Effects of Estrogen on Primary Ovine Pituitary Cell Cultures: Stimulation of Prolactin Secretion, Synthesis, and Preprolactin Messenger Ribonucleic Acid Activity*

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Abstract
Estrogen regulation of PRL synthesis and secretion was studied in primary ovine pituitary cell cultures. Medium PRL, [3H]leucine incorporation into PRL, and pre-PRL messenger RNA (mRNA) activity were determined in cultures incubated in control or 17β-estradiol (E2)-containing medium. We consistently observed a significant increase in medium PRL after 2 days of E2 treatment. A maximal response of 150–200% of control was reached after 6–8 days of E2. The response increased with increasing doses of E2 in the physiological range (10-9-10-8 M) and was maximal at about 10-9 M E2. Sodium dodecyl sulfate gel electrophoresis of the soluble proteins labeled by a 1-h incubation in radioactive leucine showed that the synthesis of only one major protein band which comigrated with authentic ovine PRL was stimulated. Immunoprecipitation of the labeled cell homogenates confirmed that PRL synthesis was increased. Five days of E2 increased medium PRL to 150% of controls, PRL synthesis to 183% of controls, and pre-PRL mRNA activity to 206% of controls. Progesterone, testosterone, and corticosterone had little effect. However, progesterone suppressed the E2-induced increase in medium PRL, PRL synthesis, and pre-PRL mRNA. A time-course study showed that 24 h were required for the increase in PRL synthesis and pre-PRL mRNA. Medium PRL accumulation was increased after 48 h. We conclude that PRL synthesis can be directly stimulated in primary cell culture by physiological levels of E2. The effect is specific for E2 and its expression requires a minimum of 24 h.

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