The combination of various interventions to obtain enhanced cardioprotection is always an important area of research focus. This randomized experiment was designed to assess whether combined fentanyl and limb remote ischemic postconditioning produced enhanced protection against myocardial ischemia/reperfusion injury in an in vivo rat model, and to determine if kappa-opioid receptors were implicated in the cardioprotection of these interventions. Seventy-two rats were exposed to a 30-min myocardial ischemia followed by a 180-min reperfusion. Half of the rats (36) were randomized into four different groups receiving control treatment, fentanyl postconditioning, limb remote ischemic postconditioning, and combined fentanyl and limb remote ischemic postconditioning. The remaining 36 rats were also randomized into four groups receiving the same interventions as the above groups following the intravenous administration of a kappa-opioid receptor antagonist, nor-binaltorphimine, before myocardial ischemia. At the end of reperfusion, both serum cardiac troponin I and infarct size were determined. Both fentanyl postconditioning and limb remote ischemic postconditioning significantly decreased the infarct size and serum cardiac troponin I level, and combined fentanyl and limb remote ischemic postconditioning produced enhanced cardioprotection on the infarct size-sparing effect. The use of nor-binaltorphimin to block kappa-opioid receptors eliminated cardioprotection by fentanyl postconditioning and enhanced cardioprotection by combined fentanyl and limb remote ischemic postconditioning, but did not change cardioprotection by limb remote ischemic postconditioning. Combined fentanyl and limb remote ischemic postconditioning produced enhanced protection against myocardial ischemia/reperfusion injury. kappa-Opioid receptors are essential for cardioprotection by fentanyl postconditioning and enhanced cardioprotection by combined fentanyl and limb remote ischemic postconditioning; however, they do not play a pivotal role in cardioprotection by limb remote ischemic postconditioning.