The binding of Neuregulin-1 (Nrg1) to its cognate receptors ErbB-3 and -4 mediates intercellular and intracellular communication. In vitro, this interaction has been shown to control prolactin (PRL) secretion from pituitary tumour cells. However, Nrg1/ErbB signalling and its function in vivo are not well understood. In the present study, we demonstrated that type I and III Nrg1 isoforms were expressed in the rat anterior pituitary. We observed that Nrg1 positive gonadotrophs can form contacts with lactotrophs, which are positive for ErbB-3 receptor. In addition, we show that gonadotroph cell-derived Nrg1 regulates the secretion of an 18 kDa form of PRL from pituitary lactosomatotroph GH3 cells in vitro. The results obtained strongly suggest that gonadotrophs are the major source of Nrg1 in the normal anterior pituitary and that Nrg1 may function as a paracrine/juxtacrine regulator of PRL secretion.