SELECTIVE ANTAGONISTS OF PAF: INTERFERENCE WITH PLATELET FUNCTION AND EFFECT ON THROMBOGENESIS INDUCED BY SUBENDOTHELIUM.
Platelet activating factor (PAF) is a very potent excitatory agonist of blood platelets but the physiological importance of this mediator in platelet thrombus formation is not known. We investigated the effect of two chemically unrelated selective inhibitors of PAF-induced platelet aggregation on thrombogenesis induced by rabbit aorta subendothelium (SE) using an ex vivo perfusion system.Ro 19-3704 is a highly potent inhibitor structurally related to PAF. This compound inhibits PAF-induced aggregation of rabbit platelets in platelet rich plasma in vitro competitively. Against 4 nM PAF, a concentration resulting in submaximal platelet aggre-gregation velocity, the IC50 was 70 nM. Inhibition was highly selective for PAF-induced aggregation, since aggregation induced by collagen (HORM, 5 yg/ml), ADP (1 yM) or thrombin (0.4 U/ml) was not inhibited even at a concentration as high as 10 yM. Bro-tizolam, a triazolobenzodiazepine reported to be a selective inhibitor of PAF-induced platelet activation, had in our system an IC50 of 200 nM. The selective benzodiazepine antagonist Ro 151788 was without effect on inhibition of PAF-induced platelet activation by brotizolam.Ro 19-3704 was given intravenously to rabbits as a bolus of 0.2 mg/kg followed by constant infusion of 0.02 mg/kg/min. This dosage provoked ex vivo a constant right shift ratio of the dose response curve for PAF-induced aggregation (RSR[PAF]) by a factor of 25 to 35. Brotizolam was given orally at a dose of 100 mg/ kg together with 300 mg/kg of Ro 15-1788 (to antagonize the central effects) 90 minutes before starting the perfusion experiment, resulting in a RSR[PAF] of 35 to 135. ADP induced platelet aggregation was not impaired by either compound. SE was exposed to the non-anticoagulated blood withdrawn from the carotid artery for 3 min at 2600 s-1 and for 20 min at 200 s-1 shear rate. Quantitative morphometric evaluation showed that SE coverage by platelets and by fibrin, thrombus area and thrombus height were all unchanged by the PAF antagonists at low and at high shear rates despite a very substantial inhibition of PAF-induced platelet aggregation. Therefore a major role of PAF in SE-induced thrombogenesis seems unlikely.