PARTICIPATION OF GUANYLATE CYCLASE, PKC EPSILON AND ERK 1/2 IN ESTRADIOL EFFECTS MEDIATED BY MEMBRANE ESTROGEN RECEPTOR IN LACTOTROPH CELLS

GUTIÉRREZ S.; DE PAUL A.L,; PETITI J.P.; SOSA L.; PALMERI C.; SOAJE M.; MASINI DE REPISO A.; TORRES A.I.

Rio de Janeiro

Congreso; 13 International Congress of Endocrinology; 2008

INTERNATIONAL SOCIETY OF ENDOCRINOLOGY

The aim of the present work was to investigate the Guanylate cyclase (GC), protein kinase C (PKC) epsilon and extracellular-signal regulated kinase (ERK) 1/2 participation in 17-beta estradiol (E2) effects on prolactin secretion and lactotroph proliferation, and to identify membrane estrogen receptors (ER) in lactotroph cells. Primary pituitary cell cultures from female rats were treated with E2, E2 conjugated to bovine serum albumin (E2?BSA) and insulin (In), alone or combined, for 15, 30, 60 min and 24 h. The cultured cells were processed for PRL secretion by RIA, quantification of lactotroph proliferation by BrDU/PRL and protein detection by western blot (PKC epsilon, ERK1/2 phosphorylated (P) and total, GC alpha-1, beta-1 and ß-actin). Membrane ERs were immunodetected with specific antibody against classic nuclear ER alpha and beta. Statistical analysis: ANOVA-Tukey. In serum free conditions, E2 and E2-BSA promoted a differential secretory activity on PRL cells but were unable to induce lactotroph cell proliferation. However, both free and conjugated E2 were competent arresting the mitogenic activity promoted by Ins. E2, E2-BSA and Ins induced opposite effects on GC alpha-1 and beta-1 expression, increasing the GC alpha-1 and inhibing GC beta-1, while the E2/Ins or E2-BSA/Ins co-incubation induced an augmentation of two GC isoforms. E2, E2-BSA and Ins stimuli increased the PKC epsilon and P-ERK1/2 expression, although combined treatments with E2/Ins or E2-BSA/Ins induced a significant reduction in these levels, in close correlation with the decrease of lactotroph cell proliferation. Pre-treatment with bisindolmaleymide I (PKC inhibitor) significantly inhibited ERK1/2 activation promoted by Ins, without modifying P-ERK1/2 expression levels induced by E2 or E2-BSA. By immuno-electron-microscopy, ER alpha was localized on the cell-surface of lactotrophs. No immunostaining for ER beta was detected. These findings show the presence of ER alpha in the plasma membrane of lactotroph cells which acts modulating PRL secretion and lactotroph proliferation through GC, PKC epsilon and ERK1/2.