We proposed the application of photodynamic therapy (PDT) as a new type of atrial fibrillation treatment. PDT is well known as a practical cancer treatment using cytotoxicity of reactive oxygen species generated by the photochemical interaction. We predicted that the photocytotoxic effect induced by PDT might cause electrical blockade in myocardial tissue. We studied the electrical blockade induced by the PDT with talaporfin sodium in vitro, ex vivo, and in vivo. The cell lethality measurement using rat cardiac myocytes confirmed the PDT-induced photocytotoxic effect and its dependence on the loading time of the photosensitizer. In ex vivo experiment using rat right ventricle, the PDT caused a shutdown of the stimulated electrical signal propagation. The long-lasting atrioventricular block induced by the PDT was obtained in vivo experiment with rat heart. These results revealed the possibility of atrial fibrillation treatment with the PDT.