Physical exercise has many health benefits, including improved cardiovascular fitness, lean muscle development, increased metabolism, and weight loss, as well as positive effects on brain functioning and cognition. Recent evidence suggests that regular physical exercise may also affect the responsiveness of reward regions of the brain to food stimuli. We examined whether the total number of minutes of self-reported weekly physical exercise was related to the responsiveness of appetite and food reward-related brain regions to visual presentations of high-calorie and low-calorie food images during functional MRI. Second, we examined whether such responses would correlate with self-reported food preferences. While undergoing scanning, 37 healthy adults (22 men) viewed images of high-calorie and low-calorie foods and provided desirability ratings for each food image. The correlation between exercise minutes per week and brain responses to the primary condition contrast (high-calorie>low-calorie) was evaluated within the amygdala, insula, and medial orbitofrontal cortex, brain regions previously implicated in responses to food images. Higher levels of exercise were significantly correlated with lower responsiveness within the medial orbitofrontal cortex and left insula to high-calorie foods. Furthermore, activation of these regions was positively correlated with preference ratings for high-calorie foods, particularly those with a savory flavor. These findings suggest that physical exercise may be associated with reduced activation in food-responsive reward regions, which are in turn associated with reduced preferences for unhealthy high-calorie foods. Physical exercise may confer secondary health benefits beyond its primary effects on cardiovascular fitness and energy expenditure.