Radioligand Binding Characteristics of the Endothelin Receptor in the Rabbit Iris

Abstract

We previously suggested the presence of functionally atypical endothelin (ET) A receptors in the rabbit iris sphincter. Here, we further characterized the ET receptor by a radioligand-receptor binding study utilizing a membrane fraction of the rabbit iris. In addition, we functionally confirm the presence of an atypical ET_A receptor in the iris dilator similar to that in the iris sphincter. In binding experiments, [¹²⁵I]ET-1 was completely displaced by ET-3 in a biphasic fashion, but only partially by BQ-123 and ET_B ligands. In the presence of RES-701, ET-3 and sarafotoxin (SRTX)-b completely displaced [¹²⁵I]ET-1 in a monophasic fashion, but with shallow slopes. Moreover, ET-1, ET-3 and SRTX-b completely displaced [³H]BQ-123 with IC₅₀ values of 0.8, 81 and 4.4 nM, respectively, but with slopes of ET-3 and SRTX-b being again shallow. In iris dilator muscles, ET-3 showed lower and SRTX-b showed higher contractile activities than ET-1. SRTX-c was inactive. BQ-123 more preferentially antagonized ET-3 and SRTX-b than ET-1, with the Schild plot slope of SRTX-b being shallow. Thus, functional experiments suggested the presence of atypical ET_A receptors in the iris dilator similar to the iris sphincter. However, the binding experiments suggested the presence of rather typical ET_A- and ET_B-like receptors. Therefore, we apparently failed to show ET binding sites corresponding to functionally atypical ET_A receptors.