The main purpose of the present study was to: 1) to investigate differences between males and females in brain activation when performing a mental rotation task, 2) investigate hemisphere differences in brain activation during mental rotation. Brain activation was measured with functional magnetic resonance imaging (fMRI). Image acquisition was performed with a 1.5 Tesla Siemens Vision MR scanner equipped with 25 m T/m gradients. Scanning of anatomy was done with a T1-weighted 3D FLASH pulse sequence. Serial imaging with 70 BOLD sensitive echo planar (EPI) whole brain measurements was done during stimulus presentations, divided into 7 blocks of 10 EPI multi-slice volume measurements each. Eleven subjects were presented with black-and-white drawings of 3-D shapes taken from the set of 3-D perspective drawings developed by Shepard and Metzler [1], alternated with 2-D white bars as control stimuli. In the experimental condition, the subjects were shown 36 pairs of 3-D drawings, presented in three blocks of 12 pairs of drawings. The drawings were always presented pairwise. On half of the trials, the two 3-D shapes were congruent but portrayed with different orientation, in the other half the two shapes were incongruent. MR data were analyzed with the SPM-96 analysis software. After subtraction of activity related to the 2-D control stimuli, clusters of significant activation were found in the superior parietal lobule (BA 7), more intensely over the right hemisphere, and bilaterally in the inferior frontal gyrus (BA 44/45). Males showed predominantly parietal activation, while the females showed inferior frontal activation. It is suggested that males and females may differ in the processing strategy used when approaching a 3-D mental rotation task, males using a 'gestalt' strategy and females using a 'serial' reasoning strategy.