Adult cardiac myocytes in primary culture: cell characteristics and insulin-receptor interaction

Calcium-tolerant adult cardiac myocytes were kept in culture under serum-free conditions in the presence of physiological concentrations of insulin. Up to 4 days, 70% of cells retained their in vivo rodshaped morphology without gross structural alterations. During that period a constant ATP-to-ADP ratio was observed with a mean value of 10.6 +/- 0.5 (n = 4). The rate of 14Cphenylalanine incorporation remained unaltered up to 63 h in culture. Insulin binding to cultured cells was found to be time-and temperature-dependent, reversible, and highly specific. Scatchard analysis of equilibrium binding data showed a curvilinear plot with a high-affinity segment yielding an apparent dissociation constant of 4.5 X 10(-10) mol/l and a receptor number of 125,000 sites/cell. Both affinity and receptor number remained unaltered between 18 and 66 h in culture. 14Cphenylalanine incorporation was stimulated by 108% in cardiocytes cultured in the presence of high concentrations of insulin (1.7 X 10(-7) mol/l) for 63 h, when compared with control cells cultured in the absence of insulin. These data demonstrate the retention of structural integrity, insulin receptors, and insulin responsiveness in primary cultured adult cardiac myocytes and provide a useful model for long-term studies on the regulation of insulin action on the heart.