A novel survival model of cardioplegic arrest and cardiopulmonary bypass in rats: a methodology paper

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The CPB circuit (Figure 2) consisted of a 4 ml Plexiglas venous reservoir, a roller pump (Masterflex; Cole-Parmer Instrument Co., Vernon Hills, IL), and a custom-designed smallvolume oxygenator (M. Humbs, Valley, Germany). The 4 ml priming volume oxygenator was built of two Plexiglas shells (12.8 cm x 12.8 cm x 2.7 cm) that carry a sterile, disposable three layer hollow fiber membrane providing a surface area for gas exchange of 558 cm2 1821. To prevent excessive heat loss, one of the shells had an integrated heat exchanger. An in-line flow probe (2N806 probe and T208 flowmeter, Transonics Systems Inc., Ithaca, NY) was used to continuously measure CPB flow. The entire circuit was primed with 10 mL of 6% hetastarch (Hextend, Hospira Inc, Lake Forest, IL). All parts were connected through single use silicone tubing. During CPB, a flow rate of 150 mL├óÔé¼╦İkg-1├óÔé¼╦İmin-1 was maintained, and an average of 3 mL of hetastach was added to compensate losses and extravasation. At the start of CPB, frequency of ventilation was lowered to 30 min-1, and 0.5-1% isoflurane was administered through the oxygenator with 70% oxygen and additional CO2 as needed (_ stat blood gas management). After 15 min of CPB, the endoaortic clamp was quickly inflated while concomitantly 0.5 ml cardioplegia was infused via an infusion pump at a rate of 200 mL├óÔé¼╦İhr-1 through the central lumen of the angioplasty catheter. Cardioplegia solution consisted of 4 parts of standard undiluted adult cardioplegia induction solution (5% dextrose in 0.225% NaCl (655mL├óÔé¼╦İL-1) potassium chloride (95mEq├óÔé¼╦İL-1), tromethamine (238mL├óÔé¼╦İL-1), and citrate-phosphate-dextrose (CPD) solutions (60mL├óÔé¼╦İL-1), and 1 part esmolol 10 mg├óÔé¼╦İml-1. Ventilation was discontinued. Cardioplegic arrest was confirmed both by ultrasound and electrocardiographically. The CPB flow rate was adjusted as needed to maintain a constant venous reservoir blood level. At 15 min of arrest, a second dose of 0.4 mL of cardioplegia was administered to uphold the arrest. After 30 min of cardioplegic arrest, the balloon was deflated and removed and ventilation restarted at a slower rate of 35 min-1 and FiO2 of 0.7. CPB was maintained for another 30 min to allow for rewarming of the animals. After discontinuation of CPB, the ventilatory rate was raised to 60 min-1. The venous cannula was removed, and the incision closed. During the entire experiment, mean arterial pressure was maintained above 45 mmHg, with the use of small doses of phenylephrine if necessary. After cessation of CPB, the remaining blood left in the CPB circuit was centrifuged for 5 min at 3000 rpm, and the supernatant discarded. Two mL of the remaining red blood cell concentrate was then re-infused. The animals were ventilated for another 60 min, after which the remaining cannulae were removed, and the wounds closed. The rats recovered in a warmed and oxygenenriched environment for at least 12 hr prior to return to their home cages. Blood gas analysis was performed before the start of CPB, at 15 min of CPB, at 15 min of arrest, at the end of 30 min arrest, at the end of the total of 75 min of CPB, and at 1 hr post CPB, using an IL-GEM Premier 3000 blood gas analyzer (Global Medical Instrumentation, Ramsey, MI).

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