Functional heterogeneity of gonadotrophs in the ovine fetus: analysis by reverse hemolytic plaque assay.
Stimulation of sheep fetal gonadotrophs with 10(-7) M luteinizing hormone releasing hormone (LHRH) for 3 h in culture wells increased luteinizing hormone (LH) release over basal values. Using the reverse hemolytic plaque assay (RHPA), we demonstrated that this increase was due to a recruitment of LH-secreting cells. During gestation, the percentage of LH-containing cells able to release and the mean size of plaques were the highest at around 100-130 days and were usually lower in females than in males. In an attempt to delineate the involvement of protein kinase C (PKC) in LH release, cells were treated with an activator of PKC, phorbol 12-myristate 13-acetate (PMA). Stimulation of cells with 10(-6) M PMA for 3 h enhanced LH release in culture wells 2- to 3-fold more than did 10(-7) M LHRH. This increase was a consequence of an enhanced number of LH-secreting cells and, in males only, of an enhancement of the mean plaque size. The percentage of LH-secreting cells among LH-containing cells and the plaque areas were maximal between 110 and 120 days of gestation in both sexes. They were usually lower in females than in males. Stimulation of cells with LHRH plus PMA enhanced LH release in culture wells in an additive manner when compared to either factor alone in both sexes and at all fetal ages. This additive effect reflected an increase in the number of secreting cells. Under these conditions, plaque sizes were larger than the plaque sizes obtained with PMA alone in males and in females in late gestation. In conclusion, our results show that LHRH stimulated LH secretion from sheep fetal cells by recruiting secreting cells when compared to controls. Both 100-120 days of gestation and were higher in males than in females. Results following treatment of cells with PMA, either alone or in combination with LHRH, suggest that these two secretagogues act on two different subpopulations of gonadotrophs and probably through different pathways.
Taragnat
C
C
Station de Physiologie de la Reproduction des Mammifères Domestiques, URA-CNRS 1291, Institut National de la Recherche Agronomique, Nouzilly, France.
Durand
P
P
D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D005260:Female; D005333:Fetus; D007987:Gonadotropin-Releasing Hormone / Q000502:physiology*; D006462:Hemolytic Plaque Technique; D007986:Luteinizing Hormone / Q000378:metabolism*; D008297:Male; D010902:Pituitary Gland / Q000196:embryology / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D011863:Radioimmunoassay; D012756:Sheep; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D007987:Gonadotropin-Releasing Hormone; D007986:Luteinizing Hormone; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate