ABSTRACTPurpose:Remote ischemic preconditioning (RIPC) confers cardioprotection against ischemia reperfusion (IR) injury. However, the precise mechanisms involved in RIPC-induced cardioprotection are not fully explored. The present study was aimed to identify the role of melatonin in RIPC-induced late cardioprotective effects in rats and to explore the role of H2S, TNF-α and mitoKATPin melatonin-mediated effects in RIPC.Methods:Wistar rats were subjected to RIPC in which hind limb was subjected to four alternate cycles of ischemia and reperfusion of 5 min duration by using a neonatal blood pressure cuff. After 24 h of RIPC or ramelteon-induced pharmacological preconditioning, hearts were isolated and subjected to IR injury on the Langendorff apparatus.Results:RIPC and ramelteon preconditioning protected the hearts from IR injury and it was assessed by a decrease in LDH-1, cTnT and increase in left ventricular developed pressure (LVDP). RIPC increased the melatonin levels (in plasma), H2S (in heart) and decreased TNF-α levels. The effects of RIPC were abolished in the presence of melatonin receptor blocker (luzindole), ganglionic blocker (hexamethonium) and mitochondrial KATPblocker (5-hydroxydecanoic acid).Conclusions:RIPC produce delayed cardioprotection against IR injury through the activation of neuronal pathway, which may increase the plasma melatonin levels to activate the cardioprotective signaling pathway involving the opening of mitochondrial KATPchannels, decrease in TNF-α production and increase in H2S levels. Ramelteon-induced pharmacological preconditioning may also activate the cardioprotective signaling pathway involving the opening of mitochondrial KATPchannels, decrease in TNF-α production and increase in H2S levels.