Changes in cytosolic Ca(2+) concentration ([Ca(2+)]c) regulate granule secretion in different cell types. Thrombin activates PAR1 and PAR4 receptors and promotes release of Ca(2+) from distinct intracellular stores, which, in turn, activates store-operated Ca(2+) entry (SOCE). A crucial step during platelet function is the release of physiological agonists stored in secretory granules to the extracellular compartment during activation. We aim to study the role of Ca(2+) mobilization from the extracellular compartment or from different intracellular stores in platelet granule secretion. By using flow cytometry, we have found that α- and δ-granules are secreted in thrombin-stimulated platelets in the absence of extracellular Ca(2+), and in a concentration-dependent manner. Our findings show that thrombin-stimulated granule secretion depends on Ca(2+) mobilization from intracellular stores. Analysis of the kinetics of granule secretion reveals that platelet stimulation with thrombin results in rapid release of α-granules which precedes the secretion of δ-granules. Incubation of platelets with a specific antibody, which recognizes the extracellular amino acid sequence 573-586 of TRPC6, inhibited thrombin-evoked δ-granule exocytosis. Our results indicate that the mechanisms underlying thrombin-induced α- and δ-granule secretion show differences in dependency on Ca(2+) mobilization.
Keywords: Calcium; Platelets; TRPC6; Thrombin; α-granules; δ-granules.
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