Although fatigue is one of the major symptoms of persons with multiple sclerosis (MS), the behavioral and neural correlates are poorly understood. The present study utilized a novel approach to cognitive fatigue examining objective behavioral performance while simultaneously monitoring brain activity using fMRI. Fifteen persons with MS and 15 healthy controls were given 4 trials of a behavioral task assessing processing speed (mSDMT) during fMRI acquisition. It was hypothesized that individuals with MS would show an abnormal pattern of activity across time in specific brain areas previously hypothesized to subserve fatigue [Chaudhuri A, Behan PO. Fatigue and basal ganglia. J Neurol Sci 2000;179:34-42]. Specifically, it was hypothesized that persons with MS would show a greater increase in cerebral activation across time during behavioral performance than that seen in healthy controls, which was interpreted as fatigue. No difference in performance accuracy on the mSDMT was observed, although the MS group was significantly slower than controls. Behavioral alterations indicative of fatigue in the MS group were associated with increased activation in the basal ganglia, frontal areas including superior, medial, middle and inferior regions, parietal regions (precuneus and cuneus), thalamus and the occipital lobes. These data provide direct support for the Chaudhuri and Behan model of "central" fatigue which hypothesizes a specific role of the "non-motor" functions of the basal ganglia.