To determine the independent contributions of estradiol and progesterone to the auditory brainstem response (ABR) latency changes associated with the menstrual cycle, we obtained ABRs on young women with premature ovarian failure who were undergoing cyclic hormone replacement therapy (HRT). We evaluated the influence of cyclic HRT on the ABRs of young women in three controlled phases of the same replacement cycles: 1) no steroid replacement, 2) estrogen-only replacement (E2-only), and 3) estrogen-plus-progesterone replacement (E2-plus-P). A significantly lengthening of wave V peak latency and I-V interpeak interval was found during E2-only replacement. Despite equivalent circulating estradiol levels, both wave V peak latencies and wave I-V interpeak intervals significantly decreased during the E2-plus-P replacement phase as compared to the E2-only replacement phase. These findings are compatible with the hypothesis that estradiol potentiates secretion of the inhibitory neurotransmitter gamma-amino-butyric acid (GABA) at auditory nerve synapses, leading to delayed synaptic conduction time. Progesterone is known to blunt E2-potentiated GABA release and may antagonize its prolongation of wave V latency.