The primary cause of Duchenne Muscular Dystrophy, a condition that affects 1 in 3500 live male births, is a deficiency of the protein dystrophin. This deficiency leads to downregulation of the neuronal nitric oxide synthase (nNOS) and lowered production of nitric oxide. This study investigated the effects of L-arginine, the substrate of nNOS, on cardiac function and fibrosis in mdx mice, a widely used animal model for Duchenne Muscular Dystrophy. Six month old mdx mice received 5 mg/g body weight L-arginine by daily oral gavage for six months while control mdx and C57BL10ScSn mice received water by oral gavage. At the completion of treatment, mice were anaesthetised with sodium pentobarbitone (70 mg/kg, i.p.) prior to euthanasia by excision of the heart. Cardiac function was assessed with the Langendorff technique at a perfusion pressure of 80 mmHg. Mdx mice had an impaired left ventricular developed pressure relative to C57BL10ScSn control mice, but this difference was not evident in L-arginine treated mdx mice. The L-arginine treated mdx mice also showed an increased coronary flow and reduced diastolic stiffness relative to untreated mdx mice (P<0.05). This reduction in stiffness was associated with a significant reduction in cardiac collagen measured as the percent of left ventricular area stained with the collagen specific dye picrosirius red (P<0.05). These results indicate that chronic treatment with L-arginine produces the beneficial effects of improving function and reducing fibrosis in the dystrophin-deficient heart.