Direct inhibitory mechanisms of halothane on human platelet aggregation.
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.
Kohro
S
S
Department of Anesthesiology, Sapporo Medical University School of Medicine, Hokkaido, Japan.
Yamakage
M
M
D000328:Adult; D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D000242:Cyclic AMP / Q000032:analysis; D004305:Dose-Response Relationship, Drug; D006221:Halothane / Q000494:pharmacology*; D006801:Humans; D015544:Inositol 1,4,5-Trisphosphate / Q000032:analysis; D007530:Isoflurane / Q000494:pharmacology; D010974:Platelet Aggregation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000032:analysis; D013917:Thrombin / Q000494:pharmacology
D018685:Anesthetics, Inhalation; D010975:Platelet Aggregation Inhibitors; D019038:Platelet Glycoprotein GPIb-IX Complex; D015544:Inositol 1,4,5-Trisphosphate; D007530:Isoflurane; D000242:Cyclic AMP; D013917:Thrombin; D002118:Calcium; D006221:Halothane