Synergy between Ca2+ and protein kinase C is the major factor in determining the level of secretion from human platelets.
The aim of this study was to establish further the role of protein kinase C in aggregation and secretion of 5-hydroxytryptamine (5-HT) from human platelets by using the selective inhibitor Ro 31-8220. Ro 31-8220 (3 microM) inhibited completely phosphorylation of pleckstrin, the major protein kinase C substrate, induced by thrombin, A23187 or phorbol dibutyrate (PDBu). Myosin light-chain phosphorylation induced by PDBu was also inhibited completely, but that induced by thrombin or A23187 was only inhibited partially. As myosin light chain is a substrate for both myosin light-chain kinase and protein kinase C, these results suggest that Ro 31-8220 is inhibiting only the protein kinase C-induced phosphorylation and that Ro 31-8220 has a greater selectivity to protein kinase C than does its structural analogue staurosporine. The stimulation of secretion of 5-HT by maximally effective concentrations of thrombin and A23187 was decreased significantly by 3 microM Ro 31-8220, but not inhibited completely. These results indicate a major role for protein kinase C in the stimulation of secretion by agonist- and ionophore-induced activation. On its own, a maximal concentration of PDBu induced a small degree of secretion (3.3 +/- 1.0%), but potentiated markedly the response to a submaximal concentration of A23187 (300 nM) to a level greater than seen with a maximal concentration of A23187. A similar set of results was also seen with aggregation, but not with shape change. We interpret these results to mean that the signalling event for secretion and aggregation is Ca2+, and this is potentiated markedly by protein kinase C. In the case of secretion, it appears that it is the synergy which is the major determining factor in influencing the extent.
Walker
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TR
Department of Pharmacology, University of Oxford, U.K.
Watson
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SP
D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D001798:Blood Proteins / Q000378:metabolism; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D009218:Myosins / Q000378:metabolism; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010750:Phosphoproteins; D010766:Phosphorylation / Q000187:drug effects; D015539:Platelet Activation; D010974:Platelet Aggregation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012701:Serotonin / Q000378:metabolism; D013997:Time Factors
D001798:Blood Proteins; D007211:Indoles; D010750:Phosphoproteins; C039077:platelet protein P47; D012701:Serotonin; D015240:Phorbol 12,13-Dibutyrate; D000001:Calcimycin; D011493:Protein Kinase C; D009218:Myosins; D002118:Calcium; C064758:Ro 31-8220