Recently, we described that free corticosterone levels in the brain of male Wistar rats, as assessed by in vivo microdialysis, show an ultradian rhythm with a pulse frequency of 1.2 pulses/h. To establish whether gender influences brain free corticosterone rhythms, we studied free corticosterone levels in the female Wistar rat under baseline and stressful conditions using microdialysis in the hippocampus. Analysis of the data with the PULSAR algorithm revealed that hippocampal free corticosterone levels show a clear ultradian pattern in female rats with a pulse frequency of 1.16 � 0.05 pulses/h between 09.00 h and 21.00 h. Further analysis showed that the pulse amplitude is significantly higher during the late afternoon/early night (15.00–21.00 h) than during the morning/early afternoon (09.00–15.00 h) phase (0.13 � 0.03 versus 0.07 � 0.01 μg/dl, respectively, P < 0.05). Pulse characteristics were extremely reproducible as demonstrated by the almost identical pulse parameters derived from two consecutive 24‐h periods [pulse frequency: 1.13 � 0.09 and 1.19 � 0.08 pulses/h; pulse amplitude: 0.11 � 0.05 and 0.10 � 0.02 μg/dl for day 1 and day 2 (09.00–21.00 h) respectively, P > 0.05]. Both exposure to a novel environment and forced swim stress increased hippocampal free corticosterone levels. However, the stress‐induced rise reached higher levels and was more prolonged after forced swimming (area under the curve: 46.84 � 9.25 and 12.08 � 1.69 arbitrary units for forced swimming and novelty stress respectively, P = 0.01). Importantly, the ultradian rhythm was rapidly restored after termination of the stress response. This is the first demonstration that the female rat brain is exposed to free corticosterone levels that follow a circadian as well as an ultradian pattern and show almost identical pulse characteristics as recently reported in male animals. These observations are of significance for further investigations into the dynamics of glucocorticoid action in the brain of both genders.