Background Previously, we showed a mouse model (ACE8/8) of cardiac renin-angiotensin system (RAS) activation has a high rate of spontaneous ventricular tachycardia (VT) and sudden cardiac death (SCD) secondary to a reduction in connexin43 (Cx43) level. Angiotensin-II activation increases reactive oxygen species (ROS) production, and ACE8/8 mice show increased cardiac ROS. We sought to determine the source of ROS and if ROS played a role in the arrhythmogenesis. Methods and Results Wild-type and ACE8/8 mice with and without two weeks of treatment with L-NIO (nitric oxide synthase inhibitor), sepiapterin (precursor of tetrahydrobiopterin), MitoTEMPO (mitochondria-targeted antioxidant), TEMPOL (a general antioxidant), apocynin (NADPH oxidase inhibitor), allopurinol (xanthine oxidase inhibitor), and ACE8/8 crossed with P67 dominant negative mice to inhibit the NADPH oxidase were studied. Western blotting, detection of mitochondrial ROS by MitoSOX Red, electron microscopy, immunohistochemistry, fluorescent dye diffusion technique for functional assessment of Cx43, telemetry monitoring, and in-vivo electrophysiology studies were performed. Treatment with MitoTEMPO reduced SCD in ACE8/8 mice (from 74% to 18%, P<0.005), decreased spontaneous ventricular premature beats, decreased VT inducibility (from 90% to 17%, P<0.05), diminished elevated mitochondrial ROS to the control level, prevented structural damage to mitochondria, resulted in 2.6 fold increase in Cx43 level at the gap junctions, and corrected gap junction conduction. None of the other antioxidant therapies prevented VT and SCD in ACE8/8 mice. Conclusions Mitochondrial oxidative stress plays a central role in angiotensin II-induced gap junction remodeling and arrhythmia. Mitochondria-targeted antioxidants may be effective antiarrhythmic drugs in cases of RAS activation.

Objectives The aim of this study was to evaluate the role of c-Src inhibition on connexin43 (Cx43) regulation in a mouse model of myocardial infarction (MI). Background MI is associated with decreased expression of Cx43, the principal gap junction protein responsible for propagating current in ventricles. Activated c-Src has been linked to Cx43 dysregulation. Methods MI was induced in 12-week-old mice by coronary artery occlusion. MI mice were treated with c-Src inhibitors (PP1 or AZD0530), PP3 (an inactive analogue of PP1), or saline. Treated hearts were compared to sham mice by echocardiography, optical mapping, telemetry ECG monitoring, and inducibility studies. Tissues were collected for immunoblotting, quantitative PCR, and immunohistochemistry. Results Active c-Src was elevated in PP3-treated MI mice compared to sham at the scar border (280%, p=0.003) and distal ventricle (346%, p=0.013). PP1 treatment restored active c-Src to sham levels at the scar border (86%, p=0.95) and distal ventricle (94%, p=1.0). PP1 raised Cx43 expression by 69% in the scar border (p=0.048) and by 73% in distal ventricle (p=0.043) compared to PP3 mice. PP1-treated mice had restored conduction velocity at the scar border (PP3: 32 cm/s, PP1: 41 cm/s, p < 0.05) and lower arrhythmic inducibility (PP3: 71%, PP1: 35%, p < 0.05) than PP3 mice. PP1 did not change infarct size, ECG pattern, or cardiac function. AZD0530 treatment demonstrated restoration of Cx43 comparable to PP1. Conclusions c-Src inhibition improved Cx43 levels and conduction velocity and lowered arrhythmia inducibility after MI, suggesting a new approach for arrhythmia reduction following MI.