Material and Methods

Abstract

Kidneys of Wistar rats (Ivanovas, Kissleg; 150–300 g BW) were fixed by total body perfusion as described previously (Kaissling and Kriz 1982). After anesthesia with thiobutabarbital (Inactin, 10 mg/100 g BW) or pentobarbital (Nembutal, 0.12 ml/100 g BW i.p.), the abdominal cavity was opened, the aorta clamped beneath the renal arteries, and a catheter was inserted into the abdominal aorta distal to the clamps in a retrograde direction. The catheter contained less than 0.5 ml of a solution prepared of 2 ml of 0.9% sodium chloride and 1 ml of heparin (5,000 IU). After the catheter was connected to the perfusion device, the inferior vena cava was broadly opened, the clamp was removed, and perfusion was started. Without prior flushing, perfusion of the fixative was carried out at a pressure of 180–220 mm Hg for 3 min at room temperature. The fixative contained 1.5% glutaraldehyde, 1.5% formaldehyde, 0.05% picric acid, and 0.08% CaCl₂ in 0.1 M cacodylate buffer (pH 7.4). Small pieces of cortical tissue were postfixed in buffered 1% OsO₄ dehydrated through a graded series of ethanol and embedded in Epon 812 by standard procedures.