title: Synthesis of intracellular histamine in platelets is associated with activation of protein kinase C, but not with mobilization of Ca2+.
abstract: In previous reports, we have provided evidence indicating that newly formed histamine is an intracellular messenger in human platelets. The involvement of protein kinase C (PKC) and intracellular calcium (Ca2+i) in the synthesis of histamine was investigated. Human platelets were stimulated by phorbol 12-myristate 13-acetate (PMA), collagen and the Ca2+ ionophore A23187, with or without the PKC inhibitor staurosporine. Aggregation, histamine synthesis and phosphorylation of pleckstrin (47 kDa; P47) and myosin light chain (20 kDa; P20) proteins were monitored. Staurosporine inhibited PMA- and collagen-induced aggregation, histamine synthesis and phosphorylation of 47 kDa and 20 kDa proteins in a dose-dependent manner. For PMA, median inhibitory concentrations (IC50 values) for staurosporine inhibition of aggregation, histamine synthesis and phosphorylation were similar, suggesting that histamine synthesis induced by this agonist may be a consequence of PKC activation. Conversely, collagen-stimulated histamine synthesis was inhibited by staurosporine at concentrations significantly higher than those required to inhibit aggregation (P less than 0.005) or pleckstrin phosphorylation (P less than 0.01), indicating the possible involvement of non-PKC mechanism(s) in the synthesis of histamine induced by this agonist. A23187 failed to induce the synthesis of intracellular histamine in platelets, whereas staurosporine blocked A23187-induced aggregation and phosphorylation of the 20 kDa protein at significantly higher concentrations than those needed to inhibit PKC. When platelets were stimulated with a combination of A23187 and PMA, the increase in platelet histamine was less than that with PMA alone. The results provide evidence that the synthesis of intracellular histamine in platelets occurs as a consequence of PKC activation and may be down-regulated under conditions where there is a substantial rise in [Ca2+]i.
authors.affiliation: Manitoba Institute of Cell Biology, Winnipeg, Canada.
authors.forename: S P
authors.initials: SP
authors.lastname: Saxena
authors.forename: C
authors.initials: C
authors.lastname: Robertson
authors.forename: A B
authors.initials: AB
authors.lastname: Becker
authors.forename: J M
authors.initials: JM
authors.lastname: Gerrard
mesh_terms: D000244:Adenosine Diphosphate / Q000037:antagonists & inhibitors; D000470:Alkaloids / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D001798:Blood Proteins / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D003094:Collagen / Q000494:pharmacology*; D016861:Cyclooxygenase Inhibitors; D004789:Enzyme Activation / Q000187:drug effects; D006632:Histamine / Q000096:biosynthesis*; D006801:Humans; D010750:Phosphoproteins; D010766:Phosphorylation; D010974:Platelet Aggregation; D011493:Protein Kinase C / Q000378:metabolism*; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
chemical_list: D000470:Alkaloids; D001798:Blood Proteins; D016861:Cyclooxygenase Inhibitors; D010750:Phosphoproteins; C039077:platelet protein P47; D000001:Calcimycin; D000244:Adenosine Diphosphate; D006632:Histamine; D003094:Collagen; D011493:Protein Kinase C; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium