IMMUNOCHEMICAL DEMONSTRATION OF THE MINERALOCORTICOID RECEPTOR IN OCULAR-TISSUES

We studied the presence of the mineralocorticoid receptor (MCR) in the eye with the aid of a number of immunochemical techniques. Immunoblot ting with a polyclonal antibody, directed against the rat renal MCR, r evealed a single band of about 102 kD in extracts prepared from whole bovine or rat retina similar to that observed in cytosol from the kidn ey and myocardium from these species. Isolated cells of the bovine ret inal pigment epithelium (RPE) similarly exhibited a 98- to 102-kD band in Western blots developed with the aid of anti-MCR antiserum. The 98 - to 102-kD band was also obtained following autoradiography of RPE cy tosol irradiated in the presence of H-3-R 5020. This fluorographic pat tern was abolished when RU 26752, an antagonist specific to the MCR, w as allowed to compete with radiolabelled promegestone. The MCR-H-3-RU 26752 complex in RPE cytosol underwent heat activation, as judged by b inding to DNA cellusose, and could also be precipitated by anti-MCR Ig G. In primary cultures, the proliferation of the RPE cells was inhibit ed by the two MCR-specific antagonists RU 26752 and ZK 91587. The loss of the MCR-specific immunofluorescence in RPE cells after only 3 pass ages in culture was associated with refractoriness to the inhibitory e ffect of both of these spironolactones. Immunohistochemistry, using MC R-specific antiserum, revealed strong fluorescence in specific areas o f the rat eye. In the retina, immunopositivity was observed in Muller cells, external and internal limiting membranes, the vitreous base lin ing and in the pigment epithelium. Epithelial cells of the ciliary bod y, iris and cornea also exhibited strong MCR-specific immunofluorescen ce. Thus, both the epithelial and the nonepithelial compartments of th e ocular tissues form interesting new targets to delineate the mechani sm of action of mineralotropic hormones.