Adenylyl cyclase (AC) superactivation is thought to play an important role in opioid tolerance, dependence, and withdrawal. In the present study, we investigated the involvement of protein kinases in chronic δ-opioid agonist-mediated AC superactivation in Chinese hamster ovary (CHO) cells stably expressing the human δ-opioid receptor (hDOR/CHO). Maximal forskolin-stimulated cAMP formation in hDOR/CHO cells increased by 472 ± 91, 399 ± 2, and 433 ± 73% after chronic treatment with the δ-opioid agonists (+)-4-[(αR)-α-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3- methoxy-benzyl]-N,N-diethyl benzamide (SNC 80), [D-Pen2,D-Pen5]-enkephalin, and deltorphin II, respectively. Concurrently, chronic SNC 80 (1 μM, 4-h) treatment augmented 32p incorporation into a 200-kDa protein immunoreactive with the ACV/VI antibody by 300 ± 60% in hDOR/CHO cell lysates. The calmodulin antagonist calmidazolium significantly attenuated chronic deltorphin II-mediated AC superactivation. Tyrosine kinase (genistein) and protein kinase C (chelerythrine) inhibitors individually had minimal effect on chronic δ-opioid agonist-mediated AC superactivation. Conversely, simultaneous treatment with both genistein and chelerythrine significantly attenuated AC superactivation. Because we showed previously that the Raf-1 inhibitor 3-(3,5-dibromo-4-hydroxybenzylidene-5-iodo-1,3-dihydro-indol-2-one (GW5074) attenuates AC superactivation, we hypothesize that parallel calmidazolium-, chelerythrine-, and genistein-sensitive pathways converge at Raf-1 to mediate AC superactivation by phosphorylating AC VI in hDOR/CHO cells.
|Original language||English (US)|
|Number of pages||7|
|Journal||Journal of Pharmacology and Experimental Therapeutics|
|State||Published - Jul 1 2003|
ASJC Scopus subject areas
- Molecular Medicine