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Direct inhibitory mechanisms of halothane on human platelet aggregation
1Department of Anesthesiology, Sapporo Medical University School of Medicine, Hokkaido, Japan.
Background:
Although halothane directly inhibits platelet aggregation, the mechanisms of this effect are still unknown. The current study aimed to clarify the inhibitory mechanisms of halothane on thrombin-induced human platelet aggregation by measuring (1) platelet-surface glycoprotein Ib expression, (2) the concentration of intracellular free Ca2+ ([Ca2+]i) measured simultaneously with aggregation, (3) the concentration of intracellular inositol 1,4,5-triphosphate, and (4) the concentration of intracellular cyclic 3',5'-adenosine monophosphate ([cAMP]i).
Methods:
Washed platelet suspensions, obtained from healthy volunteers, were preincubated with halothane (0-2 mM) for 2 min and then exposed to 0.02 units/ml thrombin for 3 min. The glycoprotein Ib bound to fluorescein-labeled antibody was measured by fluorescence flow cytometry. [Ca2+]i was measured, simultaneously with aggregation, in Fura-2 (Ca2+ indicator)-loaded platelets by use of a fluorometer. Inositol 1,4,5-triphosphate and [cAMP]i were measured by radioimmunoassay.
Results:
Halothane had no effect on glycoprotein Ib expression with or without thrombin. Halothane decreased the thrombin stimulated [Ca2+]i transient and inhibited platelet aggregation in a dose-dependent manner, both in the presence and in the absence of external Ca2+. Isoflurane had no apparent effect on either platelet aggregation or [Ca2+]i in the absence of external Ca2+. Halothane inhibited the increase in inositol 1,4,5-triphosphate induced by thrombin. Halothane moderately but significantly increased [cAMP]i, but the adenylate cyclase activator forskolin (which has the same inhibitory ability on aggregation as halothane) increased [cAMP]i to a much greater extent than did halothane.
Conclusions:
Halothane inhibits thrombin-induced human platelet aggregation by decreasing [Ca2+]i without inhibiting agonist-receptor binding; the inhibitory effect of halothane on [Ca2+]i might be mediated by a decrease in inositol 1,4,5 triphosphate and in part by an increase in [cAMP]i.