Thyrotropin-Releasing Hormone Gene Expression in Cultured Anterior Pituitary Cells: Role of Gender

Abstract

The present studies were undertaken to investigate the effect of gender on thyrotropin-releasing hormone (TRH) gene expression in cultured anterior pituitary (AP) cells. AP cells derived from 15-day-old male, female, or female pups that had been neonatally treated with testosterone propionate (TP), were cultured for up to 18 days in a modified DMEM/L-15 medium containing 10% fetal calf serum. TRH and AP hormones including GH, prolactin (PRL), luteinizing hormone (LH) and thyrotropin (TSH) were measured by RIA, proTRH mRNA was determined by in situ hybridization using a full-length riboprobe followed by quantification with a computer-assisted image analysis system. Cultures derived from female rats contained significantly (p < 0.01) higher amounts of TRH and secreted approximately twice (p < 0.01) as much TRH under basal conditions and in response to activators of the protein kinase A and C pathways, respectively. In situ hybridization studies revealed that ‘female’ cultures contained significantly higher amounts of proTRH mRNA compared to ‘male’ cultures. Computer-assisted image analysis demonstrated that proTRH mRNA levels were 3.5 times higher in ‘female’ compared to ‘male’ cultures (p < 0.01), an effect that was the result of a significantly higher number (3 times; p < 0.01) of cells expressing proTRH mRNA in ‘female’ cultures. Neonatal TP treatment did not affect either proTRH mRNA or TRH peptide levels. In vitro testosterone treatment resulted in a moderate rise (p < 0.05) of intracellular TRH accumulation in cultures from both sexes, however, proTRH mRNA levels remained unchanged. Gender-specific differences were also found in the contents of all AP hormones measured: GH and TSH were significantly higher in ‘male’ cultures, while ‘female’ cultures contained larger amounts of LH and PRL. The results show that gender determines the level of TRH gene expression in cultured AP cells. Neonatal androgen exposure does not appear to be a determinant in the sex-specific differences observed.

Paracrinicity: The Story of 30 Years of Cellular Pituitary Crosstalk

“…In culture (from 2-week-old rats), TRH expression was stimulated by thyroid hormone and potentiated by glucocorticoids ( 426 ), whereas TRH gene expression was undetectable in cultures deprived of glucocorticoid ( 427 ). Cultures from female rats contained and secreted higher amounts of TRH than those from males ( 428 ).…”

Hypothalamic Prolactin Receptor Messenger Ribonucleic Acid Levels, Prolactin Signaling, and Hyperprolactinemic Inhibition of Pulsatile Luteinizing Hormone Secretion Are Dependent on Estradiol

“…Indeed, pro-TRH-derived peptides have been characterized in long-term cultures of anterior pituitary cells (232), while pro-TRH mRNA has been found in a subpopulation of somatotrophs (234) in which the gene expression is regulated coordinately with the growth hormone gene by glucocorticoids ( 233) and thyroid hormone (235). TRH appears to be most abundant in anterior pituitary glands obtained from 15-day-old female rats (235,236). Although TRH is found in pituitary cell cultures, the peptide from this source apparently does not affect prolactin synthesis or secretion (237).…”

Thyrotropin-Releasing Hormone Gene Expression in the Anterior Pituitary. IV. Evidence for Paracrine and Autocrine Regulation

“…Briefly, AP tissue was separated from posterior/intermediate lobes, collected into sterile HBSS, enzymatically dispersed with neutral protease (1.5 U/AP; Sigma Chemical Co., St. Louis, MO), and plated in a monolayer on 16-mm wells (peptide or pituitary hormone measurements) or 35-mm wells (mRNA determination) at a density of 1000 cells/mm 2 . The cells were cultured for up to 16 days in a modified L-15/DMEM (Life Technologies, Grand Island, NY) medium (11,12,20) containing 10% FCS (Life Technologies).…”