Ischemic preconditioning (PC) induced by 1 cycle of 5-min coronary occlusion and 5-min reperfusion limits infarct size (IS) after 30-min sustained ischemia in rabbits. The shortest ischemic period that induces the PC effect in rabbits is 3 min. To establish the maximum ischemic period to induce a beneficial PC effect, we examined the effect of PC periods of 10 and 15 min on IS after sustained ischemia. The IS in control rabbit hearts after 30 min of sustained occlusion of the left anterolateral coronary artery and 48-h reperfusion was compared with that of hearts treated as follows before being subjected to PC: 5-min occlusion and 5-min reperfusion; 10-min occlusion and 5-min reperfusion; or 15-min occlusion and 5-min reperfusion. In addition, the IS after 15-min or 45-min occlusion and 48-h reperfusion was measured. There was no significant difference in blood pressure, heart rate, or area at risk (AAR) among the rabbits in 5 groups. The IS measured histologically was 40 +/- 4% of AAR in the control, 10 +/- 3% after 5-min PC, and 12 +/- 2% after 10-min PC. However, in the 15-min PC group, the IS was 77 +/- 4% of AAR, which was significantly larger than that of the controls, but similar to that of hearts subjected to 45-min ischemia and reperfusion (67 +/- 3%). As 15 min of preconditioning ischemia alone caused small infarcts (18 +/- 1% of AAR), the infarcts caused by sustained ischemia per se in the 15-min PC group was estimated to be 72 +/- 5% of AAR, which was still significantly higher than in the control groups. We conclude that the maximum period of preconditioning ischemia that induces cardioprotection in rabbits is 10 min. When the ischemic period is longer than this, the IS after sustained ischemia is increased rather than restricted. However, the infarcted size in the 15-min PC group was not higher than that in the group subjected to 45-min continuous ischemia. This may be a major limitation for any clinical application of PC.