The mechanisms responsible for the observed differences in contractile performance between immature and adult mammalian myocardium remain incompletely defined. Because of the structural and functional immaturity of the sarcoplasmic reticulum during the perinatal period, it is likely that the immature sarcolemmal membrane differs functionally from that in the adult heart. However, relatively little is known of the calcium regulatory properties of the sarcolemma with regard to perinatal development. The major specific aim of this proposal is to comprehensively characterize the transarcolemmal sodium-calcium (Na-Ca) exchange mechanism in the developing rabbit. Experimental results from four age groups will be compared: fetal (25-28 days post-conception), newborn (1-7 days of age), immature (14-21 days of age), and adult (6-10 months of age). Sarcolemmal membrane vesicles will be prepared from ventricular tissue by homogenization and sucrose gradient centrifugation. The kinetics of sodium-dependent calcium fluxes will be compared among the age groups. The effects of changes in potential regulatory factors (pH, temperature, sarcolemmal protein phosphorylation/dephosphorylation) will be examined. Na-Ca exchange will also be studied in suspensions of viable, calcium-tolerant myocytes prepared by enzymatic digestion of ventricular myocardium. Age-related comparisons of the effects of amiloride (an inhibitor of Na-Ca exchange in adult myocardium) in vesicles and myocytes will be used to assess the relative sensitivities to Na-Ca inhibition. 3H-Amiloride will be used to determine the binding characteristics of amiloride (binding affinity, binding site density) in vesicles and myocytes from each age group. The effects of changes in Na-Ca exchange activity on contractile function will be assessed in isolated right ventricular papillary muscles obtained from fetal, newborn, immature, and adult rabbits. These experiments will provide information regarding the perinatal development of sarcolemmal Na-Ca exchange. Completion of this project will provide important insights into the mechanisms involved in the perinatal maturation of myocardial contractile function.
