title: Thapsigargin modulates agonist-stimulated cyclic AMP responses through cytosolic calcium-dependent and -independent mechanisms in rat pinealocytes.
abstract: The role of mobilization of intracellular Ca2+ in the adrenergic-stimulated cAMP accumulation in rat pinealocytes was investigated with thapsigargin, an agent that inhibits endoplasmic reticulum Ca2+-ATPase. It was found that although thapsigargin alone had no effect on the basal cAMP accumulation, it potentiated the beta-adrenergic-stimulated cAMP response by isoproterenol in a dose-dependent manner. The potentiation was abolished with ethylene glycol bis(beta-aminoethyl ether)-N, N,N',N'-tetraacetic acid-acetoxymethyl ester (EGTA-AM) but persisted in the presence of isobutylmethylxanthine, indicating that thapsigargin enhances cAMP synthesis through elevation of cytosolic intracellular Ca2+ concentration ([Ca2+]i). However, when the pinealocytes were stimulated by norepinephrine, a mixed alpha 1- and beta-adrenergic agonist, thapsigargin dose-dependently inhibited the cAMP response. To investigate this inhibitory effect of thapsigargin, we substituted ionomycin, a [Ca2+]i-elevating agent, and 4 beta-phorbol-12-myristate 13-acetate, an activator of protein kinase C, for the alpha(1)-adrenergic component of the norepinephrine-stimulated response. Although thapsigargin had no effect on the potentiation of the isoproterenol-stimulated cAMP accumulation by ionomycin, it significantly inhibited the potentiation by 4 beta-phorbol-12-myristate 13-acetate. Furthermore, the inhibitory effect of thapsigargin was not affected by cotreatment with EGTA-AM or ionomycin, suggesting that this effect is independent of [Ca2+]i. Similar results were obtained when cyclopiazonic acid was used to inhibit the Ca(2+)-ATPase. Taken together, our results indicate that thapsigargin enhances the beta-adrenergic-stimulated cAMP accumulation through its action in elevating [Ca2+]i but inhibits the potentiation of the beta-adrenergic-stimulated cAMP response by protein kinase C, as a consequence of Ca(2+)-ATPase inhibition.
authors.affiliation: Department of Physiology, Faculty of Medicine, University of Alberta, Edmonton, Canada. aho@physio.med.ualberta.ca
authors.forename: A K
authors.initials: AK
authors.lastname: Ho
authors.forename: T
authors.initials: T
authors.lastname: Ogiwara
authors.forename: C L
authors.initials: CL
authors.lastname: Chik
mesh_terms: D000818:Animals; D002118:Calcium / Q000502:physiology*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors*; D000242:Cyclic AMP / Q000096:biosynthesis*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D007211:Indoles / Q000494:pharmacology; D007545:Isoproterenol / Q000494:pharmacology; D008297:Male; D009638:Norepinephrine / Q000494:pharmacology; D010870:Pineal Gland / Q000166:cytology / Q000378:metabolism*; D011493:Protein Kinase C / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013729:Terpenes / Q000494:pharmacology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019284:Thapsigargin
chemical_list: D004791:Enzyme Inhibitors; D007211:Indoles; D013729:Terpenes; D019284:Thapsigargin; D000242:Cyclic AMP; D011493:Protein Kinase C; D000252:Calcium-Transporting ATPases; D007545:Isoproterenol; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D009638:Norepinephrine; C000543:cyclopiazonic acid