Abstract 12964: Over Expression of miR1 Leads to EAD Development in Cardiomyocytes by Increasing CaMKII and Calcium Current
Introduction: miR1 has been shown to induce cardiac arrhythmias in animal models but the mechanism is unclear. We overexpress miR1in rat cardiomyocytes and investigated the cellular mechanism of arrhythmia development.
Methods: Rat cultured ventricular cardiomyocytes were infected with adenovirus-miR1 and adenovirus-GFP (control). Voltage and current clamp were used to record calcium current and action potentials at 35°C. Cells were loaded with Fura2-AM for measurement of calcium transients.
Results: Action potential duration at 90% repolarization (APD90) was 542.4% longer in ad-miR1 cardiomyocytes compared to control (377.1±137.2 ms vs. 58.7±7.2 ms vs., n=7, p<0.05) at 0.25Hz. This increase in APD90 was due, in part, to a 174% increase in the incidence of early afterdepolarizations (EADs) in ad-miR1 cardiomyocytes (22±2 EADs/100 beats vs. 8±1 EADs/100 beats, n=25, P<0.05). There was no difference in the incidence of delayed afterdepolarizations (DADs) at 2Hz. To understand the mechanism of this arrhythmogenesis, we measured calcium current. Ad-miR1 cardiomyocytes had a calcium current density than in control cells (18.7±0.7 pA/pF vs.14.2±1.8 pA/pF, n=7, p<0.05). Calcium transient peak height increased in ad-miR1 cardiomyocytes compared to control. To investigate the role of Ca2+/calmodulin-dependent protein kinase II (CamKII) activation, we blocked CamKII with KN93 (1μ M) in ad-miR1 cardiomyocytes. Calcium current decreased from 18.7±0.7 pA/pF to 10.0±1.0 pA/pF, (n=11, p<0.05) with the addition of KN93 and a corresponding decrease was also seen in APD90 (377.1±137.2 ms to 73.4±13.7 ms, n=11, p<0.05). EAD incidence decreased to the levels of control cells with the addition of KN93. Nifedipine was used to block calcium current. APD90 decreased in ad-miR1 cells, but remained longer than in control cells (67.3±1.7 ms vs 18.8±1.6 ms, n=16 and 13, p<0.05). These results suggest that increased calcium current contributes to the prolonged APD90 and that other currents may also involved in this prolongation in ad-miR1 infected cardiomyocytes.
Conclusions: Overexpression of miR1 induced EADs in rat cardiomyocytes, in part, by increasing calcium current and CaMKII, which may be a mechanism of arrhythmias by miR1 overexpression in animal models.
- © 2011 by American Heart Association, Inc.