Examination of Steroid-Induced Changes in LHRH Gene Transcription Using ³³P- and ³⁵S-Labeled Probes Specific for Intron 2

We previously demonstrated that estrogen, but not progesterone, induces an increase in LHRH messenger RNA levels between 0800 and 1200 h in neurons of the rostral preoptic area/organum vasculosum of the lamina terminalis (rPOA/OVLT). To determine whether these changes are a result of increased transcription, we used in situ hybridization histochemistry to examine the temporal effects of estradiol (E₂) and progesterone on cellular levels of intron 2-containing LHRH heteronuclear RNA transcripts. To make our method sufficiently sensitive to detect changes in these levels, we used ³³P/³⁵S probes to minimize both background and grain stacking over labeled cell nuclei. We found that levels of LHRH heteronuclear RNA rose between 0800 and 1000 in OVX, E₂-treated rats, remained elevated at 1200 h, and declined by 1400 h. Progesterone did not affect these E₂-induced changes. Our results suggest that an E₂-dependent signal initiates LHRH gene transcription in neurons of the rPOA/OVLT hours before LHRH release begins and that progesterone does not affect this signal.