Pre-pubertal exposure of the developing reproductive tract (RT) to estrogen or xenoestrogens can have long-term consequences that compromise the reproductive performance of dairy cattle. This research examined the effects of the selective estrogen receptor modulator, tamoxifen (TAM), on gene and protein expression in the pre-pubertal RT with particular focus on signaling pathways that affect RT morphology. Tamoxifen was administered to heifer calves (n = 7) daily (0.3 mg/kg subcutaneously) from 28 to 120 d of age. Control calves (CON; n = 7) received an equal volume of excipient. Weight, gross measurements and samples of RT tissues were collected upon sacrifice at 120.7 ± 0.3 d of age. Protein and mRNA were extracted from snap frozen samples of vagina, cervix, uterine body, ovary and oviduct. Data were analyzed using the proc mixed procedure of SAS version 9.1.3 (SAS Institute Inc., Cary, NC) followed by the Tukey-Kramer test as a multiple comparison test. As we previously reported, overall weight of the RT was dramatically decreased following TAM treatment (P = 0.01). Both estrogen receptor α (ERα) protein and gene expression were dramatically reduced in the uterus, cervix and vagina (P < 0.01). In oviduct, ERα protein was reduced in the TAM treated animals (P < 0.01) while ERα gene expression was not affected. Similarly, the phosphorylated form of ERα protein was reduced in the cervix and vagina (P < 0.01) and tended to be reduced in the uterus (P = 0.09) of the TAM group. In contrast to other RT tissues, phosphorylated ERα protein abundance was greater in the ovaries of the TAM animals (P < 0.05). Even though insulin-like growth factor-I (IGFI) gene expression was higher in the uterus, oviduct and vagina of the TAM group (P < 0.01), IGFI receptor protein and gene expression did not differ. Total mitogen-activated protein kinases (MAPK) protein was higher in the oviduct, vagina and ovary (P < 0.01) but was lower in the uterus of TAM treated heifers (P < 0.01). The phosphorylated form of MAPK protein was similarly increased in the ovary but was decreased in the cervix of the TAM group (P < 0.05). In conclusion, the bovine pre-pubertal RT is affected by TAM treatment. Further research is needed to determine if these effects have long-term consequences for reproductive performance.