Epinephrine can in certain in vitro conditions induce the aggregation of human platelets and could play an important role in vivo in the appearance of thrombotic disorders when catecholamine levels are increased. This study examines some functional and biochemical responses to epinephrine. Epinephrine induces the aggregation and serotonin secretion of human platelets in citrated plasma. This is not due to a direct effect of citrate itself, such as the lowering of plasma free Ca2+ but more likely to the generation of traces of thrombin during blood collection, as suggested by abrogation of these platelet responses when hirudin was added before citrate. When washed human platelets suspended in Tyrode buffer containing 2 mM Ca2+, 0.35% albumin and apyrase, and 0.1-100 microM epinephrine were used, no shape change, aggregation, or secretion of serotonin was observed, nor was the platelet ultrastructure modified. Epinephrine does not modify platelet membrane fluidity, as studied with the lipophilic fluorescent probe trimethylammonium-diphenylhexatriene. It has no direct effect on fibrinogen binding to intact platelets, intracellular Ca2+ levels measured by quin2, or protein phosphorylation. Epinephrine potentiates the action of all types of aggregating agents on aggregation, secretion, intracellular Ca2+ levels, membrane fluidity, fibrinogen binding, or protein phosphorylation. These effects are mediated by alpha 2-adrenergic agonists and inhibited by alpha 2-adrenergic antagonists. This study shows that epinephrine alone does not induce modifications of morphology, metabolism, or function of intact and functional washed human platelets and that it cannot be considered per se as an aggregating agent. However, epinephrine interacts with alpha 2-adrenergic receptors on human platelets and potentiates biochemical and aggregatory responses induced by other platelet agonists.

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SummaryIncorporation into human platelets of the calcium fluorescent indicators quin2 or fura-2 at low concentrations used to measure intracellular free calcium leads to the potentiation of the effects of agonists on platelets. This was shown by increased aggregatory and secretory responses of quin2 or fura-2 loaded platelets after stimulation with ADP, PAP and with low concentrations of thrombin, collagen, the endoperoxide analog U-46619 and the calcium ionophore A 23187. Quin2 and fura-2 mediated platelet sensitisation could be due to altered arachidonic acid metabolism since it was inhibited by prior treatment with the cydooxygenase inhibitor acetylsalicylate. In contrast, platelets loaded with higher concentrations of calcium chelators exhibited diminished aggregation responses to all aggregating agents. This latter effect was accompanied by increased fluidity of the platelet plasma membrane bilayer and by the exposure of a new pool of membranes to the outer surface of platelets, as monitored with trimethylammonium- diphenylhexatriene (TMA-DPH) in platelets loaded with the non-fluorescent calcium probe analog MAPT. In contrast, low concentrations of quin2 did not potentiate shape change of platelets activated with ADP. Thus, shape change and aggregation can be influenced separately by intracellular Ca2+ chelators. We conclude that platelet responses are altered by the incorporation of intracellular calcium chelators at concentrations used to monitor intracellular calcium changes.

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