The participation of type I GnRH receptor (GnRH-R) on GnRH-II-induced gonadotropin secretion in rat pituitary cells was investigated. Furthermore, we extended the study of GnRH-II action to ovarian cells. The GnRH-II was able to mobilize inositol triphosphate (IP₃) and to induce LH and FSH release in a dose-dependent manner in pituitary cells and in a GnRH-I-like manner. The GnRH-analog 135-18 (agonist for type II GnRH-R and antagonist for type I GnRH-R) was unable to elicit any cellular response tested in these pituitary cells. The GnRH-II responses were blocked by the type I GnRH-R-antagonists CRX or 135-18, suggesting that these effects were mediated by the type I GnRH-R. In contrast to pituitary cells, GnRH-I, but not GnRH-II, elicited an IP₃ response in superovulated ovarian cells; 135-18 also had no effect. However, GnRH-II as well as GnRH-I presented antiproliferative effects on these cells. Surprisingly, 135-18 had stronger antiproliferative effects than either GnRH peptide. The 135-18 analog, but not GnRH-I or GnRH-II, increased progesterone secretion in superovulated ovarian cells. These results strongly suggest that GnRH-II is able to stimulate rat pituitary cells through the type I GnRH-R, with no evidence for the presence of type II GnRH-R. On the other hand, our results indicate a putative GnRH-R in superovulated ovarian cells with response characteristics that differ from those of the GnRH-R in the pituitary.