Abstract
In rat astrocyte-enriched culture, C2 ceramide dose- and time-dependently increased proenkephalin (proENK) mRNA; the significant increase began at 6 h after 30 microM C2 ceramide treatment (about 13-fold) and at 12 h after treatment (about 21-fold). In addition, C2 ceramide also increased AP-1 proteins, such as Fra-1, c-Jun, JunB and JunD, and phosphorylation of CREB. The blocking of protein synthesis by cycloheximide (CHX) evokes a further increase of C2 ceramide-induced proENK mRNA and phospho-CREB level, while C2 ceramide-induced increases of AP-1 protein levels were reduced by CHX. The C2 ceramide-induced proENK mRNA expression was not changed significantly by the pretreatment with H89 (a PKA inhibitor), KN62 (a calcium/calmodulin-dependent protein kinase II inhibitor), and PD98059 (an ERK pathway inhibitor). However, calphostin C (a PKC inhibitor) and or SB203580 (a p38 inhibitor) partially but significantly reduced C2 ceramide-induced proENK mRNA expression as well as phospho-CREB level. These results suggest that, in the rat astrocyte-enriched culture, C2 ceramide increases proENK mRNA expression via phosphorylation of CREB rather than the increases of AP-1 protein levels. Additionally, the activations of PKC and p38, but not PKA, calcium/calmodulin-dependent protein kinase II, and ERK, by C2 ceramide play important regulatory roles in C2 ceramide-induced proENK mRNA expression via activating the CREB.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / analogs & derivatives
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1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / pharmacology
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Animals
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Astrocytes / cytology
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Astrocytes / enzymology*
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Cells, Cultured
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Cyclic AMP Response Element-Binding Protein / metabolism*
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Cycloheximide / pharmacology
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Enkephalins / genetics*
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Enzyme Inhibitors / pharmacology
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Flavonoids / pharmacology
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Gene Expression / drug effects
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Imidazoles / pharmacology
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Isoquinolines / pharmacology
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Mitogen-Activated Protein Kinases / metabolism*
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Naphthalenes / pharmacology
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Phosphorylation
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Protein Precursors / genetics*
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Protein Synthesis Inhibitors / pharmacology
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Pyridines / pharmacology
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RNA, Messenger / metabolism
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Rats
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Rats, Sprague-Dawley
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Sphingosine / analogs & derivatives
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Sphingosine / pharmacology*
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Stimulation, Chemical
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Sulfonamides*
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Transcription Factor AP-1 / metabolism
Substances
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Cyclic AMP Response Element-Binding Protein
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Enkephalins
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Enzyme Inhibitors
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Flavonoids
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Imidazoles
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Isoquinolines
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N-acetylsphingosine
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Naphthalenes
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Protein Precursors
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Protein Synthesis Inhibitors
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Pyridines
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RNA, Messenger
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Sulfonamides
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Transcription Factor AP-1
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proenkephalin
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KN 62
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1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine
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Cycloheximide
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Mitogen-Activated Protein Kinases
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calphostin C
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N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide
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Sphingosine
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SB 203580
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2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one