Ml 870

Rats were sacrificed and the chest was opened to obtain the trachea. The trachea was dissected and excess connective tissue and fat were removed. Then, the trachea was cut into rings of 3–4 mm width each having about four cartilages for the formation of tracheal ring. Each tracheal ring was hung between two Nichrome hooks inserted into the lumen, and placed in a 10 mL organ bath containing Krebs-Henseliet solution (KHS; composition (mM): NaCl 120, KCl 4.72, KH₂PO₄ 1.2, MgSO₄·7H₂O 0.5, CaCl₂·2H₂O 2.5, NaHCO₃ 25 and dextrose 11). This solution was maintained at 37 ± 0.5 °C and constantly bubbled with 5% CO₂-95% O₂. Tissue was suspended under isotonic tension of 1 g and allowed to equilibrate for at least 1 h while it was being washed with KHS solution every 15 min. In all experiments, contraction responses were measured using an isotonic transducer (MLT0202, AD Instruments, Australia) which was connected to a power lab system (PowerLab 8/30, ML870, AD Instruments, Australia).

To determine electrical effect of exogenous L5, 8-week-old C57B6/J mice (wild-type mice) were injected with 5 mg/kg of L1 or L5 (n = 5 per group) daily through the tail vein for 1 week. LOX-1 knockout (LOX-1^−/−) mice (n = 5) from the laboratory of Dr. Tatsuya Sawamura were also injected with L5 daily for 1 week to determine the role of LOX-1. All animal research was approved by the Mackay Medical College Institutional Animal Care and Use Committee (A1020016), and all procedures were conducted in accordance with the Guide for the Care and Use of Laboratory Animals by the US National Institutes of Health. After 1 week, the lead II surface electrocardiogram was recorded in anesthetized animals at the sampling rate of 1000 Hz by using PONEMAH real-time acquisition interface P3P Plus coupled to a digital converter (ML-870, ADInstruments, Colorado Springs, CO, USA). Data were analyzed by the software Lab Chart 7 plus (ADInstruments), and all paramters were measured as the average of five consecutive cycles. The rate-corrected QT interval was calculated according to the Mitchell’s approach verified previously in rodents: $$\ln \left(\mathrm{QT}0\right)=\ln \left(\mathrm{QTc}\right)+\mathrm{y}\ln \left(\mathrm{RR}100\right)$$ where QT0, QTc, y, RR100 are the observed QT, rate-corrected QT interval, value of the exponent, and normalized RR interval, respectively^{53 (link)}.

Rats were anesthetized with intraperitoneal injection of thiopental sodium (60–80 mg/kg ip). Right carotid artery was cut down to insert the microtip 2.0 F Pressure-Volume (PV) catheter (SPR-838, Millar Instruments; Houston, TX). After arterial pressure was recorded, the catheter was advanced to the LV guided by pressure tracing waves as described previously [17 (link)]. After stabilization, signals of pressure and volume were continually recorded by using a P–V conductance system (MPVS Ultra, emka TECHNOLOGIES, Paris, France) coupled to a digital converter (ML-870, ADInstruments, Colorado Springs, CO). Hemodynamic parameters were measured under different preloads, which were elicited by transiently compressing the abdominal inferior vena cava.