Hypercholesterolemia (mean plasma cholesterol: 15 mM) was induced in rabbits by the feeding of a chow diet enriched with a low amount (0.25%, w/w) of cholesterol only. Platelet size and protein content decreased significantly, but the whole blood platelet count did not change. The platelets became enriched in cholesterol, as indicated by a significant increase in the choiesterol:phospholipid molar ( C P ) ratio. Specific responses of washed platelets stimulated with various agonists were studied to determine the effects of hypercholesterolemia on the various pathways of platelet aggregation in the absence of plasma components. In platelets from hypercholesterolemic rabbits compared with controls: aggregation induced by ADP was not altered; collagen-induced responses (aggregation, secretion of [ 14C]serotonin from prelabelled platelets, thromboxane A 2 (TXA 2) formation, mobilization of [ 3H]arachidonate from prelabelled platelets) were enhanced; with aspirin-treated platelets, aggregation induced by the TXA 2 mimetic U46619 was enhanced; and thrombin-induced responses of both untreated platelets (aggregation, secretion of granule contents, TXA 2 formation) and aspirin-treated platelets (aggregation) were enhanced. Thus, platelets from cholesterol-fed rabbits not only form more TXA 2, but they aggregate more extensively when stimulated by its mimetic. In addition, it has not been previously recognized that these platelets are also hypersensitive to thrombin-induced aggregation that is independent of TXA 2.