Baseline and stress-responsive glucocorticoid (GC) levels were characterized during the active period in free-ranging male and reproductive female little brown myotis (Myotis lucifugus). Bats were trapped and blood was sampled within 3 min of capture at two maternity sites during the summer and at one swarming site prior to hibernation in New England. Both GC hormones, cortisol and corticosterone, were detected, with cortisol accounting for an average of approximately 95% of total circulating GCs. Samples collected at the dusk emergence and after the first return from feeding showed significant seasonal differences across the active period (early pregnancy, mid-to-late pregnancy, lactation [and comparable mid-summer times for males], and pre-hibernation) within and between each sex. Elevated baseline values were found in mid-to-late pregnancy females at emergence, and in both males and females at the swarming site compared to other groups. Female GC values during mid-to-late pregnancy and during the pre-hibernation period were greater than those for males. Significantly higher GC levels following 15 min of restraint were exhibited by all animals in the summer and prior to hibernation. There was little variation between groups or sexes in the total GC levels reached following restraint. Taken together, these results suggest that: (1) GCs may be involved in the increased feeding and/or fat deposition characteristic of pregnancy and the pre-hibernation period, (2) GCs may be related to mating and to the generally increased levels of activity that occur during the pre-hibernation period, and (3) regardless of sex or reproductive condition, all animals maximally respond to restraint stress.