title: Chemical differentiation of type I and type II receptors for adrenal steroids in brain cytosol.
abstract: Studies outlined here compare the properties of mineralocorticoid (Type I) and glucocorticoid (Type II) receptors in cytosol from adrenalectomized mouse brain. Pretreating cytosol with dextran-coated charcoal (DCC) produced a 4.7-fold increase in the subsequent macromolecular binding of the mineralocorticoid, [3H]aldosterone (20 nM ALDO, in the presence of a 50-fold molar excess of the highly specific synthetic glucocorticoid, RU 26988), whereas it produced a 55% decrease in the binding of the glucocorticoid, [3H]triamcinolone acetonide (20 nM TA). Scatchard analyses revealed that DCC pretreatment had no effect on the affinity or maximal binding of Type I receptors for [3H]ALDO (in the presence of a 0-, 50- or 500-fold excess of RU 26988), whereas it produced a 3- to 6-fold increase in the Kd, and an 8-43% decrease in the maximal binding, of Type II receptors for [3H]TA and [3H]dexamethasone. Optimal stability of unoccupied Type I receptors at 0 degree C was found to be achieved in buffers containing glycerol, but lacking molybdate. Although the addition of molybdate was found to reduce the loss in Type I receptor binding observed after incubating unlabelled cytosol at 12 or 22 degrees C, this stabilization was accompanied by a concentration-dependent reduction in the binding of [3H]ALDO at 0 degree C. Scatchard analyses showed that this reduction was due to a shift in the maximal binding, and not the affinity, of the Type I receptors for [3H]ALDO. The presence or absence of dithiothreitol in cytosol appeared to have little effect on the stability of Type I receptors. In contrast to our finding for Type I receptors, it was possible to stabilize the binding capacity of unoccupied Type II receptors, even after 2-4 h at 12 or 22 degrees C, if the glycerol containing buffers were supplemented with both molybdate and dithiothreitol. In summary, these results indicate distinct chemical differences between Type I and Type II receptors for adrenal steroids.
mesh_terms: D000315:Adrenalectomy; D000450:Aldosterone / Q000378:metabolism; D000732:Androstanols / Q000378:metabolism; D001923:Brain Chemistry; D002606:Charcoal; D003600:Cytosol / Q000032:analysis*; D003907:Dexamethasone / Q000378:metabolism; D003911:Dextrans; D004229:Dithiothreitol / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D007700:Kinetics; D008982:Molybdenum / Q000494:pharmacology; D011965:Receptors, Glucocorticoid / Q000032:analysis*; D018161:Receptors, Mineralocorticoid; D011987:Receptors, Steroid / Q000032:analysis*; D014222:Triamcinolone Acetonide / Q000378:metabolism
chemical_list: D000732:Androstanols; D003911:Dextrans; D011965:Receptors, Glucocorticoid; D018161:Receptors, Mineralocorticoid; D011987:Receptors, Steroid; C044659:molybdate; D002606:Charcoal; D000450:Aldosterone; C038341:RU 26988; D003907:Dexamethasone; D008982:Molybdenum; D004492:Edetic Acid; D014222:Triamcinolone Acetonide; D004229:Dithiothreitol