Transcranial Doppler (TCD) sonography is a frequently employed technique for quantifying cerebral blood flow by assuming a constant arterial diameter. Given that exercise increases arterial pressure by sympathetic activation, we hypothesized that exercise might induce a change in the diameter of large cerebral arteries. Middle cerebral artery (MCA) cross-sectional area was assessed in response to handgrip exercise by direct magnetic resonance imaging (MRI) observations. Twenty healthy subjects (11 female) performed three 5 min bouts of rhythmic handgrip exercise at 60% maximum voluntary contraction, alternated with 5 min of rest. High-resolution 7 T MRI scans were acquired perpendicular to the MCA. Two blinded observers manually determined the MCA cross-sectional area. Sufficient image quality was obtained in 101 MCA-scans of 19 subjects (age-range 20-59 years). Mixed effects modelling showed that the MCA cross-sectional area decreased by 2.1 ± 0.8% (p = 0.01) during handgrip, while the heart rate increased by 11 ± 2% (p < 0.001) at constant end-tidal CO (p = 0.10). In conclusion, the present study showed a 2% decrease in MCA cross-sectional area during rhythmic handgrip exercise. This further strengthens the current concept of sympathetic control of large cerebral arteries, showing in vivo vasoconstriction during exercise-induced sympathetic activation. Moreover, care must be taken when interpreting TCD exercise studies as diameter constancy cannot be assumed.