Muscle oxygenation responses are reportedly greater in the distal muscle region than in the proximal muscle region. We combined near infrared spectroscopy and electromyography (EMG) to determine whether regional differences in oxygenation are associated with differences in (1) muscle activation and/or (2) fatigue development. Nine males performed 2-min sustained isometric knee extensions at 15% and 30% maximum voluntary contraction during which oxygenation and EMG were recorded simultaneously from proximal and distal locations of the vastus lateralis muscle. Near infrared spectroscopy variables for oxygen saturation (StO2%) were initial slope at contraction onset, peak drop, and recovery slope at contraction end. Electromyography produced the root mean square to indicate muscle activation and mean power frequency changes over time (decreasing slope) to indicate fatigue development. For StO2%, significantly greater peak drop and steeper recovery slope were found for the distal muscle region than for the proximal muscle region. Root mean square, however, was not different between locations. Mean power frequency decreased throughout the contractions but changes were not different between locations. Our results indicate that for modest submaximal contractions, regional differences in oxygenation are not associated with differences in muscle activation or with fatigue development (as interpreted by changes in mean power frequency over time).