The oviduct plays a crucial role in mammalian reproduction by providing an optimal environment for the final maturation and transport of gametes, fertilization, and early embryonic development. It is now recognized that these reproductive events in vitro can be either negatively or positively affected by reactive oxygen species such as hydrogen peroxide and lipid hydroperoxides. In the current study, we analyzed the expression of the phospholipid hydroperoxide glutathione peroxidase (PHGPx or GPx-4), a selenoenzyme that directly reduces membrane-bound lipid hydroperoxides in the bovine oviduct. Using in situ hybridization, we demonstrated that GPx-4 expression is almost restricted to the oviductal luminal epithelium in contrast to GPx-1, which is widely distributed, and GPx-2 and -3, which are mainly detected in the epithelial cells and lamina propria. Interestingly, real-time quantitative RT-PCR analysis showed that GPx-4 expression was highest during the follicular and postovulatory phases. In addition, GPx-4 expression was highest in the isthmus proximal to the dominant follicle during the follicular stage and remained high during the postovulatory period. This increased in expression of GPx-4 corresponded to increased GPx-4 enzymatic activity. Based on intrauterine infusion of estradiol, we determined that the increase in expression and activity of GPx-4 is estrogen mediated. This work clearly demonstrates that GPx-4 gene expression is influenced by the proximity of the dominant follicle in the oviduct in vivo. We propose that GPx-4 has an important role in the physiological control of peroxide tone in the bordering cells of the oviductal lumen.