title: Orphanin FQ/nociceptin and mu-opioid receptor mRNA levels in human SH-SY5Y neuroblastoma cells: effects of activating the cAMP-PKA signal transduction pathway.
abstract: Responses to opioid agonists vary, depending on past opioid exposure and the physiological state. The intracellular signaling pathway mediated by cAMP and protein kinase A (PKA) has been linked to regulation of opioid receptor responsiveness. The role of the cAMP-PKA pathway in regulating opioid receptor gene expression is incompletely defined. Mu-opioid receptor (MuOR) and orphanin FQ/nociceptin receptor (ORL(1)) transcripts were measured after activating this pathway in human neuroblastoma cells. Human SH-SY5Y neuroblastoma cells were maintained in continuous monolayer culture. Cells were incubated with combinations of agents which activate the cAMP-PKA signal transduction pathway, including forskolin and choleratoxin (CTX). MuOR and ORL(1) transcript levels were measured by hybridization to specific probes. Activation of the cAMP-PKA signal transduction pathway with forskolin in the presence of phosphodiesterase inhibitors was associated with a time-dependent decrease in the level of MuOR mRNA; partial recovery was observed with prolonged incubations. Forskolin effects were mimicked by CTX, but not by dideoxyforskolin. The PKA inhibitor H89 blunted the actions of forskolin. However, forskolin responses persisted despite coincubation with protein synthesis inhibitors. ORL(1) transcript levels did not significantly change, but vasoactive intestinal polypeptide (VIP) transcripts exhibited substantial increases, in the presence of forskolin or CTX. These observations support a role for cAMP in regulating MuOR responsiveness through actions at the level of receptor gene expression. ORL(1) transcript levels are not effected, suggesting that the cAMP-PKA pathway has differential effects on the expression of mRNA for different, but biochemically closely related, opioid receptor subtypes.
authors.affiliation: Beecher Memorial Laboratory, Department of Anesthesia and Critical Care, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114-2696, USA.
authors.forename: Cara-Louise
authors.initials: CL
authors.lastname: Wallington
authors.forename: Dawn M
authors.initials: DM
authors.lastname: Peters
authors.forename: Carol R
authors.initials: CR
authors.lastname: Gelb
authors.forename: Robert A
authors.initials: RA
authors.lastname: Peterfreund
mesh_terms: D000701:Analgesics, Opioid / Q000494:pharmacology*; D002490:Central Nervous System / Q000187:drug effects* / Q000378:metabolism; D002772:Cholera Toxin / Q000494:pharmacology; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000187:drug effects* / Q000378:metabolism; D015536:Down-Regulation / Q000187:drug effects / Q000235:genetics; D004361:Drug Tolerance / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D006801:Humans; D009447:Neuroblastoma; D015345:Oligonucleotide Probes; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D011957:Receptors, Opioid / Q000235:genetics*; D017450:Receptors, Opioid, mu / Q000096:biosynthesis / Q000235:genetics*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology; D014407:Tumor Cells, Cultured; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology; D014660:Vasoactive Intestinal Peptide / Q000378:metabolism; D000094942:Nociceptin Receptor
chemical_list: D000701:Analgesics, Opioid; D004791:Enzyme Inhibitors; D015345:Oligonucleotide Probes; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D011957:Receptors, Opioid; D017450:Receptors, Opioid, mu; D005576:Colforsin; D014660:Vasoactive Intestinal Peptide; D002772:Cholera Toxin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D000094942:Nociceptin Receptor; C000722283:OPRL1 protein, human