Non-Linear Reduction of Ion Currents in Cultured Cardiac Myocytes: Correlation with a Loss of T-Tubules?

Background: Downregulation of K+ currents and loss of T-tubule density is well documented during heart failure. A similar reduction of both has been observed in cultured ventricular myocytes. We examined whether the loss of K+ and Ca2+ currents in cultured cardiac myocytes correlates with a loss of T-tubules.Methods: Atrial and ventricular myocytes were isolated and cultured up to 48 hours. Current and voltage clamp recordings were made using patch electrodes. Confocal scans of myocytes stained with di-8-ANEPPS were made to visualize myocyte ultrastructure.Results: Action potential (AP) recordings from control ventricular myocytes displayed a large phase 1 and spike-and-dome morphology. After 48 hours culture, the AP duration was greatly prolonged, and loss of the spike-and-dome morphology was noted. Voltage clamp analysis of Ito, IKr, and ICa revealed that the magnitude of all currents was significantly reduced after 48 hours in culture. However, IKr was reduced the largest after 48 hours (61% for IKr, 34% for Ito and 19% for ICa). IK1 was not significantly altered. Recovery from inactivation of Ito was greatly slowed in cultured ventricular cells. Confocal scans of T-tubule structure in ventricular cells showed a loss of T-tubules with prolonged days in culture. The loss of ion currents in ventricular cells may be related to a loss of T-tubules. We next measured Ito in atrial cells (which lack T-tubules) and found a downregulation of Ito when cultured or isolated from failing hearts.Conclusion: In cultured ventricular myocytes, there was a non-uniform loss of ion currents similar to what is observed in heart failure. Moreover, the loss of ion currents in ventricular cells does not appear to correlate with a loss of T-tubules since atrial cells (which lack T-tubules) exhibited a similar loss.