Anesthetics, especially propofol, are discussed to influence ischemic preconditioning. We investigated whether cardioprotection by milrinone or levosimendan is influenced by the clinically used anesthetics propofol, sevoflurane or dexmedetomidine. Hearts of male Wistar rats were randomised, placed on a Langendorff system and perfused with Krebs–Henseleit buffer (KHB) at a constant pressure of 80 mmHg. All hearts underwent 33 min of global ischemia and 60 min of reperfusion. Three different anesthetic regimens were conducted throughout the experiments: propofol (11 μM), sevoflurane (2.5 Vol%) and dexmedetomidine (1.5 nM). Under each anesthetic regimen, pharmacological preconditioning was induced by administration of milrinone (1 μM) or levosimendan (0.3 μM) 10 min before ischemia. Infarct size was determined by TTC staining. Infarct sizes in control groups were comparable (KHB-Con: 53 ± 9%, Prop-Con: 56 ± 9%, Sevo-Con: 56 ± 8%, Dex-Con: 53 ± 9%; ns). Propofol completely abolished preconditioning by milrinone and levosimendan (Prop-Mil: 52 ± 8%, Prop-Lev: 52 ± 8%; ns versus Prop-Con), while sevoflurane did not (Sevo-Mil: 31 ± 9%, Sevo-Lev: 33 ± 7%; p < 0.05 versus Sevo-Con). Under dexmedetomidine, results were inconsistent; levosimendan induced infarct size reduction (Dex-Lev: 36 ± 6%; p < 0.05 versus Dex-Con) but not milrinone (Dex-Mil: 51 ± 8%; ns versus Dex-Con). The choice of the anesthetic regimen has an impact on infarct size reduction by pharmacological preconditioning.