Duchenne Muscular Dystrophy (DMD) is a degenerative muscle disorder that is associated with myofibre necrosis. Increased generation of reactive oxygen species (ROS) (Haycock et al., 1996) and the delocalisation of neuronal nitric oxide synthase (nNOS) from the sarcolemma (Brenman et al., 1995) are two pathophysiological events that have been reported in dystrophic muscle. The aim of this study was to investigate whether the delocalisation of nNOS in mdx mice results in aberrant ROS generation in skeletal muscle. 2-4 month old wild type (WT)(n=6) and mdx(n=5) mice were anaesthetised using sodium pentobarbital(10mg/100g) via an intraperitoneal injection. Two microdialysis probes were inserted into the gastrocnemius muscle and perfused with saline for the measurement of nitrate/nitrite formation as a measure of NO release, or cytochrome c for the measurement of superoxide (McArdle et al., 2001). Following insertion of the microdialysis probes, baseline microdialysis perfusates were collected for 1 hour to allow the baseline to stabilise. Mice were then subjected to a 15-min period of isometric contractions (McArdle et al. 2001). Perfusates were collected during contractions and over the subsequent 1-hour recovery period. Immediately following the final collection, mice were killed by cervical dislocation and muscle samples were removed for analysis of nNOS expression. Total nNOS expression in muscle homogenates was determined by western blotting, and the localisation of nNOS was assessed through immunohistochemical studies. Data was analysed using a mixed design ANOVA (P<0.05). Superoxide generation during contraction was significantly increased in muscles of mdx mice compared with generation during contraction in muscles of WT mice (mean = 0.38 (SD 0.08) versus 0.25 (SD 0.11) nmol/15min, P0.05). Total nNOS levels were reduced in mdx skeletal muscle homogenates. Immunohistochemical studies revealed the absence of nNOS from the sarcolemma in mdx mice. The data is consistent with the theory that the loss of nNOS into the cytosol of the skeletal muscle fibre results in an alteration of nNOS function, leading to increased superoxide release. The effects of this altered nNOS function may have potential cytotoxic consequences that may be implicated in the pathogenesis of DMD.