Using antibodies against the hormone binding domain of the trout estrogen receptor (ER), the distribution of ER-immunoreactive (ER-IR) cells was studied in the brain of maturing diploid and triploid female rainbow trout using a streptavidin-biotin-peroxidase method followed by a nickel-intensified diaminobenzidine reaction. This technique resulted in an excellent signal/background ratio allowing unambiguous identification of positive cells. In all animals, ER-IR cells were consistently located in three brain regions, the ventral telencephalon, the anterior ventral preoptic region, and the mediobasal hypothalamus. About 250 ER-IR cells were observed in the ventral and dorsal parts of the ventral telencephalon. In the anterior nucleus preopticus periventricularis, about 2400 ER-IR cells were observed surrounding the preoptic recess. In the posterior hypothalamus, approximately 2700 ER-IR cells were located in the anterior, posterior and inferior divisions of the nucleus lateralis tuberis and in the nucleus saccus vasculosus. In these regions cell nuclei exhibiting different densities of staining were observed and absolutely no labeling of cytoplasmic processes was detected. These results are in partial agreement with those obtained either after injection of tritiated-estradiol in other teleots species or in situ hybridization of ER mRNAs in trout. In particular, no immunoreactivity was observed in the thalamic region nor in the nucleus posterioris periventricularis. These data indicate that target cells for estradiol are essentially located in brain regions involved in the neuroendocrine control of pituitary functions and having direct connections with the hypophysis.