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 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.