Pressure transducer

To measure LVP, a water-filled latex balloon attached to a metal cannula was placed in the left ventricle through the pulmonary vein and connected to a pressure transducer (EMKA Technologies, Paris, France). ECGs were recorded using two surface electrodes (EMKA Technologies, Paris, France) held with a spring against the epicardium. One electrode was placed on the right ventricle near the atrium-ventricle ring and the second on the left ventricle in a position. All hemodynamic parameters were recorded and evaluated using iox2 software (EMKA Technologies, Paris, France) for 10 - 15 min before and after reperfusing KMU-191 with buffer. The QTc in milliseconds was calculated as follows: Bazett: QTcB = QT/RR^1/2; Fridericia: QTcF = QT/RR^1/3. KMU-191 was perfused serially from low concentration (1 μM) to high concentration (10 μM). The procedures used in this study were reviewed and approved by the Institutional Animal Care and Use Committee of the Korea Institute of Toxicology (RS21006).

Rats were placed in a temperature-controlled plethysmograph (body temperature was kept at 37 °C) in which they were ventilated (ventilation rate: 90 beats/min; volume 2 mL/beat) at 270 min after LPS administration. The previously placed tracheostomy was replaced by a small catheter that was connected to a pressure transducer fixed on the plethysmograph box (EMKA Technologies, Paris, France). Transpulmonary pressure (cm H₂O) was determined by measuring pressure differences in the cannula in the trachea. Increasing doses of methacholine (acetyl-β-methyl-choline chloride, Sigma-Aldrich, Zwijndrecht, The Netherlands) (0.37–50 mg/mL, 10% puff for 10 s) were administered by aerosol generated in a nebulizer (EMKA Technologies, Paris, France) connected in between the plethysmograph and the ventilator (EMKA Technologies, Paris, France). After the first dose of methacholine, pulmonary resistance (in cm H₂O/(mL·s⁻¹)) was measured for 3 min. This procedure was repeated for all doses. Subsequently, rats were sacrificed (at 300 min as previously described).