Off-pump coronary artery bypass grafting (CABG) has become a popular procedure. However, temporary occlusion of the target vessel is sometimes a threat to the patients. Although ischemic preconditioning (IP) has been proposed to reduce myocardial injury, its effects remain controversial. The coronary veins represent an alternate route for delivery of therapeutic agents and arterial blood to the acutely ischemic myocardium. The aim of this study was to investigate the protective effect against myocardial ischemia and reperfusion injury of combined IP and synchronized coronary venous retroperfusion (SCVR) in an off-pump CABG model. Twenty-one pigs were assigned to 3 groups of 7 animals. In the control group, the left anterior descending coronary artery (LAD) was occluded for 45 min followed by 2 h of reperfusion using a left intrathoracic artery (LITA) bypass circuit. In the IP group, LAD occlusion was done for 5 min with 15 min of reperfusion, followed by 45 min of LAD occlusion. In the SCVR group, pretreatment before LAD occlusion was the same as in the IP group. Then, SCVR was commenced just after the start of LAD occlusion for 45 min. The percent systolic shortening of ischemic myocardium (measured by sonomicrometry) after reperfusion via the LITA was significantly (p < 0.001) greater in the SCVR group (14.6 +/- 3.3%) than in the control group (-1.6 +/- 5.6%, 95%CI: -24.3 - -8.1) or the IP group (0.7 +/- 8.0%, 95%CI: -22.0 - -5.8) after 30 min of reperfusion, and this difference persisted throughout the reperfusion period. Infarct size (expressed as a percentage of the area at risk) was significantly (p < 0.001) smaller in the SCVR group (2.4 +/- 2.7%) than in the control group (83.0 +/- 2.3%, 95%CI: -99.0 - -62.4) or the IP group (42.0 +/- 23.0%, 95%CI: -58.0 - -21.3). Combined SCVR and IP had a potent myocardial protective effect in the present off-pump CABG model. This method may be clinically feasible and may be able to prolong a safe coronary occlusion.