Effects of chronic nociceptin/orphanin FQ exposure on cAMP accumulation and receptor density in Chinese, hamster ovary cells expressing human nociceptin/orphanin FQ receptors

Nociceptin/orphanin FQ (N/OFQ) is the endogenous ligand for the Gi-coupled N/OFQ receptor (NOP). We have examined the effects of chronic exposure of Chinese hamster ovary cells expressing the recombinant human NOP receptor (CHOhNOP) to 1 nM N/OFQ for up to 48 h in the absence and presence of the NOP selective antagonist [Nphe1]N/OFQ (1–13)NH2 ([Nphe1]). 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 [125I]Y14N/OFQ. There was a time-dependent reduction in pEC50 (without a change in maximum) for N/OFQ with significant differences observed following > 24 h of exposure (control pEC50f9.5; 48 h pretreatmentf8.7). In cells co-exposed to N/ OFQ+[Nphe1] for 48 h, there was no reduction in pEC50. There was a compensatory (f2.5-fold), [Nphe1]-sensitive increase in cAMP mass in cells exposed to N/OFQ for 24–48 h. N/OFQ pretreatment also resulted in a time-dependent [Nphe1]-sensitive loss of cell surface receptors. At 48 h, Bmax was reduced from f2.0 to f1.3 pmol mg1 protein without a change in pKd for N/OFQ. There was a positive correlation between pEC50 for cAMP inhibition and Bmax. The lack of effect on maximum cAMP response probably results from receptor overexpression and the creation of a receptor reserve.