In rats, hormonal fluctuations during the estrus cycle may have numerous behavioral and neurobiological consequences. The aim of this study was to investigate the effects of estrus cycles and citalopram on behavior, ultrasonic vocalizations, anxiety levels, and c-fos expression in rats. With this aim, the rats were grouped into two: (1) a control group (n = 16) and (2) a citalopram group (n = 16), which received daily intraperitoneal 20 mg/kg citalopram from baseline (D0) to the 10th day (D10). Behavioral analysis and ultrasonic vocalization (USV) recordings were made on D0 and D10. Next, the rats were further subgrouped according to estrus phases identified through a vaginal smear (8 proestrus rats and 8 non-proestrus rats, in each group). The rat's anxiety levels were analyzed with an elevated plus maze (EPM), and their c-fos expression was measured at the cingulate cortex, the amygdala, and the paraventricular thalamic nucleus. Our results showed that the citalopram group showed significantly more grooming behaviors on D10 than the control group (p = 0.002). USVs on D0, D10 and during the EPM did not show any significant differences between the groups. Proestrus rats in the control group showed significantly less anxiety-like behavior during the EPM than the non-proestrus rats in the control group (p = 0.028 for time spent in open arms, and p = 0.011 for entries into open arms). There was no significant difference in anxiety-like behavior between the control and citalopram groups, and between the proestrus and non-estrous rats in the citalopram group. C-fos expression at the amygdala (p = 0.013) and the paraventricular thalamic nucleus (p = 0.014) was significantly inhibited in the citalopram group. We concluded that estrus cycles have a significant effect on anxiety levels in rats, which may be suppressed behaviorally and neurobiologically by citalopram. (C) 2014 Elsevier Inc. All rights reserved.