The histological distribution of gonadotrophs containing either LH or FSH, but not both gonadotropins, has been demonstrated before in the juvenile and adult chicken throughout the caudal and cephalic anterior pituitary lobes. In the present investigation, the distribution of FSH- and/or LH-containing gonadotrophs was further investigated in the chicken embryo by use of the same homologous antibodies as used in our earlier study. Fluorescent dual-labeling immunohistochemistry revealed that during embryogenesis LH and FSH reside exclusively in separate gonadotrophs, as has been described before in the post hatch bird. LH-immunoreactive cells were observed for the first time at day 9 of embryogenesis. This is as much as 4 days earlier than the FSH-immunoreactive cells, which appeared at day 13 of embryogenesis. Our results confirm that FSH- and LH-containing gonadotrophs are distributed throughout both lobes of the anterior pituitary. No conspicuous differences were observed between the sexes in any of the aspects investigated. The described situation is unique in that it seems to imply the existence of separate cell lineages for FSH- and LH-producing cells, as opposed to the single gonadotrope lineage described in all other species studied so far, with the exception of bovine. Our data indeed raise the question as to which signaling and/or transcription factors may cause the unique dichotomy observed in the chicken gonadotrophs.