Estradiol Modulates Protein Kinase C Activity in the Rat Pituitary in Vivo and in Vitro

Abstract

17.beta.-Estradiol (E2) alters different functions of pituitary cells, including cell sensitivity to several neurohormones such as LHRH, TRH, somatostatin, or dopamine, presumbly by affecting receptor coupling mechanisms. Attempting to pinpoint the membrane processes underlying this modulation, we studied the effect of E2 on pituitary kinase-C (PKC) activity, a major signal transduction enzyme. The distribution of calcium-and phospholipid-dependent partially purified PKC (chromatography on DEAE-52 cellulose columns) was evaluated in membrane and cytosol fractions from anterior pituitaries of ovariectomized (OVX) or OVX plus E2-treated rats. E2 administration by implants to OVX animals increased significantly both soluble and particulate enzyme activity. The effect increased progressively from 24 h to 5 days after E2 treatment. Administration of 17.alpha.-estradiol, an inactive stereoisomer of E2, was ineffective, pointing to stereospecific interactions. Total destruction of neural connections to the pituitary (complete hypothalamic lesions) did not modify the enzyme response to E2 administration, indicating a direct effect of the steroid on pituitary PKC activity. A direct E2 (10-9 M) effect was confirmed in primary mixed cultures of pituitary cells; it was time dependent (15-96 h) and specific, and reflects a genomic E2 action. E2 treatment for shorter times had no effect on the enzyme levels or the membrane redistribution of PKC activity. In contrast, under the same experimental conditions phorbol esters (12-O-tertadecanoyl-phorbol-13-acetate (TPA)) induced a rapid and sustained translocation of the enzyme. PKC activity was found in all pituitary cell types, with maximal activity in fractions of gonadotropes and thyrotropes, as evaluated in cultures enriched in certain types of pituitary cells separated by means of unit gravity gradient sedimentation. E2 treatment (10-9 M; 72 h) significantly increased both soluble and particulate enzyme levels in all cell types. In addition, administration of E2 (10-9 M; 72 h) to cell cultures strongly increased the TPA-evoked LH and PRL release. These results indicate that E2-induced changes in pituitary function include selective effects of the steroid on PKC activity involved at different levels in the coupling mechanisms.