IDENTIFICATION AND CHARACTERIZATION OF ENDOTHELIN RECEPTOR SUBTYPE-B IN RAT RETINA

The presence of immunoreactive (IR) endothelin (ET)-l and ET-1 recepto rs in rat retina has been studied by radioimmunoassay and receptor ass ay, respectively. The specific binding of I-125-ET-1 to rat retinal pa rticulate preparations was saturable. Apparent equilibrium conditions were established within 120-140 min. Scatchard analysis of binding dat a indicated a single class of high-affinity binding sites with a K(D) of 35 +/- 11 pM and a B(max) of 1 68 +/- 60 fmol/mg of protein. I-125- ET-1 binding to retinal particulate preparations was not inhibited by 1 muM concentrations of somatostatin, atrial natriuretic factor, brain natriuretic peptide, thyroid-stimulating hormone, growth hormone, or insulin. The three endothelin isoforms, ET-1, -2, and -3, had similar affinity for the receptor. Cross-linking of I-125-ET-1 to retinal part iculate preparations with disuccinimidyl suberate resulted in the labe ling of two bands with apparent molecular masses of 52 and 34 kDa. We have established a highly sensitive and specific radioimmunoassay for ET-1. The concentration of IR-ET-1 in rat retina was 35 +/- 10 fmol/g wet weight. The demonstration of specific high-affinity ETB receptors and the presence of IR-ET-1 suggest that the peptide may act as a neur otransmitter or neuromodulator in the retina.