Abstract
The μ-opioid receptor (MOR) couples to multiple G proteins, of which coupling to Gs has long been debated. As expected, in opioid naive Chinese hamster ovary cells expressing recombinant MOR, the predominant action of [d-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin (DAMGO) is inhibitory. However, inactivation of Gi/Go proteins via pertussis toxin (PTX) unmasks its ability to facilitate forskolin activation of adenylyl cyclase (AC) activity. Tolerance develops to this effect of DAMGO, which can also be attenuated by cholera toxin (CTX). The latter suggests G mediation. d-Phe-Cys-Tyr-d-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP), previously considered to be a neutral MOR antagonist, also produces a facilitation of forskolin (FSK) activation of AC that is augmented by chronic morphine. Facilitative effects of CTAP in naive as well as its augmentation in tolerant membranes are both substantially reduced by CTX. This suggests that not only Gs mediation but also Gsα-linked signaling is critical to the chronic morphine-induced enhanced facilitative action of CTAP. Interestingly, the (augmented) CTAP facilitation of FSK-stimulated AC activity that is observed in opioid tolerant (but not in naive) membranes is also sensitive to PTX. This can best be explained by postulating the involvement of Gi-derived Gβγ, which would stimulate type 2 ACs, conditional on the presence of activated Gsα. The emergence of a Gβγ dimension of AC stimulation by CTAP after chronic morphine could explain its ability to augment the stimulatory action of CTAP on AC. These results support putative MOR coupling to Gs and underscore the multifaceted nature and plasticity of MOR G protein coupling.
Footnotes
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DOI: 10.1124/jpet.104.066837.
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ABBREVIATIONS: GPCR, G protein-coupled receptor; MOR, μ-opioid receptor; CTAP, d-Phe-Cys-Tyr-d-Trp-Arg-Thr-Pen-Thr-NH2; ICI174864, N,N-diallyl-Tyr-Aib-Aib-Phe-Leu; CHO, Chinese hamster ovary; MOR-CHO, Chinese hamster ovary cells stably transfected with μ-opioid receptors; PTX, pertussis toxin; CTX, cholera toxin; DAMGO, [d-Ala2,N-Me-Phe4,Gly5-ol]-enkephalin; FSK, forskolin; AC, adenylyl cyclase; ANOVA, analysis of variance.
- Received February 10, 2004.
- Accepted March 2, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
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