We studied seasonal profiles of circulating testosterone concentrations among male and female adult Tree Swallows (Tachycineta bicolor) breeding in nest-box colonies near Ames, Iowa, USA. Mean plasma testosterone in males was elevated during nest establishment (0.63 ± 0.86 ng/ml) and incubation stages (0.28 ± 0.26 ng/ml), and was significantly lower after hatching (0.03 ± 0.05 ng/ml) when males began provisioning nestlings. Male swallows do not incubate and high testosterone during the incubation stage may facilitate pursuit of extra-pair matings. Female testosterone concentrations were an order of magnitude lower than those of males (nest establishment, mean  =  0.06 ± 0.09 ng/ml) and did not change significantly over the breeding season. These testosterone profiles support the hypothesis that elevated testosterone in males is associated with defense behaviors and obtaining additional mating opportunities during the first part of the breeding season, but is incompatible with parental care once the eggs have hatched.