Salidroside (Sal), a natural extract of Rhodiola rosea, shows a latent effect on protecting cardiovascular system. Our study explored the effect of salidroside on ischemia-reperfusion (I/R) injury in rat heart. I/R was performed on Wistar rat hearts, and Sal pretreatment was performed in I/R rats. Cardiac marker enzyme, myocardial infarct size, malondialdehyde (MDA) and superoxide dismutase (SOD) content were then measured. Compared with the untreated group, Sal pretreatment observably ameliorated the cardiac function, decreased the myocardial infarct size, reduced the levels of cardiac lactate creatine kinase-MB (CK-MB) and dehydrogenase (LDH), and inhibited the anti-oxidative stress. In addition, Sal treatment also significantly inhibited autophagy and apoptosis, which could be partially reversed by Rapamycin (RAPA), an autophagic agonist. Furthermore, Sal treatment attenuated autophagy by up-regulating the expression of hsa_circ_0000064 (circ-0000064) and Rapamycin (RAPA) treatment abolished it. Our study showed that Sal protected the heart from I/R injury, which might be related to the upregulation of circ-0000064 and the inhibition of autophagy.