Acute and Subacute Effects of Injury on the Canine Alveolar Septum: Methods (2)
Prcparation of Lung Specimens
Six lung specimens were obtained for ultrastructural study as follows: two were control lungs from normal dogs (untreated controls); one was from the animal into which 24 ml of physiologic saline solution containing 1 mg of India ink had been instilled into the right lung (saline control); the fourth lung was obtained from an animal 4 h after exposure to papain in the right lung (4 h papain); a fifth, two weeks after exposure; and the sixth, one month after exposure to papain (two-week papain and four-week papain dogs, respectively).
For ultrastructural studies the animals were sacrificed at the prescribed intervals following exposure to papain, up to four weeks. Anesthesia was induced (or maintained, if sacrifice occurred on the same day as the exposure to saline or papain), and the sternum was split, allowing access to the mediastinum. Vascular perfusion-fixation was carried out as previously described. Ventilation was maintained with room air using a positive inspiratory pressure of 20 cm of water with a Harvard respirator. The main pulmonary artery was cannulated via the right ventricular outflow tract. In some instances, the right and left pulmonary arteries were individually cannulated, so that the right and left lungs could be perfused separately. buy prednisone
A cannula for drainage was inserted into the left ventricle. In order to wash out the blood, perfusion with heparinized Ringer s lactate containing 2.0 g of procaine per liter of solution or with a solution as modified by Sacks et al was begun immediately, at a perfusion pressure of 30 to 38 cm of water. After 5 to 10 min, the perfusate was changed to 2.5 percent glutaraldehvde in 0.1-M phosphate buffer and continued for 25 min. The pulmonary veins then were ligated, while maintaining perfusion pressure; the trachea, arterial cannulas and left ventricular drainage tubes were clamped; and the lungs were removed vn bloc and floated in the fixative for 24 h at 4°C.