Sensitization of the Adenylyl Cyclase System in Cloned k-Opioid Receptor-Transfected Cells Following Sustained Agonist Treatment. A Chimeric Study Using G Protein .ALPHA.i2/.ALPHA.q Subunits.

Chronic and/or sustained opioid treatment has been shown to result in development of sensitization of the adenylyl cyclase (AC) system or cAMP overshoot. In this study, we investigated the molecular mechanism responsible for sensitization of the AC system using CHO cells co-expressing cloned kappa-opioid receptor and some chimeric G protein alpha(i2)/alpha(q) subunits. In CHO cells co-expressing the kappa-opioid receptor and pertussis toxin-insensitive chimeric alpha(i2)/alpha(q) subunits with alpha(i2) residues Met244-Asn331, despite pretreatment with pertussis toxin, acute treatment with the kappa-opioid-receptor-selective agonist U69,593 suppressed forskolin-stimulated cAMP accumulation, while sustained treatment with U69,593 (4 h) induced cAMP overshoot over the naive level by the kappa-opioid-receptor-selective antagonist norbinaltorphimine (sensitization of the AC system). On the other hand, in CHO cells co-expressing the kappa-opioid receptor and pertussis toxin-insensitive chimeric alpha(i2)/alpha(q) subunits without alpha(i2) residues Met244-Asn331, pretreatment with pertussis toxin completely blocked these acute and sustained effects of U69,593 on cAMP accumulation. These results suggested that the presence of the specific region of alpha(i2) (Met244-Asn331), which was reported to be responsible for the inhibition of AC, and continuous inhibition of AC by alpha(i2) is necessary for the development of sensitization.