Antibodies to the estrogen receptor-α modulate rapid prolactin release from rat pituitary tumor cells through plasma membrane estrogen receptors

Andrea M. Norfleet, Charlotte H. Clarke, Bahiru Gametchu, Cheryl S. Watson

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    100 Scopus citations


    Antibodies (Abs) raised against the estrogen receptor-α (ERα) were used to investigate the role of ERα proteins located at the plasma membrane in mediating the rapid, estrogen-stimulated secretion of prolactin (PRL) from rat pituitary GH3/B6/F10 cells. Exposure of the cells to 1 nM 17β-estradiol (E2) significantly increased PRL release after 3 or 6 min. When ERα Abs that bind specifically to ERα but are too large to diffuse into cells were tested for activity at the cell membrane, Ab R4, targeted to an ERα hinge region sequence, increased PRL release in a time- and concentration-dependent fashion. Ab H151, directed against a different hinge region epitope, decreased PRL release and blocked the stimulatory action of E2. Abs raised against the DNA binding domain (H226) or the carboxyl terminus (C542) were not biologically active. When each Ab was examined for recognition of ERα on the cell surface by immunocytochemistry, all except H151 generated immunostaining in aldehyde-fixed cells. In live cells, however, Ab H151 but not Ab R4 blocked the membrane binding of fluorescently tagged E2-BSA. Overall, the data indicate that plasma membrane ERα proteins mediate estrogen-stimulated PRL release from GH3/B6/F10 cells. These results may also convey information about conformationally sensitive areas of the membrane form of ERα involved in rapid, nongenomic responses to estrogens.

    Original languageEnglish (US)
    Pages (from-to)157-165
    Number of pages9
    JournalFASEB Journal
    Issue number1
    StatePublished - 2000



    • Immunocytochemistry
    • Membrane steroid receptors
    • Nongenomic effects

    ASJC Scopus subject areas

    • Agricultural and Biological Sciences (miscellaneous)
    • Biochemistry, Genetics and Molecular Biology(all)
    • Biochemistry
    • Cell Biology

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