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Division of Genetics, Department of Medicine, Brigham and Women's Hospital, Howard Hughes Medical Institute, Harvard Medical School Boston, Massachusetts 02115
Address requests for reprints to: Margaret A. Shupnik, Box 511, University of Virginia Health Sciences Center, Charlottesville, Virginia 22908.
Abstract
Our previous work demonstrated that in vivo estradiol (E2) administration to ovariectomized rats suppressed the transcription of the LH subunit genes within 4 h. To determine whether these effects were mediated directly at the level of the gonadotrope, the transcription rates of the LHβ, FSHβ, and
-subunits were measured in short term cultures of pituitary fragments from female rats in various physiological states. In each in vitro experiment, fragments from matched sets of hemipituitaries were used for control and treatment (10–8ME2) groups. In pituitaries from ovariectomized animals, treated in vitro with or without E2, there was no significant effect of 2 h or 6 h of E2 on FSHβ or
-subunit gene transcription, but a consistent 2- to 3-fold stimulation of LHβ mRNA synthesis. In pituitaries from intact randomly cycling rats, E2 in culture had no effect on the transcription rate of
-subunit, FSHβ, or TSHβ genes, but stimulated LHβ and PRL transcription 2- fold. Thyroid hormone treatment specifically suppressed
-subunit (38% of control) and TSHβ (17% of control) mRNA synthesis, indicating that the culture system is responding in an appropriate physiological manner and that decreases in transcription can be easily and accurately measured with this system. Basal and E2-treated transcription rates for each gonadotropin subunit gene were measured as a function of stage of the estrous cycle. Basal transcription of the
-subunit gene did not vary significantly throughout the cycle and did not respond to E2 treatment in vitro. FSHβ mRNA synthesis was lowest at metestrus and diestrus (18–25 ppm), increased slightly on the morning and evening of proestrus (40–50 ppm), and reached its highest level on the morning and evening of estrus (70–80 ppm). However, E2 treatment in vitro had no significant effect on transcription. LHβ mRNA synthesis was lowest at metestrus (25 ppm), increased continuously to its highest level on the evening of proestrus (130 ppm), and returned to low levels by the evening of estrus (35 ppm). At each stage of the cycle, E2 stimulated LHβ transcription 2- to 3-fold. Thus, E2 specifically stimulates the LHβ gene, but not the FSHβ or
-subunit gene, directly at the level of the pituitary cell. The data suggest that the negative regulatory effects of E2 on LHβ mRNA synthesis in vivo may be due to steroid-hypothalamic interactions.
Received for publication November 15, 1988. Revision received December 12, 1988. Accepted for publication December 12, 1988.
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