Ventilation-perfusion (V̇a/Q̇) mismatch during exercise may result from interstitial pulmonary edema if increased pulmonary vascular pressure causes fluid efflux into the interstitium. If present, the increased fluid may compress small airways or blood vessels, disrupting V̇a/Q̇ matching, but this is unproven. We hypothesized that V̇a/Q̇ mismatch would be greatest in basal lung following heavy upright exercise, consistent with hydrostatic forces favoring edema accumulation in the gravitationally dependent lung. We applied new tools to reanalyze previously published magnetic resonance imaging data to determine regional V̇a/Q̇ mismatch following 45 min of heavy upright exercise in six athletes (V̇o2max = 61 ± 7 mL·kg−1·min−1). In the supine posture, regional alveolar ventilation and local perfusion were quantified from specific ventilation imaging, proton density, and arterial spin labeling data in a single sagittal slice of the right lung before exercise (PRE), 15 min after exercise (POST), and in recovery 60 min after exercise (REC). Indices of V̇a/Q̇ mismatch [second moments (log scale) of ventilation (LogSDV) and perfusion (LogSDQ) vs. V̇a/Q̇ distributions] were calculated for apical, middle, and basal lung thirds, which represent gravitationally nondependent, middle, and dependent regions, respectively, during upright exercise. LogSDV increased after exercise only in the basal lung (PRE 0.46 ± 0.06, POST 0.57 ± 0.14, REC 0.55 ±0.14, P = 0.01). Similarly, LogSDQ increased only in the basal lung (PRE 0.40 ± 0.06, POST 0.51 ± 0.10, REC 0.44 ± 0.09, P = 0.04). Increased V̇a/Q̇ mismatch in the basal lung after exercise is potentially consistent with interstitial pulmonary edema accumulating in gravitationally dependent lung during exercise. NEW & NOTEWORTHY We reanalyzed previously published MRI data with new tools and found increased ventilation-perfusion mismatch only in the basal lung of athletes following 45 min of cycling exercise. This is consistent with the development of interstitial edema in the gravitationally dependent lung during heavy exercise.