The epidermal growth factor receptor (ErbB) family and its ligands are essential for the regulation of multiple cellular processes required for mammalian reproduction. The objectives of this study were to investigate the expression and localisation of ErbB subtypes (ErbB1-4) and selected ligands, namely epidermal growth factor (EGF), amphiregulin (AREG) and neuregulin (NRG), in the cervix and vagina of cycling cows and to determine possible steroid hormone-dependence of their expression using immunohistochemistry. All four ErbBs and EGF, AREG and NRG proteins were found to be localised in the nucleus and cytoplasm of different cells in the cervix and vagina, and their expression differed during the oestrous cycle. During the follicular phase, in both the cervix and vagina, ErbB1, ErbB2, ErbB3, ErbB4 and EGF expression was higher in the luminal epithelium (LE) than in stromal and smooth muscle (SM) cells (P<0.05). During the luteal phase, the expression of ErbB1, ErbB3 and EGF in the LE was significantly different from that in stromal and SM cells in the cervix, whereas the expression of EGF and AREG differed in the vagina compared to the cervix (P<0.05). Throughout the oestrous cycle, in both the cervix and vagina, although ErbB2/human epidermal growth factor receptor 2 expression in the LE and SM cells was significantly higher than in the stromal cells (P<0.05), NRG expression was similar in the LE, stromal and SM cells (P>0.05). Overall, these results suggest that all four ErbBs and the EGF, AREG and NRG proteins may collectively contribute to several cellular processes in the bovine cervix and vagina during the oestrous cycle.