Tc. Pappas et al., MEMBRANE ESTROGEN-RECEPTORS IN GH3 B6 CELLS ARE ASSOCIATED WITH RAPIDESTROGEN-INDUCED RELEASE OF PROLACTIN/, Endocrine, 2(9), 1994, pp. 813-822
The mechanisms responsible for such rapid actions of estrogen, as the
rapid release of prolactin from CH3/B6 cells, have not yet been elucid
ated, but may be mediated by a membrane form of the estrogen receptor.
Two polyclonal, affinity-purified antibodies generated to a peptide f
rom rat estrogen receptor (amino acids 270-284) specifically recognize
d native and denatured estrogen receptor from GH(3) cells. About 13% o
f live GH(3)/BC cells reacted with affinity-purified antibody at 2 deg
rees C and revealed punctate membrane labeling. Anti-actin controls sh
owed that cells were not permeabilized by the live-labeling procedure.
Nuclear labeling was also evident in cells made permeable with deterg
ent. Both membrane and intracellular labeling were competed with the p
eptide used to generate the antibody. Release of prolactin (measured b
y RIA) by CH3/B6 cells in response to 17 beta-estradiol was as rapid a
s 1 min, maximal at nM 17-beta-estradiol, and specific for estradiol.
Presence of serum in the medium caused a higher basal level of prolact
in release but abolished the estrogen-induced release. Limited dilutio
n cloning of heterogeneous CH3/B6 cells showed that immunocytochemical
presence of estrogen receptor in the plasma membrane could be correla
ted to the rapid response to estrogen.