Modulation of the nitric oxide producing system (demonstrated via the NADPH-diaphorase histochemical reaction) by oestradiol has been established in several structures of the rat brain. The present study aimed to explore the possible regulation of NADPH-diaphorase activity by oestradiol in neurones of the supraoptic (SON) and paraventricular (PVN) nuclei and the role of oestrogen receptors (ERα and ERβ) in this regulation. Adult ovariectomised rats were divided into six groups and injected either with vehicle or a single dose of oestradiol, a selective ERα agonist-PPT [4,4′,4″-(4-propyl-[1H]-pyrazole-1,3,5-triyl)trisphenol], a selective ERβ agonist-DPN [2,3-bis(4-hydroxyphenyl)-propionitrile], a selective ERα antagonist-MPP [1,3-bis(4-hydroxyphenyl)-4-methyl-5-[4-(2-piperidinylethoxy)phenol]-1H-pyrazole dihydrochloride] or a selective ERβ antagonist-PHTPP (4-[2-phenyl-5,7-bis(trifluoromethyl)pyrazolo[1,5-a]pyrimidin-3-yl]phenol). The number of NADPH-diaphorase positive elements in the SON and the PVN was modulated by both ERs but, depending on the nucleus, ERα and ERβ ligands induced different effects. These results suggest that the regulation of nitrergic system by ERs may play a role in the control of oestrogen-dependent physiological mechanisms regulated by the SON and the PVN.