Ketamine suppresses platelet aggregation possibly by suppressed inositol triphosphate formation and subsequent suppression of cytosolic calcium increase

Background: Ketamine has been shown to suppress platelet aggregation, but its mechanisms of action have not been defined. The purpose of (lie current study is to clarify the effects of ketamine on human platelet aggregation and to elucidate the underlying mechanisms of its action. Methods: Platelet aggregation was measured using an eight-channel aggregometer, and cytosolic free calcium concentration was measured in Fura-2/Am-loaded platelets using a fluorometer. Inositol 1,4,5-triphosphate (IP3) was measured with use of a commercially available IP3 assay kit. To estimate thromboxane A(2) (TXA(2)) receptor binding affinity and expression, Scatchard analysis was performed using [H-3]S145, a specific TXA(2) receptor antagonist. TX2 agonist binding assay was also performed. The membrane-bound guanosine 5'-triphosphatase activity was determined using [gamma-P-32]guanosine triphosphate by liquid scintillation analyzer. Results: Ketamine (500 mum) suppressed aggregation induced by adenosine diphosphate (0.5 mum), epinephrine (1 mum), (+)-9,11-epithia-11,12-methano-TXA(2) (STA(2)) (0-5 mum), and thrombin (0.02 U/ml) to 39.1 +/- 30.9, 46.3 +/- 4.3, -2.0 +/- 16.8, and 86.6 +/- 1.4% of zero-control, respectively. Ketamine (250 mum-1 mM) also suppressed thrombin- and STA(2)-induced cytosolic free calcium concentration increase dose dependently. Although ketamine (2 mM) had no effect on TXA(2) receptor expression and its binding affinity, it (1 mM) suppressed intracellular peak IP3 concentrations induced by thrombin and STA(2) from 6.60 +/- 1.82 and 4.39 +/- 2.41 to 2.41 +/- 0.98 and 1.90 +/- 0.86 pmol/10(9) platelets, respectively, and it suppressed guanosine triphosphate hydrolysis induced by thrombin (0.02 units/ml.) and STA., (0-5 muM) to 50.3 +/-3.2 and 67.5 +/- 5-54% versus zero-control, respectively. Conclusion. Ketamine inhibits human platelet aggregation possibly by suppressed IP3 formation and subsequent suppression of cytosolic free calcium concentration. The site of action of ketamine is neither TXA(2) nor thrombin binding sites but possibly receptor-coupled mechanisms, including G-protein.