Elevation of cyclic AMP decreases phosphoinositide turnover and inhibits thrombin-induced secretion in human platelets

Elevation of cyclic AMP (cAMP) in platelets inhibits agonist-induced, G protein-mediated responses and activation of polyphosphoinositide-specific phospholipase C (PLC) by ill-defined mechanism(s). Signal transduction steps downstream of PLC are inhibited by elevated cAMP, suggesting an inhibitory effect of cAMP, via protein kinase A, on PLC. In [P-32](i)-prelabeled platelets, forskolin increased intracellular cAMP (104 nmol/10(11) cells at 10(-5) M forskolin) and [P-32]phosphatidylinositol 4-phosphate (Delta[P-32]PIP) (30% at 10(-7)-10(-6) M forskolin). The thrombin-induced (0.1 U/ml) increase in production of [P-32]PA, 'overshoots' in [P-32]PIP and [P-32]PIP2 ([P-32]phosphatidylinositol 4,5-bisphosphate), and the increase in [P-32]PI and secretion of ADP+ATP were abolished by forskolin (10(-7) M). Forskolin stimulated total [P-32]P-i uptake in resting platelets (48%), increased P-32 incorporation into PIP (110%), and inhibited P-32 incorporation into PI (50%). The latter inhibition was most likely considerably greater due to the forskolin-induced stimulation of [P-32]P-i uptake. The changes in radioactive PA, PIP and PIP2 are regarded as being proportional with their masses in the prelabeled platelets, while the increase in PI (phosphatidylinositol) is regarded as a change in specific radioactivity, and hence in its synthesis. The results suggest that cAMP elevation inhibits the flux in the polyphosphoinositide cycle through both inhibition of PIP 5-kinase and PI synthesis. The inverse relation between forskolin-produced Delta PIP and [P-32]PA production suggests that the PLC reaction is inhibited by elevated cAMP through reduction of substrate (PIP2) resynthesis, and not by inhibition of the PLC enzyme. (C) 1998 Elsevier Science B.V. All rights reserved.