Characterization of opioid agonist efficacy in a C6 glioma cell line expressing the μ opioid receptor

P. J. Emmerson, M. J. Clark, A. Mansour, H. Akil, J. H. Woods, F. Medzihradsky

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174 Scopus citations


In C6 glioma cells stably expressing a homogeneous population of the cloned rat μ opioid receptor, the binding affinities of opioid agonists and subsequent activation of G protein were examined. Opioid receptor number in membranes of these cells was high (10-30 pmol/mg protein [3H]diprenorphine binding sites). Opioids were found to bind to the receptor with high affinity [Tyr-D-Ala-Gly-(Me)Phe-Gly-ol (DAMGO) 0.23 nM; sufentanil 0.034 nM; morphine 0.16 nM]. Activation of G protein by opioid agonists was examined by measuring the stimulation of guanosine-5'-O-(3-[35S]thio)triphosphate ([35S]GTPγS) binding. Sufentanil increased [35S]GTPγS binding by 326% with an EC50 value of 2.39 nM. Agonist stimulation of [35S]GTPγS binding was stereoselective, naltrexone-reversible, and pertussis toxin-sensitive. The 'intrinsic activity' of opioids at the μ receptor was reflected by the magnitude of agonist-mediated activation of G protein. The rank order of the stimulation of [35S]GTPγS binding was etonitazene = sufentanil = DAMGO = PLO17 = fentanyl > morphine > profadol > meperidine > butorphanol = nalbuphine = pentazocine > cyclazocine = nalorphine > levallorphan > naltrexone. High affinity binding of ligands to the μ opioid receptor was reduced by the addition of sodium and guanosine diphosphate at concentrations used in the [35S]GTPγS binding assay. Ligand affinity was reduced in a manner correlating with 'intrinsic activity': DAMGO, 1229-fold, nalbuphine 35-fold, naltrexone, 3-fold. The results presented show that the stable expression of the rat μ opioid receptor in C5 cells provides an effective tool to examine opioid receptor signal transduction mechanisms and evaluate the activity of novel opioids at the μ receptor.

Original languageEnglish (US)
Pages (from-to)1121-1127
Number of pages7
JournalJournal of Pharmacology and Experimental Therapeutics
Issue number3
StatePublished - Sep 1996
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology


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