Background— We tested the hypothesis that ischemia-induced ventricular fibrillation (VF) is facilitated by platelets, trapped regionally in the ischemic zone and activated to release arrhythmogenic secretome. Methods and Results— In a randomized study in blood-free, buffer-perfused isolated rat hearts, ischemic zone territory (34±1% of left ventricle) was selected so that ischemia evoked VF in only 42% of controls. VF incidence was increased to 91% ( P <0.05) by coronary ligation–induced trapping of freshly prepared autologous platelets (infused before and during coronary ligation, with trapping confirmed by ¹¹¹ In-labeled platelet autoradiographic imaging). Trapping of platelet secretome prepared ex vivo, or platelet-sized fluorospheres, did not increase ischemia-induced VF incidence. Secretome alone did, however, evoke VF in 2 sham coronary-ligated hearts. Perfusion did not activate infused platelets in sham coronary-ligated hearts, whereas ligation activated trapped platelets (assessed by thromboxane release). In a separate study, trapping whole-heparinized blood mimicked the ability of trapped platelets to increase VF incidence. This effect was not prevented by >5 days oral pretreatment in vivo with clopidogrel (10 mg/kg per day) or indomethacin (2.4 mg/kg per day). Conclusions— Platelets facilitate VF during acute ischemia independently of their ability to participate in occlusive thrombosis. Moreover, the effect is unresponsive to antiplatelet drugs commonly used. Labile secretome constituents appear to be responsible. This opens a novel avenue for antiarrhythmic drug research.