The aim of this study was to investigate the use of [18F]fluorodeoxyglucose and positron emission tomography (FDG PET) for quantitative evaluation of glucose metabolism in skeletal muscle during walking. Ten young males underwent FDG PET twice during walks, which were done with or without an automated stride assistance system (SAS). Walk ratios were significantly increased by the SAS in seven subjects. Regional glucose metabolism in muscles between the crista iliaca and the planta was clearly visualized in all ten subjects. Glucose utilization increased significantly in the tibialis posterior and medial gastrocnemius muscles of the seven subjects in whom walk ratios were increased by the SAS. FDG PET is useful for analysis of muscle activity during exercise and rehabilitation.