Pictures from the International Affective Picture System were used in a functional magnetic resonance imaging study to assess gender differences in brain activation in ten male and ten female volunteers. The affectively positive, negative and neutral pictures were presented for 750 ms in a single event design and were carefully matched for arousal, valence and stimulus content. Men and women showed no significant difference in valence, arousal, skin conductance response and startle modulation. Only in men was amygdala activation observed in the pleasant condition. Furthermore, men showed a stronger brain activity for positive visual stimuli than women in the frontal lobe (inferior and medial frontal gyrus). In women, stronger brain activation for affectively negative pictures was observed in the anterior and medial cingulate gyrus. These results indicate that it is crucial to take gender differences into account when emotional paradigms are used in functional brain imaging.