Distinct specificity for corticosteroid binding sites in amphibian cytosol, neuronal membranes, and plasma

To address mechanisms of corticosteroid action, one needs tools for disting uishing between the major classes of corticosteroid binding sites: neuronal membrane-associated receptors, intracellular ligand-activated transcriptio n factors, and corticosteroid binding globulins (CBG) in plasma. We charact erized the binding parameters for three classes of binding sites in an amph ibian Ambystoma tigrinum, and found that each class had a distinct pharmaco logical specificity. Equilibrium saturation and kinetic experiments indicat ed that [H-3]corticosterone binds to neuronal membranes with high affinity (K-d approximate to 0.37 nM). Aldosterone and two synthetic ligands for mam malian intracellular receptors, dexamethasone and RU486, displayed low affi nity for brain membrane sites. In cytosol prepared from brain and liver, [H -3]corticosterone bound to a single class of receptors with high affinity ( K-d approximate to 0.75 and 4.69 nM, respectively) and the rank order poten cies for steroid inhibition of [H-3]corticosterone binding was RU486 > dexa methasone approximate to corticosterone > aldosterone. In kidney and skin c ytosol, [3H]corticosterone binding was best fit with a model having a high- affinity and a lower-affinity site; these sites are not consistent with the pharmacology of mammalian Type I (MR) and Type II (GR) receptors. [H-3]Cor ticosterone also bound to presumed CBG in plasma with high affinity (K-d ap proximate to 2.7 nM), but dexamethasone and androgens bound to plasma CBG w ith equivalently high affinity. These data demonstrate that pharmacological specificity can be a useful tool for distinguishing corticosteroid binding to different classes of binding sites. These data also indicate that there may be marked species differences in the specificity of corticosteroid bin ding sites. (C) 2000 Academic Press.