Gonadotropin-releasing hormone (GnRH) neurosecretion is subject to regulation by insulin, IGF-1, leptin, and other neuroendocrine modulators whose effects may be conveyed by activation of phosphoinositide 3-kinase (PI3K)-mediated pathways. It is not known, however, if any of these regulatory actions are exerted directly, via activation of PI3K in GnRH neurons, or if they are primarily conveyed via effects on afferent circuitries governing GnRH neurosecretion. To investigate the role of PI3K signaling in GnRH neurons we used conditional gene targeting to ablate expression of the major PI3K regulatory subunit, p85, in GnRH neurons. Combined in situ hybridization and immunohistochemistry confirmed reduction of p85 mRNA expression in GnRH neurons of GnRH-p85KO animals. Females of both genotypes exhibited estrous cyclicity and had comparable serum luteinizing hormone (LH), estradiol-17 and follicle stimulating hormone (FSH) levels. In male GnRH-p85KO mice, serum LH, testosterone and sperm counts were significantly reduced compared to WT. To investigate the role of the other major regulatory subunit, p85 on the direct control of GnRH neuronal function, we generated mice with a GnRH-neuron-specific p85 deletion on a global KO background. No additional reproductive effects in male or female mice were found, suggesting that p85 does not substitute p85 activity toward PI3K function in GnRH neurons. Our results suggest that p85, and thus PI3 kinase activity, participates in the control of GnRH neuronal activity in male mice. The sex-specific phenotype in these mice raises the possibility that PI3K activation during early development may establish sex differences in GnRH neuronal function.