In goldfish, circulating LH and growth hormone (GH) levels surge at the time of ovulation. In the present study, changes in gene expression of salmon GnRH (sGnRH), chicken GnRH-II (cGnRH-II), somatostatin (SS) and pituitary adenylate cyclase activating polypeptide (PACAP) were analyzed during temperature- and spawning substrate-induced ovulation in goldfish. The results demonstrated that increases in GHRH/PACAP gene expression during ovulation are best correlated with the GH secretion profile. These results suggest that GHRH/PACAP, instead of GnRH, is involved in the control of GH secretion during ovulation. Increases of two of the SS transcripts during ovulation are interpreted as the activation of a negative feedback mechanism triggered by high GH levels. The results showed a differential regulation of sGnRH and cGnRH-II gene expression during ovulation, suggesting that sGnRH controls LH secretion whereas cGnRH-II correlates best with spawning behavior. This conclusion is further supported by the finding that non-ovulated fish induced to perform spawning behavior by prostaglandin F₂ treatment increased cGnRH-II expression in both forebrain and midbrain, but decreased sGnRH expression in the forebrain.