Opioid efficacy in a C6 glioma cell line stably expressing the human kappaopioid receptor

A C6 glioma cell line stably transfected with the human kappa opioid recept or (kappa OR) was used to characterize receptor binding and G protein activ ation via the kappa OR by a comprehensive series of opioid ligands. The lig and-binding affinity for [H-3]5 alpha,7 alpha,8 beta(-)-N-methyl-N-(7-Cl-py rrolidinyl)-1-oxaspiro(4,5)dec-8-yl)benzene acetamide (U69593) was similar to that observed in monkey brain membranes and was 10-fold lower in the pre sence of sodium and GDP. Both peptide and nonpeptide agonists maximally sti mulated [S-35]GTP gamma S binding. The stimulation of [S-35]GTP gamma S bin ding was blocked by pretreatment of cells with pertussis toxin. Partial sti mulation of [S-35]GTP gamma S binding via the kappa OR was observed for sev eral ligands that are antagonists at the mu opioid receptor, suggesting an additional mechanism of drug action. The ability of isomers of tifluadom an d levallorphan to stimulate [S-35]GTP gamma S binding indicates that the ch iral carbon of levallorphan, a benzomorphan derivative, imparts a greater d egree of stereoselectivity than does the chiral carbon in the benzodiazepin e derivative tifluadom. In addition, (-)tifluadom, the less potent isomer o f tifluadom, which is also a gamma-aminobutyric acid(A) receptor agonist, s timulated [S-35]GTP gamma S binding. in contrast, d-pentazocine, (+)SKF1004 7, (+)cyclazocine, and d-ethylketocyclazocine displayed no agonist activity . kappa OR-selective antagonist norbinaltorphimine competitively inhibited the stimulation of [S-35]GTP gamma S binding by the active isomers of ethyl ketocyclazocine, cyclazocine, and nalorphine to the same degree, indicating that all three ligands are eliciting an effect via the kappa OR. The resul ts suggest that these cells express a homogeneous population of kappa OR, a nd that their [S-35]GTP gamma S-binding properties make them an excellent m eans to assess kappa OR efficacy.