Responses to irradiation in the mammary epithelium are enhanced by hormonal treatments. (a) The timeline summarizes treatment schedules of mice. Pellets obtained from Innovative Research of America (Sarasota, FL, USA) were used to deliver either E+P to mice for 14 days (neonatal E+P or mature E+P). Placebo groups were implanted with control pellets at these times. Mice were mated starting at 5 weeks of age and nursed pups for 1 week, and then mammary glands were allowed to involute for 4 weeks post-weaning to produce parous females. Tissues were harvested to determine basal levels of p53 and TUNEL labeling or irradiated 6 hours prior to harvest to determine responses following DNA damage. (b) Radiation-induced accumulation of p53 was increased significantly by neonatal E+P and mature E+P compared with the placebo-treated controls. The proportion of p53+ mammary epithelial cells in the E+P-treated groups did not differ from that in parous mice. The tissues from Trp53-/- mice were devoid of immunoreactive p53. (c) Radiation-induced TUNEL labeling also showed significant increases following treatment with neonatal E+P and mature E+P compared with the placebo-treated controls. Radiation-induced apoptosis was diminished in Trp53-/- tissues compared with Trp53+/+. However, the proportion of TUNEL-positive epithelial cells in parous Trp53-/- was increased compared with the placebo-treated Trp53-/- controls. (d) Levels of spontaneous apoptosis were low in nulliparous mice but increased significantly by neonatal E+P and were similar to the levels observed in tissues from parous mice. No significant difference in TUNEL labeling was detected in Trp53-/- tissues from nulliparous and neonatal E+P-treated mice. Bars indicate statistical differences between means: *P < 0.05; **P < 0.01. E+P, estradiol and progesterone; Trp53, transformation-related protein 53 (gene in mouse encoding the p53 tumor suppressor protein); TUNEL, terminal uridine deoxynucleotidyl transferase dUTP nick-end labeling.