The phorbol ester TPA potentiates cholera toxin- and isoproterenol-stimulated cyclic AMP-synthesis in primary astrocyte cultures.
Cellular responses to changes in the extracellular environment are mediated by intracellular signaling systems. One of the most extensively studied systems is adenylate cyclase which generates the second messenger molecule cAMP. Another one is the phosphatidylinositol (PI) second messenger system giving rise to IP3 and diacylglycerol, the latter stimulating protein kinase C. Recently, a third potential signaling system has attracted increased scientific attention: the phospholipase A2 system which generates arachidonic acid. This substance may be used for eicosanoid synthesis or serve as a second messenger molecule. The present report gives more evidence about mechanisms how these signaling pathways interact in cultured astrocytes. Substances commonly used for stimulation of arachidonic acid release and prostaglandin synthesis in these cultures (A23187, TPA) had no influence on intracellular cAMP levels. Pertussis toxin that had previously been shown to inhibit prostaglandin synthesis, had no influence on cAMP levels either. Cholera toxin, however, raised intracellular cAMP significantly, although much less than the beta-adrenoceptor agonist isoproterenol. Cholera toxin also caused a marked change in astroglial morphology even at reduced concentrations (1-10 ng/ml). A23187 used in combination with Ctx had a moderate stimulatory effect on cAMP synthesis. In contrast, in the presence of Ctx, the PKC-activating phorbol ester TPA synergistically stimulated cAMP production, raising cAMP levels as high as isoproterenol-stimulated levels. The TPA effect was concentration-dependent. It was also dependent on an intact PKC since preincubation of cells with the phorbol ester completely abolished the synergistic effect. The synergistic effect of the phorbol ester was also observed at subthreshold concentrations of isoproterenol. The data reveal that the sole activation of most Gs molecules is a necessary but not sufficient prerequisite to achieve maximal adenylate cyclase activity. The fine-tuning of this activity apparently occurs at the catalytic subunit which is under the (partial) control of phosphorylation by PKC.
Gebicke-Haerter
P J
PJ
Institute of Pharmacology, University of Freiburg, Germany.
Seregi
A
A
Schobert
A
A
Hertting
G
G
D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D001253:Astrocytes / Q000187:drug effects / Q000378:metabolism*; D000001:Calcimycin / Q000494:pharmacology; D002478:Cells, Cultured; D002772:Cholera Toxin / Q000494:pharmacology*; D000242:Cyclic AMP / Q000096:biosynthesis*; D004357:Drug Synergism; D007545:Isoproterenol / Q000494:pharmacology*; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D000318:Adrenergic beta-Agonists; D000001:Calcimycin; D002772:Cholera Toxin; D000242:Cyclic AMP; D011493:Protein Kinase C; D007545:Isoproterenol; D013755:Tetradecanoylphorbol Acetate