In vitro development of placode-derived LHRH neurons: possible involvement of alpha-fetoprotein

Using the olfactory placode (NAP) of 12.5-day-old and vomeronasal organ (VNO) of 14.5-day-old (E12.5, E14.5) rat embryos, we examined in vitro the roles of brain tissues in the development of LHRH neurons. The culture was performed singly or in combination with various brain tissues of E12.5 and E14.5 embryos. Furthermore, a serum-free basal culture medium was supplemented with a water extract of cerebral cortex or septopreoptic-medial basal hypothalamus (S-MBH) of E18.5 embryos and 2-day-old (N2) newborns. In cocultures, many LHRH cells derived from the NAP or VNO migrated into the collocated tissues of the forebrain vesicles and medial basal hypothalamus, especially into the latter, along neural cell adhesion molecule (NCAM)-positive fibers projecting from the NAP and/or the brain tissues, especially in the case of the medial basal hypothalamus. The effects of the brain extracts were evaluated by differentiation, migration, neurite extension, and survival of LHRH neurons. E18.5 S-MBH extract was most effective. In the analysis of the protein composition of the extracts, alpha-fetoprotein (AFP) was determined only in the E18.5 S-MBH extract. When anti-AFP IgG was added to the cultures containing E18.5 S-MBH extract, the stimulative effects of the extract were reduced to the level of the N2 S-MBH extract. When we cultured the NAP by adding AFP in the basal culture medium, the development of LHRH cells was somewhat promoted. The actual roles of AFP thus remain to be further elucidated.