Role for G protein-coupled receptor kinase in agonist-specific regulation of μ-opioid receptor responsiveness

Abstract

The G protein-coupled -opioid receptor (OR) mediates the physiological effects of endogenous opioid peptides as well as the structurally distinct opioid alkaloids morphine and etorphine. An intriguing feature of OR signaling is the differential receptor trafficking and desensitization properties following activation by distinct agonists, which have been proposed as possible mechanisms related to opioid tolerance. Here we report that the ability of distinct opioid agonists to differentially regulate OR internalization and desensitization is related to their ability to promote G protein-coupled receptor kinase (GRK)-dependent phosphorylation of the OR. Although both etorphine and morphine effectively activate the OR, only etorphine elicits robust OR phosphorylation followed by plasma membrane translocation of ␤-arrestin and dynamin-dependent receptor internalization. In contrast, corresponding to its inability to cause OR internalization, morphine is unable to either elicit OR phosphorylation or stimulate ␤-arrestin translocation. However, upon the overexpression of GRK2, morphine gains the capacity to induce OR phosphorylation, accompanied by the rescue of ␤-arrestin translocation and receptor sequestration. Moreover, overexpression of GRK2 also leads to an attenuation of morphine-mediated inhibition of adenylyl cyclase. These findings point to the existence of marked differences in the ability of different opioid agonists to promote OR phosphorylation by GRK. These differences may provide the molecular basis underlying the different analgesic properties of opioid agonists and contribute to the distinct ability of various opioids to induce drug tolerance.

Internalization and trafficking of opioid receptor ligands in rat cortical neurons

“…Scale bar ϭ 10 m. the addition of the inhibitor of clathrin-mediated endocytosis, phenylarsine oxide. Ligand-induced, clathrinmediated internalization of OR had previously been documented in both neuronal Sternini et al, 1996Sternini et al, , 2000Trafton et al, 2000) and nonneuronal cells (von Zastrow et al, 1993;Burford et al, 1998;Arden et al, 1995;Keith et al, 1996;Gaudriault et al, 1997;Whistler et al, 1999;Zhang et al, 1998). ␦OR had also been shown to internalize in a clathrinmediated fashion, but only in transfected cell systems (Chu et al, 1997;Zhang et al, 1999) and in a neural cell line (Ko et al, 1999).…”

Brain region-specific N-glycosylation and lipid rafts association of the rat mu opioid receptor

“…Thus, a question arises as whether the heterogeneity of MOR in N-glycosylation and/or raftsdistribution plays a role in such differential MOR phosphorylation and desensitization in different brain regions. In in vitro cellular systems, expression of G protein-coupled receptor kinase 2 (GRK2) enhanced phosphorylation and regulation of MOR [34,35], and most of GRK2 was shown to be not associated with rafts [36]. Since GRK2 is ubiquitously expressed in the brain, this regulation was expected to occur in vivo [37].…”

Understanding the Effect of Different Assay Formats on Agonist Parameters: A Study Using the µ-Opioid Receptor

“…17 in the case of the µ-opioid receptor, we and others have reported that compounds can display clearly different profiles in G-protein activation and β-arrestin recruitment, with morphine exhibiting low emax values in β-arrestin recruitment/receptor internalization but high emax values in assays measuring G-protein activation. [18][19][20][21] However, emax values are not always a good indicator of efficacy. Herein, we have shown that some ligands do show agonist-directed conformational states; for instance, morphine does not differentiate from the other ligands in terms of efficacy (tau) for G-protein activation but does differentiate in terms of efficacy (emax) in the β-arrestin assay.…”