Primary cultures of rat glial cells were established from newborn rat forebrains.
A mixed population of oligodendrocytes and astrocytes was obtained, as confirmed by
indirect immunofluorescence staining with specific markers for each cell type. Receptors
were measured 3 weeks after primary culture in glial cells cultured in the presence
or not of 50 nM estradiol and we have identified progesterone, glucocorticoid, estrogen,
and androgen receptors (PR, GR, ER and AR), but only PR was inducible by the estrogen
treatment. This estrogen-induction of PR was more dramatic in glial cells derived
from female offsprings than from males, as measured by binding studies and by immunohistochemical
techniques with the KC 146 anti-PR monoclonal antibody. The antiestrogen tamoxifen
inhibited the estrogen induction, but had no effect by itself on PR concentration.
Specific binding sites for PR, GR, ER and AR were measured by whole cell assays after
labeling cells with, respectively, [3H]R5020, [3H]dexamethasone, [3H]OH-tamoxifen
or [3H]R1881. PR and GR were also analyzed by ultracentrifugation and after exposure
of cells to agonists, both receptors were recovered from cytosol as a 9S form, and
from the nuclear high-salt, tungstate ions-containing fraction as a 4-6S form. In
contrast, when the antiprogestin- and antiglucocorticosteroid RU486 was used as a
ligand, a non-activated 8.5S receptor complex was found for both receptors in this
nuclear fraction. The 8.5S complex of the GR was further analyzed in the presence
of specific antibodies and, in addition to GR, the presence of the heat shock protein
hsp90 and of a 59 kDa protein was found. During primary culture, the effects of progesterone
(P) and estradiol (E2) were tested on glial cell multiplication, morphology and differentiation.
Cell growth was inhibited by P and stimulated by E2. Both hormones induced dramatic
morphologic changes in oligodendrocytes and astrocytes and increased synthesis of
the myelin basic protein in oligodendrocytes and of the glial fibrillary acidic protein