GnRH, the main regulator of reproduction, is produced in a variety of tissues outside of the hypothalamus, its main site of synthesis and release. We aimed to determine whether GnRH produced in the female rat pituitary and ovaries is involved in the processes leading to ovulation. We studied the expression patterns of GnRH and GnRH receptor (GnRH-R) in the same animals throughout the estrous cycle using real-time PCR. Hypothalamic levels of GnRH mRNA were highest at 1700 h on proestrus, preceding the preovulatory LH surge. No significant changes in the level of hypothalamic GnRH-R mRNA were detected, although fluctuations during the day of proestrus are evident. High pituitary GnRH mRNA was detected during the day of estrus, in the morning of diestrus 1, and at noon on proestrus. Pituitary GnRH-R displayed a similar pattern of expression, except on estrus, when its mRNA levels declined. Ovarian GnRH mRNA levels increased in the morning of diestrus 1 and early afternoon of proestrus. Here, too, GnRH-R displayed a somewhat similar pattern of expression to that of its ligand. To the best of our knowledge, this is the first demonstration of a GnRH expression pattern in the pituitary and ovary of any species. The different timings of the GnRH peaks in the three tissues imply differential tissue-specific regulation. We believe that the GnRH produced in the anterior pituitary and ovary could play a physiological role in the preparation of these organs for the midcycle gonadotropin surge and ovulation, respectively, possibly via local GnRH-gonadotropin axes.