THYROTROPIN-RELEASING-HORMONE GENE-EXPRESSION IN THE ANTERIOR-PITUITARY .2. STIMULATION BY GLUCOCORTICOIDS

The present studies were undertaken to determine whether glucocorticoi ds (GC) regulate TRH gene expression in cultured anterior pituitary (A P) cells. AP cells derived from 15-day-old male rats were cultured for up to 18 days in Dulbecco's Modified Eagle's Medium-L-15 medium suppl emented with 1) fetal calf serum (FCS), 2) charcoal-treated FCS, 3) no rmal rat serum, or 4) serum from rats that were adrenalectomized, rend ered hypothyroid, and gonadectomized (ATG rat serum). Dexamethasone (D ex) or corticosterone (Cort) was added to the culture medium at variou s concentrations with exposure times ranging from 4-18 days. TRH and p repro-TRH-(25-50) in cellular extracts and release media were measured by RIA, and pro-TRH mRNA was determined by Northern blot analysis and in situ hybridization. Dex substantially stimulated cellular TRH and prepro-TRH-(25-50) accumulation under all culture conditions investiga ted, i.e. in medium supplemented with any of the four sera. TRH gene e xpression did not occur in medium supplemented with charcoal-treated F CS or ATG rat serum. Pretreatment with 10(-8) M Dex caused a significa nt increase in basal as well as cAMP- or phorbol ester-stimulated rele ase of the peptide. Steady state pro-TRH mRNA levels rose 6.8- and 4.2 -fold (both P<0.01) after treatment with 10(-8) M Dex for 4 and 12 day s, respectively. In situ. hybridization experiments revealed that this rise in pro-TRH mRNA levels was probably the result of an increase in the number of AP cells expressing pro-TRH. Both Dex and Cort caused a dose-dependent increase in TRH accumulation, but Cort was approximate ly 40 times less potent than Dex. These results indicate that GC stimu late TRH gene expression in cultured AP cells. The presence of GC in c ulture medium is a prerequisite for the occurrence of TRH gene express ion in the AP. As GC have been reported to reduce pro-TRH mRNA levels in the hypothalamus in vivo, our results may provide an example of the tissue-specific effects of GC on TRH gene expression.