{"title":"Elevation of cyclic AMP decreases phosphoinositide turnover and inhibits thrombin-induced secretion in human platelets","authors":"Anita Ryningen, Baard Olav Jensen, Holm Holmsen","doi":"10.1016/S0005-2760(98)00106-4","DOIUrl":null,"url":null,"abstract":"<div><p>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 [<sup>32</sup>P]<sub>i</sub>-prelabeled platelets, forskolin increased intracellular cAMP (104 nmol/10<sup>11</sup> cells at 10<sup>−5</sup> M forskolin) and [<sup>32</sup>P]phosphatidylinositol 4-phosphate (Δ[<sup>32</sup>P]PIP) (30% at 10<sup>−7</sup>–10<sup>−6</sup> M forskolin). The thrombin-induced (0.1 U/ml) increase in production of [<sup>32</sup>P]PA, ‘overshoots’ in [<sup>32</sup>P]PIP and [<sup>32</sup>P]PIP<sub>2</sub> ([<sup>32</sup>P]phosphatidylinositol 4,5-bisphosphate), and the increase in [<sup>32</sup>P]PI and secretion of ADP+ATP were abolished by forskolin (10<sup>−7</sup> M). Forskolin stimulated total [<sup>32</sup>P]P<sub>i</sub> uptake in resting platelets (48%), increased <sup>32</sup>P incorporation into PIP (110%), and inhibited <sup>32</sup>P incorporation into PI (50%). The latter inhibition was most likely considerably greater due to the forskolin-induced stimulation of [<sup>32</sup>P]P<sub>i</sub> uptake. The changes in radioactive PA, PIP and PIP<sub>2</sub> 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 ΔPIP and [<sup>32</sup>P]PA production suggests that the PLC reaction is inhibited by elevated cAMP through reduction of substrate (PIP<sub>2</sub>) resynthesis, and not by inhibition of the PLC enzyme.</p></div>","PeriodicalId":100162,"journal":{"name":"Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"1998-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/S0005-2760(98)00106-4","citationCount":"35","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0005276098001064","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
引用次数: 35
Abstract
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 [32P]i-prelabeled platelets, forskolin increased intracellular cAMP (104 nmol/1011 cells at 10−5 M forskolin) and [32P]phosphatidylinositol 4-phosphate (Δ[32P]PIP) (30% at 10−7–10−6 M forskolin). The thrombin-induced (0.1 U/ml) increase in production of [32P]PA, ‘overshoots’ in [32P]PIP and [32P]PIP2 ([32P]phosphatidylinositol 4,5-bisphosphate), and the increase in [32P]PI and secretion of ADP+ATP were abolished by forskolin (10−7 M). Forskolin stimulated total [32P]Pi uptake in resting platelets (48%), increased 32P incorporation into PIP (110%), and inhibited 32P incorporation into PI (50%). The latter inhibition was most likely considerably greater due to the forskolin-induced stimulation of [32P]Pi 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 ΔPIP and [32P]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.