Ex vivo Langendorff-perfused rat heart

Adult male Wistar rats weighing 250–350 g were heparinized (4 IU/g i.p.) and anaesthetized by intraperitoneal administration of an overdose of sodium thiopental (200 mg/Kg). The hearts were quickly removed, mounted on the aortic cannula of the Langendorff perfusion system apparatus and perfused with an oxygenated Krebs- Henseleit buffer (en mM; 118 NaCl, 4.7 KCl, 1.25 CaCl₂, 1.2 KH₂PO₄, 1.2 MgSO₄, 25 NaHCO₃, and 5 glucose) as described previously [9, 17]. Before each experimental protocol was initiated, the isolated hearts were set at a mean arterial pressure of 60–80 mmHg and were allowed to stabilize at 37°C for 40 to 60 minutes. Chart Powerlab software (ADInstruments) was used for continuous recording throughout the experiments of heart rate, left ventricular developed pressure (LVDP), and maximum positive and negative derivative of left ventricular pressure (±dP/dt). The heart contractility under different treatments was evaluated by the analysis of +dP/dt, which corresponds to % increase of +dP/dt normalized to basal value after the period of stabilization.

The standard protocol of ischemia/reperfusion in perfused hearts was followed as described previously [16]. Group 1 of I/R: After stabilization, rat hearts were exposed to global ischemia (without aorta perfusion) during 40 minutes and 1 hour of reperfusion with freshly oxygenated solution. Group 2 corresponds to the pharmacological preconditioning with Ucn-1 (10 nM). After stabilization, Ucn-1 was applied 20 minutes before ischemia and 30 minutes at the beginning of reperfusion.