Membrane alterations resulting from states of myocardial ischemia have been hypothesized as a major factor in the development of irreversible cellular damage. In this proposal, studies will be performed to investigate the mechanism(s) initiating membrane damage as well as the functional, structural and biochemical consequences of membrane damage. Isolated perfused heart preparations will be used for these studies. The components of myocardial ischemia that will be tested for their involvement in the development of membrane alterations will include: 1) an evaluation of damage due to the generation of oxygen-derived free radicals resulting in lipid peroxidation of biological membranes; 2) an examination of membrane phospholipid alterations due to the activation of phospholipases; and 3) an evaluation of cellular and subcellular electrolytes, including documentation of shifts that occur relatively early during ischemia, and evaluation of the role of sodium alterations in producing calcium overloading by sodium-calcium exchange. Specific interventions including free radical scavengers, such as superoxide dismutase, catalase and Alpha-tocopheral, and phospholipase inhibitors, such as chlorpromazine or newer agents, will be evaluated for their ability to protect against specific alterations thereby providing information on the involvement of various components of ischemia in the damage process. Indices of damage to be evaluated will include: measurements of lipid peroxidation products; phospholipid degradation; alterations in the levels of free fatty acids; alterations in triacylglycerol; electrolyte alterations; structural integrity; function; levels of adenine nucleotides; and enzyme loss. Statistical comparisons will be made between control, ischemic, and reperfused hearts, and hearts treated with various interventions. A calcium paradox model will be evaluated for alterations in phospholipids and compared to ischemia to determine if phospholipid alterations may be a general type of injury resulting in myocardial damage. The proposed studies will provide important insights concerning the significance of membrane lipids in the maintenance of cell survival, the influence of electrolyte alterations in the development of irreversible myocardial injury, and the ability of interventions to protect against specific aspects of the damage process and provide information as to the relative importance of these phenomenon in myocardial injury.
