Competitive antagonism at thromboxane receptors in human platelets

The inhibitory effects of three prostanoid analogues, EP 045, EP 092 and pinane thromboxane A2 (PTA2), on the aggregation of human platelets in vitro have been investigated. In diluted platelet-rich plasma (PRP), EP 045 (20 microM) and EP 092 (1 microM) completely inhibited irreversible aggregation responses to thromboxane A2 (TXA2), prostaglandin H2 (PGH2) and five chemically stable thromboxane mimetics, including 11,9-epoxymethano-PGH2 and 9,11-azo-PGH2. Reversible aggregation produced by the prostanoid analogue, CTA2, was also inhibited. The block of the stable agonist action was surmountable. In plasma-free platelet suspensions EP 045 and EP 092 were more potent antagonists. Schild analysis indicated a competitive type of antagonism for EP 045 (affinity constant of 1.1 X 10(7) M-1); the nature of the EP 092 block is not clear. Primary aggregation waves induced by ADP, platelet activating factor (Paf) and adrenaline were unaffected by EP 045 and EP 092, whereas the corresponding second phases of aggregation were suppressed. Aggregation and 5-hydroxytryptamine (5-HT) release induced by either PGH2 or 11,9-epoxymethano-PGH2 were inhibited in a parallel manner by EP 045. Inhibition of thromboxane biosynthesis is not involved in these effects. EP 045 and EP 092 did not raise adenosine 3':5'-cyclic monophosphate (cyclic AMP) levels in the platelet suspensions. In plasma-free platelet suspensions PTA2 produced a shape change response which could be blocked by EP 045. PTA2, therefore, has a thromboxane-like agonist action. The block of the aggregatory action of 11,9-epoxymethano-PGH2 by PTA2 appears to be mainly due to competition at the thromboxane receptor. However, PTA2 produced a slight rise in cyclic AMP levels; this could be due to a very weak stimulant action on either PGI2 or PGD2 receptors present in the human platelet. Functional antagonism by PTA2 may therefore augment its thromboxane receptor blocking activity. The results are discussed in terms of (a) the specificity of antagonism produced by EP 045, EP 092 and PTA2, (b) the validity of affinity constant determinations for receptor antagonists when aggregation is the biological response, and (c) the characteristics of the human platelet thromboxane receptor in comparison with those of thromboxane receptors in smooth muscle.