Abstract 20283: Calmodulin Kinase and Protein Kinase C Mediate the Effect of Increased Intracellular Calcium to Augment Late Sodium Current in Ventricular Myocytes
Objective: An increase in intracellular calcium concentration ([Ca2+]i) leads to augmentation of late sodium current (late INa) by activating Ca2+-calmodulin-dependent protein kinase II (CaMK-II). The objective of this study was to confirm our hypothesis that both CaMK-II and protein kinase C (PKC) are the signaling molecules whose activation result in increased late INa in the presence of an increased [Ca2+]i.
Methods: Whole-cell and open-cell patch clamp techniques were used to record late INa in rabbit ventricular myocytes dialyzed with pipette solutions containing various concentrations (0.1, 0.3 and 0.6 μM) of [Ca2+]i in the absence and presence of CaMK-II (KN-93), PKC (bisindolylmaleimide, Bim) and MAPK (U0126) inhibitors in the bath solution.
Results: Increases in [Ca2+]i from 0.1 to 0.6 μM resulted in an augmentation of late INa from 0.30 ± 0.03 to 0.45 ± 0.03 pA/pF (n=8–10, p<0.01) in a concentration dependent manner, which was associated with an increase in mean Na+ channel open probability from 0.0067 ± 0.0009 to 0.0621 ± 0.0047 and prolongation of channel mean open-time. In the presence of 0.6 μM [Ca2+]i, either KN-93 (10 μM) or Bim (2 μM) decreased late INa by 33.3 % and 60.0 % from 0.45 ± 0.03 and 0.44 ± 0.02 to 0.40 ± 0.04 and 0.36 ± 0.03 pA/pF, respectively (n=8 and 10, p<0.01 vs. 0.6 μM [Ca2+]i). A combination of KN-93 (10 μM) and Bim (2 μM) completely reversed the increase in late INa induced by 0.6 μM [Ca2+]i from 0.46 ± 0.03 to 0.30 ± 0.03 pA/pF (n=8, p< 0.001), as well as the changes in Na channel mean open probability and mean open-time. In addition, PKC activator phorbol myristoyl acetate (2 μM) increased the amplitude of late INa from 0.30 ± 0.05 to 0.39 ± 0.02 pA/pF (n=10, p<0.01). U0126 did not change the amplitude of late INa in the presence of increased [Ca2+]i levels.
Conclusions: Both CaMK-II and PKC are involved in the pathway by which an increased [Ca2+]I leads to augmentation of late INa in rabbit ventricular myocytes.
- © 2010 by American Heart Association, Inc.