title: Effects of chronic nociceptin/orphanin FQ exposure on cAMP accumulation and receptor density in Chinese hamster ovary cells expressing human nociceptin/orphanin FQ receptors.
abstract: Nociceptin/orphanin FQ (N/OFQ) is the endogenous ligand for the G(i)-coupled N/OFQ receptor (NOP). We have examined the effects of chronic exposure of Chinese hamster ovary cells expressing the recombinant human NOP receptor (CHO(hNOP)) to 1 nM N/OFQ for up to 48 h in the absence and presence of the NOP selective antagonist [Nphe(1)]N/OFQ (1-13)NH(2) ([Nphe(1)]). Then, either a concentration-response curve for N/OFQ inhibition of cAMP formation was constructed or the cells were homogenized and membrane receptor density was determined using [(125)I]Y(14)N/OFQ. There was a time-dependent reduction in pEC(50) (without a change in maximum) for N/OFQ with significant differences observed following >24 h of exposure (control pEC(50) approximately 9.5; 48 h pretreatment approximately 8.7). In cells co-exposed to N/OFQ+[Nphe(1)] for 48 h, there was no reduction in pEC(50). There was a compensatory (approximately 2.5-fold), [Nphe(1)]-sensitive increase in cAMP mass in cells exposed to N/OFQ for 24-48 h. N/OFQ pretreatment also resulted in a time-dependent [Nphe(1)]-sensitive loss of cell surface receptors. At 48 h, B(max) was reduced from approximately 2.0 to approximately 1.3 pmol mg(-1) protein without a change in pK(d) for N/OFQ. There was a positive correlation between pEC(50) for cAMP inhibition and B(max). The lack of effect on maximum cAMP response probably results from receptor overexpression and the creation of a receptor reserve.
authors.affiliation: University Department of Anaesthesia, Critical Care and Pain Management, Leicester Royal Infirmary, Leicester, LE1 5WW, UK.
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Hashimoto
authors.forename: Girolamo
authors.initials: G
authors.lastname: Calo'
authors.forename: Remo
authors.initials: R
authors.lastname: Guerrini
authors.forename: Graham
authors.initials: G
authors.lastname: Smith
authors.forename: David G
authors.initials: DG
authors.lastname: Lambert
mesh_terms: D000818:Animals; D016466:CHO Cells; D005576:Colforsin / Q000494:pharmacology; D006224:Cricetinae; D000242:Cyclic AMP / Q000378:metabolism*; D015536:Down-Regulation / Q000187:drug effects; D006801:Humans; D007700:Kinetics; D009292:Narcotic Antagonists; D018847:Opioid Peptides / Q000378:metabolism / Q000494:pharmacology*; D011480:Protease Inhibitors / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000187:drug effects; D011957:Receptors, Opioid / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D000094942:Nociceptin Receptor; D000097629:Nociceptin
chemical_list: D005576:Colforsin; D000242:Cyclic AMP; D009292:Narcotic Antagonists; D018847:Opioid Peptides; D011480:Protease Inhibitors; D011956:Receptors, Cell Surface; D011957:Receptors, Opioid; D011994:Recombinant Proteins; D000094942:Nociceptin Receptor; D000097629:Nociceptin