INTERACTION OF ALPHA-T3-1 CELLS WITH LACTOTROPES AND SOMATOTROPES OF NORMAL PITUITARY IN-VITRO

A pituitary cell population of 14-day-old female rats, containing lact otropes and somatotropes but deprived of gonadotropes, was prepared by unit gravity sedimentation through a serum albumin gradient and allow ed to reaggregate either as such or after mixing this population with cells of the gonadotropic alpha T3-1 cell line. In a perifusion system gonadotropin-releasing hormone (GnRH) had no effect on prolactin (PRL ) and growth hormone (GH) release in the former aggregates but stimula ted PRL release in the latter. In the alpha T3-1 cell-containing aggre gates GnRH showed a biphasic effect on GH release: inhibition during e xposure to GnRH followed by a rebound secretion upon removal of the pe ptide. The aggregation capacity of alpha T3-1 cells with the normal pi tuitary cells was demonstrated by using an alpha T3-1 cell clone stabl y transfected with the reporter gene beta-galactosidase. Perifusion of the gonadotrope-deprived aggregates with medium conditioned by alpha T3-1 cell provoked a rapid stimulation of PRL release and a biphasic e ffect on GH release. Medium conditioned by the corticotropic cell line AtT20 also stimulated PRL release but had no concomitant effect on GH release. Medium conditioned by alpha T3-1 cells, when added for 40 h to aggregates of 14-day-old rat pituitary, provoked an increase in the number of H-3-thymidine (H-3-T)-labelled lactotropes and a decreased in the number of H-3-T-labelled somatotropes. The conditioned medium w as concentrated on Sep-Pak C18 and ultrafiltrated through an Amicon me mbrane with 3-kD molecular weight cut-off and the retained molecules s eparated by reversed-phase HPLC. The material stimulating H-3-T labell ing of lactotropes eluted from the column with a different retention t ime than material inhibiting H-3-T labelling of somatotropes, suggesti ng that the effect on lactotropes is mediated by (a) molecule(s) diffe rent from that affecting somatotropes. The effects of alpha T3-1 cells and their secretion products on lactotropes and somatotropes were com parable to those we previously observed using enriched populations of normal gonadotropes. The HPLC elution profiles of the substances affec ting H-3-T incorporation as well as the specificity of these effects w ere also similar to that of the substances isolated previously from go nadotrope-conditioned medium. The present data, therefore, support pre vious conclusions on the paracrine control of the lactotrope/somatotro pe lineage by the gonadotropes. Further purification and chemical char acterization of the growth factors with selective action on lactotrope s and somatotropes may lead to a better understanding of the developme nt of the latter lineage.