Opioid efficacy in a C6 glioma cell line stably expressing the delta opioid receptor

A C6 glioma cell line stably transfected with the rat delta opioid receptor (C6δ) was used to characterize receptor binding and G protein activation by both peptide and nonpeptide delta opioid ligands. The ligand binding affinities for [³H]naltrindole and [³H]pCl-[D-Pen²,D- Pen⁵]enkephalin (DPDPE) were similar to those observed in monkey brain membranes. The nonpeptide agonists, BW373U86 and SNC80, as well as peptide agonist [D-Ser²,L-Leu⁵]enkephalyl-Thr maximally stimulated [³⁵S]GTPγS binding by 640, 654 and 576%, respectively, over basal. The peptide agonists, DPDPE and deltorphin II, both stimulated [³⁵S]GTPγS binding by 375%. Etorphine, diprenorphine, oxymorphindole and 7-spiroindanyloxymorphone were also partial agonists in this assay, although they were less efficacious than deltorphin II. Stimulation of [³⁵S]GTPγS binding by agonists was blocked completely by pertussis toxin pretreatment. Both delta-1 and delta-2 selective antagonists 7-benzylidenenaltrexone and a benzofuran analog of naltrindole displayed high affinity for the cloned receptor (0.04 and 0.08 nM) and antagonized the stimulation of [³⁵S]GTPγS binding by BW373U86 and DPDPE with similar potencies. Other evidence suggesting the lack of receptor subtypes includes the finding that stimulation of [³⁵S]GTPγS binding by receptor subtype selective ligands DPDPE and deltorphin II was not additive. BW373U86, SNC8O and DPDPE maximally inhibited forskolin-stimulated adenylyl cyclase. These cells highly express a homogeneous population of delta opioid receptor that couple to inhibitory G(o)/G(i) proteins. Ligand affinity for the delta opioid receptor correlates with ligand EC₅₀ values for stimulation of [³⁵S]GTPγS binding.