A crossover design experiment was conducted to examine the effects of GnRH on hormonal concentrations and changes in ovarian structures of lactating dairy cows during the estrous cycle. Cows were assigned randomly to receive either 100 micrograms of GnRH or saline 12 h after estrus (d 0) was first observed. Ultrasonographic examinations of the ovaries performed daily throughout the estrous cycle detected either three (n = 7) or four (n = 6) follicular waves. Transient increases in FSH generally preceding the follicular wave containing the dominant (preovulatory) follicle, and the emergence of the dominant follicle before the subsequent estrus occurred later in GnRH-treated than in saline-treated cows with either three or four follicular waves. Day of emergence of dominant follicles within follicular waves was preceded 83% of the time by a transient increase in FSH and was followed consistently by increased serum estradiol-17 beta, which generally reached peak concentrations concomitant with the peak diameter of the dominant follicle. Although clearly defined pulses were detected in 19 out of 26 sampling periods in d 7 and 14, number of FSH pulses and percentage of cows with pulses of FSH were similar in GnRH-treated and saline-treated cows with three or four follicular waves. In conclusion, treatment of cows with GnRH 12 h after the onset of estrus delayed the appearance of the next ovulatory follicle in cows with either three or four follicular waves. Regardless of treatment, emergence of dominant follicles was preceded in most cases by a transient increase in FSH and followed consistently by increased estradiol-17 beta.