Atrial fibrillation (AF) is a growing cardiovascular epidemic lacking effective treatment. Reactive oxygen species are believed to contribute to AF pathophysiology, yet general antioxidants have limited effectiveness. Since mitochondria are abundant in the heart and a major ROS producer, mitochondrial oxidative stress (MitoOxS) could be a therapeutic target. To determine this, rat inducible immortalized atrial myocytes (iAMs) were tachypaced to mimic AF, followed by assessment of calcium transients, contractility, mitochondrial respiration and morphology, and ROS damage. Cells were pretreated with MitoTEMPO or Sul-238 to assess their protective effects. In Drosophila, heart wall contractions were analyzed to assess arrhythmogenesis after mitochondrial antioxidant pretreatment. Using the GAL4-UAS system, mitochondrial ROS levels and the effect of SOD1 or SOD2 knockdown or overexpression on arrhythmogenesis were evaluated. Tachypacing induced contractile dysfunction and arrhythmogenesis, mitochondrial impairment, and ROS damage in iAMs and increased mitochondrial ROS and arrhythmogenesis in Drosophila. Both MitoTEMPO and Sul-238 treatments prevented mitochondrial dysfunction and arrhythmogenesis in iAMs and rescued arrhythmia in Drosophila. Underscoring the potential to target MitoOxS specifically, SOD2 knockdown promoted arrhythmogenesis in iAMs and Drosophila, whereas SOD2 overexpression rescued tachypacing-induced arrhythmia. MitoOxS is thus a key driver of tachypacing-induced contractile dysfunction and arrhythmia. Mitochondria-targeted antioxidants, such as MitoTEMPO or Sul-238, represent promising therapeutic strategies for AF.
