Rabbit aortae were removed from exsanguinated rabbits, washed, everted on probes, treated with thrombin, washed to remove unbound thrombin and used to measure the accumulation of 51Cr-labeled platelets in vitro. Thrombin pretreatment of normal rabbit aortae did not cause platelet accumulation on the endothelium; platelets appeared to accumulate only at sites where the subendothelium had been exposed. The quantitative data obtained with 51Cr-labelled platelets was reinforced by observations by scanning electron microscopy. 125I-labelled thrombin became associated with the endothelium and also with de- endothelialized vessels, and some of it could be displaced by high concentrations of heparin. Exposure of vessels to heparin after thrombin treatment eliminated the enhanced platelet accumulation caused by the thrombin treatment, probably because heparin displaced thrombin from the aortae, as demonstrated in experiments with 125I-thrombin. Inhibition of PGI2 production by aspirin treatment of the vessels did not enhance platelet accumulation on normal or thrombin-treated aortae. Thus, although thrombin treatment of the endothelium does not cause platelets to adhere to it, thrombin does cause increased platelet accumulation on the areas where the subendothelium is exposed or where endothelial cells are damaged.