Oxidative stress plays an important role in the pathogenesis of many disease states. In the heart, reactive oxygen species are linked with cardiac ischemia/reperfusion injury, hypertrophy, and heart failure. While this correlation between ROS and cardiac pathology has been observed in multiple models of heart failure, the independent role of hydrogen peroxide (H₂O₂) in vitro and in vivo is unclear, owing to a lack of tools for precise manipulation of intracellular redox state. Here we apply a chemogenetic system based on a yeast D-amino acid oxidase to show that chronic generation of H₂O₂ in the heart induces a dilated cardiomyopathy with significant systolic dysfunction. We anticipate that chemogenetic approaches will enable future studies of in vivo H₂O₂ signaling not only in the heart, but also in the many other organ systems where the relationship between redox events and physiology remains unclear.