OXYGEN-INDUCED ENZYME-RELEASE AFTER IRREVERSIBLE MYOCARDIAL INJURY - EFFECTS OF CYANIDE IN PERFUSED RAT HEARTS

Abstract

The effects of 5 mM potassium cyanide (KCN) on creatine phosphokinase (CPK) release and cellular morphology were studied. Rat hearts were perfused with substrate-deficient media gassed with O2 or N2 (O2 medium, N2 medium) at 37 C, and effluent was collected for CPK analysis. Tissue fixation was with glutaraldehyde for EM and light microscopy. Experiments included were: continuous perfusion with O2- or N2-medium in the presence of KCN; 45 or 60 min of perfusion with N2-medium followed by O2 medium for 15 or 18 min, respectively; 45 min of perfusion with N2-medium with KCN added 15 min before reoxygenation with O2-medium plus KCN; 60 min of N2-medium plus KCN followed by O2-medium plus KCN for 180 min; as a control for irreversible injury, 21 min of perfusion with Ca-free O2-medium followed by 2.5 mM Ca-O2-medium ("Ca paradox"). Initial CPK release occurred about 30 min later from hearts perfused with O2-medium plus KCN than from hearts perfused with N2-medium plus KCN; upon reoxygenation after either 45 or 60 min of anoxia, hearts had a sudden peak of O2-induced CPK release. Most irreversibly injured cells were massively swollen and had sarcolemmal defects and contraction bands. Reversibly injured cells in the same hearts resembled normal myocardium. A previously unrecognized 3rd population of cells is described. These cells were characterized by contraction bands but were not swollen, had intact sarcolemma, and contained both normal and damaged mitochondria with intramatrical Ca accumulation granules. It could not be determined if these cells were reversibly injured or in an early stage of irreversible injury. KCN added 15 min before reoxygenation of hearts after 45 min of anoxia inhibited the sudden peak of O2-induced CPK release but not a slow sustained release. Small to moderate numbers of cells in these hearts contained contraction bands. After 60 min, KCN completely inhibited both O2-induced CPK release and contraction band formation. Addition of Ca to Ca-free hearts caused both massive CPK release and contraction band formation. It was concluded that the beginning of CPK release from oxygenated KCN-inhibited hearts requires about 30 min longer than from anoxic hearts; KCN can inhibit both O2-induced CPK release and contraction bands in irreversibly injured rat myocardial cells; sudden contracture of myocardial cells as occurs in the Ca paradox can result in massive CPK release; contraction bands occur in nonswollen cells, hence contraction bands can occur independently of massive cell swelling or membrane rupture. There may be 2 stages of irreversible myocardial injury, loss of control of contraction and progressive loss of mitochondrial and membrane integrity.