To examine the effects of ischemic preconditioning on ischemia-reperfusion-induced changes in the sarcoplasmic reticulum (SR) function, isolated rat hearts were either perfused with a control medium for 30 min or preconditioned with three episodes of 5-min ischemia and 5-min reperfusion before sustained ischemia for 30 min followed by reperfusion for 30 min was induced. Preconditioning itself depressed cardiac function (left ventricular developed pressure, peak rate of contraction, and peak rate of relaxation) and SR Ca2+-release and -uptake activities as well as protein content and Ca2+/calmodulin-dependent protein kinase (CaMK) phosphorylation of Ca2+-release channels by 25-60%. Global ischemia for 30 min produced marked depressions in SR Ca2+-release and -uptake activities as well as SR Ca2+-pump protein content in control hearts; these changes were significantly attenuated by preconditioning. Compared with the control preparations, preconditioning improved the recovery of cardiac function and SR Ca2+-release and -uptake activities as well as Ca2+-release channel and Ca2+-pump protein contents in the ischemic-reperfused hearts. Unlike the protein kinase A-mediated phosphorylation in SR membranes, the CaMK-mediated phosphorylations at Ca2+-release channels, Ca2+ pump, and phospholamban were depressed in the ischemic hearts; these changes were prevented by preconditioning. These results indicate that ischemic preconditioning may exert beneficial effects on ischemia-reperfusion-induced alterations in SR function by preventing changes in Ca2+-release channel and Ca2+-pump protein contents in the SR membrane.