title: Coculturing posterior pituitary and GH3 cells: dramatic stimulation of prolactin gene expression.
abstract: Recent evidence suggests that the posterior pituitary (PP), also called the neurointermediate lobe, regulates PRL release. We previously reported that cocultures of anterior pituitary and PP cells resulted in a 2- to 3-fold increase in PRL content and release. For this study we chose GH3 cells (a somatomammotroph tumor cell line) to determine whether coculturing GH3 with PP cells: 1) stimulates the release and cell content of PRL as compared with GH; 2) increases GH3 cell proliferation; and 3) affects PRL messenger RNA (mRNA) levels. Exp 1. GH3 cells (25,000 cells per well; 25K) were cocultured with PP cells (0K, 12.5K, or 25K) from male rats in serum-free media for 1, 2, 4, and 7 days; hormones were measured by RIA. Coculturing resulted in 5- to 10-fold increases in both media and cell PRL that were linear with time and dependent on the number of PP cells. In contrast, media GH increased only 1.5- to 2-fold, and GH cell content reduced by 75%. Exp 2. GH3, PP, and GH3 + PP cells were cultured for 1, 2, and 4 days and then incubated with [3H]thymidine for 5 h. The incorporation of [3H]thymidine in GH3 cells remained constant over time and showed a small, early increase in cocultures. In contrast, incubation of PP cells alone resulted in a 50- to 60-fold rise in [3H]thymidine incorporation from days 1-4 in culture. Exp 3. Cytoplasmic mRNA was determined by slot blot hybridization with 32P-labeled complementary DNA probes for PRL and GH. After coculturing 25K GH3 cells with 12.5K and 25K PP cells for 4 days, PRL mRNA levels increased 15- and 30-fold, respectively, whereas GH mRNA levels rose less than 2-fold. Neither PRL nor GH mRNA were detected in PP cells.
  
  
  CONCLUSIONS
  1) coculturing GH3 with PP cells dramatically stimulates PRL gene expression, synthesis, and release; 2) this response is specific for PRL, has little effect on GH, and is not due to increased GH3 cell proliferation; and 3) we speculate that a subpopulation of intermediate lobe cells, possibly with a proliferative capacity, is responsible for inducing these effects.
authors.affiliation: Department of Physiology and Biophysics, Indiana University School of Medicine, Indianapolis 46202.
authors.forename: A
authors.initials: A
authors.lastname: Corcia
authors.forename: R
authors.initials: R
authors.lastname: Steinmetz
authors.forename: J W
authors.initials: JW
authors.lastname: Liu
authors.forename: N
authors.initials: N
authors.lastname: Ben-Jonathan
mesh_terms: D000818:Animals; D015152:Blotting, Northern; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D015870:Gene Expression; D013006:Growth Hormone / Q000235:genetics / Q000378:metabolism; D008297:Male; D009693:Nucleic Acid Hybridization; D010904:Pituitary Gland, Posterior / Q000502:physiology*; D011388:Prolactin / Q000235:genetics* / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D014407:Tumor Cells, Cultured
chemical_list: D012333:RNA, Messenger; D011388:Prolactin; D013006:Growth Hormone; D004247:DNA