Although the interactions between sex steroids and GnRH have been extensively studied, little is known about the mechanism of estradiol (E2) effects on GnRH secretion. In the present study, we used retrochiasmatic hypothalamic explants of 50 day-old male rats and we observed that E2 significantly increased the glutamate-evoked GnRH secretion in vitro within 15 min, in a dose dependent manner. E2 also significantly increased the L-arginine-evoked GnRH secretion. E2 effects were time-dependent as the initially ineffective 10^-9 M concentration became effective after 5 h. The E2 effects involved the estrogen receptor (ER) since they were similarly obtained with the specific ER agonist PPT. The use of glutamate receptor agonists and antagonists indicated that E2 effects on GnRH secretion evoked by both glutamate and L-Arginine involved the AMPA/kainate receptors. Similar E2 effects on the kainate-evoked secretion were observed throughout development in both sexes. The observation of similar E2 effects using explants containing the median eminence (ME) alone indicated that the ME was a direct target for E2 rapid effects on the glutamate-evoked GnRH secretion. The signaling pathways involved in E2 effects included an increase in intracellular calcium and the activation of PKA, PKC and MAPK. It is concluded that E2 can stimulate the glutamate- and NO-evoked GnRH secretion in vitro through a rapid pathway involving the ER and kainate receptor as well as through a slower mechanism responding to lower E2 concentrations.