Cardiotoxicity of Isoproterenol and Cu 2+ 195 Cardiovascular Toxicology Humana Press Volume 1, 2001 195 *Author to whom all correspondence and reprint requests should be addressed: Dr. Fernando Remião, ICETA/CEQUP, Toxicology Department, Faculty of Pharmacy, Rua Aníbal Cunha, 164, 4050/047 Porto, Portugal. E-mail: remiao@ff.up.pt Cardiovascular Toxicology, vol. 1, no. 3, 195–204, 2001 Copper Enhances Isoproterenol Toxicity in Isolated Rat Cardiomyocytes Effects on Oxidative Stress Fernando Remião, Helena Carmo, Félix Carvalho, and M. Lourdes Bastos Toxicology Department, ICETA/CEQUP, Faculty of Pharmacy, University of Porto, Rua Aníbal Cunha, 164, 4050/047 Porto, Portugal Cardiovascular Toxicology (2001) 01 195–204 $12.50 (http://www.cardiotox.com) © Copyright 2001 by Humana Press Inc. All rights of any nature whatsoever reserved. 1530-7905/01 Humana Press Abstract Sustained high levels of circulating catecholamines may result in cardio- toxicity. Although cardiotoxicity could occur primarily via adrenoceptor acti- vation, there is increasing evidence that it may also occur through oxidative mechanisms. In fact, catecholamines can be converted into aminochromes by auto-oxidation, enzymatically or metal catalyzed, with the concomitant pro- duction of reactive intermediates and free radicals. Nevertheless, there is only scarce information concerning the effects of the catecholamine oxida- tion process on isolated cardiomyocytes. The aim of this work was to evaluate the cardiotoxic effects of isoproterenol (ISO) and its oxidation process in freshly isolated adult rat cardiomyocytes by assessing the cell shape, lactate dehydrogenase leakage, reduced and oxi- dized glutathione content, and glutathione reductase, peroxidase, and trans- ferase activities. ISO was incubated at concentrations of 0.1, 0.5, and 1 mM in cardiomyocyte suspensions at subphysiological and physiological Ca 2+ concentrations for 4 h. The same study was repeated in the presence of 20 µM of Cu 2+ . The levels of ISO in the incubation medium were monitored through- out the assays. Isoproterenol (1 mM) induced both glutathione oxidation and conjuga- tion, but this effect decreased at subphysiological Ca 2+ concentrations. The concomitant incubation with Cu 2+ increased ISO oxidation and increased the glutathione oxidation but decreased the extent of glutathione conjugation. Although only a partial ISO oxidation was observed for all studied ISO con- centrations in the presence of copper, the underlying oxidative process or its oxidation products, or both, were sufficient to induce a loss of cardiomyocyte viability and a decrease in the glutathione reductase, peroxidase, and trans- ferase activities. Thus, the results suggest that the oxidation of catecholamines could be a major mechanism for catecholamine-induced cardiotoxicity. Key Words: Isoproterenol; cardiotoxicity; oxidative stress; cardiomyocytes; quinone; aminochrome; catecholamines; heart; copper; glutathione.