Evidence for specific progesterone receptors in rat brain cytosol

Cytosol prepared from rat brains perfused with saline possesses saturable macromolecular components which form unstable complexes with [3H]progesterone in vitro. The components can be distinguished from the serum protein corticosteroid-binding globulin (CBG) which also binds progesterone. Unlike CBG, the cytosol components did not bind corticosterone and were more unstable in the presence of Sephadex LH-20 than was CBG. Furthermore, the components were precipitated by a concentration of ammonium sulphate in which CBG is soluble. Unlabelled progesterone and testosterone competed equipotently with [3H)progesterone, and 5 alpha-pregnane-3,20-dione and deoxycorticosterone competed to a lesser extent, whereas oestradiol, corticosterone and 11 beta-hydroxy-pregn-4-en-3,20-dione were not bound. Although present in all brain tissues examined and in uterine and anterior pituitary tissue, the saturable binding was not detectable in cytosols from the spleen or diaphragm. Complete exchange occurred in the incubation at 0 degrees C between [3H)progesterone and unlabelled progesterone and the saturable binding sites and the reaction attained apparent equilibrium within 2 h at this temperature. An incubation temperature of 30 degrees C resulted in an almost complete loss of saturable binding. Scatchard plots obtained from binding isotherms were curvilinear and yielded at least three dissociation constants (Kd), two of higher affinity (Kd congruent to 10(-8) mol/l) and one about sixty times lower. From these results it is concluded that rat brain cytosol possesses progesterone-selective components which fulfil some of the criteria required for steroid hormone receptors.