Received for publication July 6, 2004.
Revised October 18, 2004.
Accepted for publication October 19, 2004.

RECIPROCAL REGULATION OF AGONIST AND INVERSE AGONIST SIGNALLING EFFICACY UPON SHORT-TERM TREATMENT OF THE HUMAN DELTA-OPIOID RECEPTOR WITH AN INVERSE AGONIST

Abstract

Rapid regulation of receptor signaling by agonist ligands is widely accepted, whereas short-term adaptation to inverse agonists has been little documented. In the present study, [³⁵S]GTPS binding and cAMP accumulation assays were used to assess the consequences of 30 min exposure to the inverse agonist ICI174864 (1µM), on delta opioid receptor (OR) signaling efficacy. ICI174864 pretreatment increased maximal effect (Emax) for the partial agonist TIPP at the two levels of the signaling cascade while Emax for more efficacious agonists like SNC-80 and bremazocine, were increased in [³⁵S]GTPS binding but not in cAMP accumulation assays. Preexposure to ICI174864 also induced a shift to the left in dose response curves for bremazocine and TIPP. Conversely, Emax for the inverse agonist TICP was reduced in both assays, but no changes in potency were observed. For the weaker inverse agonist naloxone, Emax in [³⁵S]GTPS binding was drastically modified since the drug turned from inverse agonist to agonist following ICI174864 pretreatment. Similarly, ICI174864 turned from inverse agonist to agonist when tested in cAMP accumulation assays. In both cases, inversion of efficacy was concomitant with marked increase in potency for agonist effects. Together with functional changes, short-term treatment with ICI174864 reduced basal receptor phosphorylation and increased immunoreactivity for G_i3 in membrane preparations. Functional consequences of ICI174864 pretreatment were simulated in the cubic ternary complex model by increasing receptor-G protein coupling or G protein amount available for interaction with the receptor. Taken together, these data show that inverse agonists may induce rapid regulation in receptor signaling efficacy.

Key words: Neuropeptides, Opioid, Gi family, Adenylyl cyclases, Phosphorylation/Dephosphorylation, Receptor binding studies, Opioids