effects
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Promoter Regions, Genetic / physiology
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RNA, Messenger / drug effects
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RNA, Messenger / metabolism
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Rats
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Rats, Sprague-Dawley
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Response Elements / drug effects
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Response Elements / physiology
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Transcriptional Activation / drug effects
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Transcriptional Activation / physiology
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Up-Regulation / drug effects
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Up-Regulation / physiology
Substances
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Central Nervous System Depressants
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Enzyme Inhibitors
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RNA, Messenger
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Colforsin
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Ethanol
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Corticotropin-Releasing Hormone
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Cyclic AMP
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Luciferases
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Cyclic AMP-Dependent Protein Kinases