Previously we correlated the efficacy for G protein activation with that for arrestin recruitment for a number of agonists at the μ-opioid receptor (MOPr) stably expressed in HEK293 cells. We suggested that the endomorphins (endomorphin-1 and -2) might be biased toward arrestin recruitment. In the present study, we investigated this phenomenon in more detail for endomorphin-2, using endogenous MOPr in rat brain as well as MOPr stably expressed in HEK293 cells. For MOPr in neurons in brainstem locus ceruleus slices, the peptide agonists [d-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin (DAMGO) and endomorphin-2 activated inwardly rectifying K+ current in a concentration-dependent manner. Analysis of these responses with the operational model of pharmacological agonism confirmed that endomorphin-2 had a much lower operational efficacy for G protein-mediated responses than did DAMGO at native MOPr in mature neurons. However, endomorphin-2 induced faster desensitization of the K+ current than did DAMGO. In addition, in HEK293 cells stably expressing MOPr, the ability of endomorphin-2 to induce phosphorylation of Ser375 in the COOH terminus of the receptor, to induce association of arrestin with the receptor, and to induce cell surface loss of receptors was much more efficient than would be predicted from its efficacy for G protein-mediated signaling. Together, these results indicate that endomorphin-2 is an arrestin-biased agonist at MOPr and the reason for this is likely to be the ability of endomorphin-2 to induce greater phosphorylation of MOPr than would be expected from its ability to activate MOPr and to induce activation of G proteins.
This work was supported by the National Institutes of Health National Institute on Drug Abuse [Grant DA020836]; the Medical Research Council UK [Grant G0600943]; the Biotechnology and Biochemical Sciences Research Council [Grant BB/D012902/1]; and by a research fellowship from the Basque Government [Grant BFI08.131] (to G.R.).
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
- G protein-coupled receptor
- μ-opioid receptor
- locus ceruleus
- G protein-coupled, inwardly rectifying, K+ channel current
- yellow fluorescent protein
- cyan fluorescent protein
- G protein-coupled receptor kinase 2
- 4′,6-diamidino-2-phenylindole dihydrochloride
- guanosine 5′-O-(γ-thiotriphosphate)
- fluorescence resonance energy transfer.
- Received March 15, 2012.
- Accepted May 2, 2012.
- Copyright © 2012 The American Society for Pharmacology and Experimental Therapeutics