Localization of Physiological Receptor Sites for Aldosterone in the Bladder of the Toad,Bufo tnarinus

Abstract

Nuclei were isolated from the mucosal epithelial cells of the toad (B. marinus) bladder by 3 methods using: citric acid and Tween-80, 0.17M sucrose and Triton X-100 and 2.2 M sucrose. Binding studies, using a competitive displacement technique, were carried out using these nuclear isolation methods. Nuclei separated by the detergent methods gave no evidence of bound aldosterone, but nuclei isolated in 2.2 M sucrose were shown to contain bound hormone. Binding studies using other mineralocorticoids, an aldosterone antagonist and inactive steroids as displacers, were carried out to demonstrate the physiological activity of the nuclear receptor sites. 9[alpha]-Fluorocortisol, deoxycorticosterone, cortisol and spirolactone SC 14266 all displaced 3H-d-aldosterone from its receptor sites. Testosterone and cholesterol gave no indication of displacement ability. The isolated nuclei contained a large fraction of the aldosterone bound in the whole tissue.