Recent evidence indicates that the GnRH receptor (GnRH-R) gene is expressed in a number of tissues besides the anterior pituitary gland, suggesting that GnRH may serve other functions in addition to its role as a hypothalamic releasing factor. In particular, high levels of GnRH-R transcripts have been detected in rat and human ovarian granulosa cells. To better understand the role of the GnRH-R in the ovary under physiological conditions and to determine which follicles are potentially responsive to the actions of GnRH, we used in situ hybridization histochemistry and quantitative reverse transcriptase-polymerase chain reaction for measurement of ovarian GnRH-R messenger RNA (mRNA) expression during the rat ovulatory cycle. Reverse transcriptase-polymerase chain reaction analyses revealed that total ovarian GnRH-R mRNA levels were elevated significantly at 1800 h on proestrus and again at 0900 and 1800 h on estrus compared to metestrus 0900 h levels. In situ hybridization analysis of GnRH-R gene expression at different stages of follicular maturation revealed significant variation in GnRH-R mRNA levels with respect to the degree of follicular development as well as the estrous cycle stage. GnRH-R gene expression was greatest in the granulosa cells of Graafian and atretic follicles, with lower levels of expression present in preantral and small antral follicles and corpora lutea. GnRH-R mRNA levels in atretic follicles showed substantial variation across the 4-day rat estrous cycle, with mRNA levels increasing 3-fold on the day of proestrus coincident with the preovulatory gonadotropin surges. A second peak of expression in atretic follicles was observed on the morning of estrus. Levels of GnRH-R gene expression in corpora lutea also varied significantly during the estrous cycle, with gene expression increasing 3-fold between the morning of metestrus and the afternoon of proestrus. These results demonstrate that the level and localization of ovarian GnRH-R mRNAs change significantly during the rat ovulatory cycle. The finding that atretic follicles exhibit the greatest degree of GnRH-R gene expression is consistent with a role for GnRH in the induction of follicular atresia.