Adult female rats show greater behavioral activation in response to both acute and repeated injections of amphetamine (AMPH) than adult male rats. The present experiments investigated whether life-long exposure to ovarian hormones in females contributes to their enhanced responsiveness to AMPH as adults and whether this effect depends on exposure to testosterone (T) in the neonatal period. In Experiment 1, female rats were ovariectomized either prior to puberty (EARLY) or in adulthood (ADULT). Locomotor activity in response to either 1.5 mg/kg AMPH or the saline vehicle was measured for 2 hr every third day on five occasions. On the sixth occasion all animals received 0.75 mg/kg AMPH in a test for sensitization. Each animal received either 5.0 micrograms estradiol benzoate (EB) or the oil vehicle (OIL), 30-35 min prior to each session. Experiment 2 was similar except that all animals were treated with T at birth. In Experiment 1, during the repeated treatment period, group EARLY had lower AMPH-induced activity scores than group ADULT, in both OIL and EB conditions. EB-treated rats had higher levels of AMPH-induced activity and showed greater changes in activity over days of testing than OIL-treated rats, regardless of time of ovariectomy. Animals previously exposed to AMPH showed higher levels of activity on the test for sensitization than animals receiving the drug for the first time, but this effect did not vary as a function of hormone condition or time of ovariectomy. Exposure of females to T at birth reduced responsiveness to AMPH and EB in adulthood, and ovariectomy prior to puberty had little effect in these animals.