Identification of endothelin receptor subtype (ETB) in human cerebral cortex using subtype-selective ligands.

Specific endothelin (ET) binding sites were characterized in membranes prepared from human cerebral cortices using binding assay and cross-linking analysis. The presence of immunoreactive (IR) ET-1 was studied by radioimmunoassay. Saturation binding experiments revealed that the KD and Bmax for 125I-ET-1 and 125I-ET-3 to membranes from gray matter were 25 +/- 6 pM and 115 +/- 15 fmol/mg of protein and 24 +/- 5 pM and 108 +/- 13 fmol/mg of protein, respectively. Similar results were obtained for white matter. In the presence of 10 nM sarafotoxin-6c, which is selective for ETB receptors, 125I-ET-1 and 125I-ET-3 binding was totally abolished. However, in the presence of 1 microM BQ123, which is selective for ETA receptors, both bindings were not affected. These results suggest that the human cerebral cortex contains only ETB receptors. Cross-linking of 125I-ET-1 and 125I-ET-3 to membranes with disuccinimidyl suberate resulted in the labeling of two bands of 48 and 31 kDa. Concentrations of IR-ET-1 in the gray and white matter were 7.0 +/- 3.2 and 2.5 +/- 1.7 fmol/g wet weight, respectively. The demonstration of high-affinity ETB receptors and the presence of IR-ET-1 suggest that the peptide may act as a neurotransmitter or neuromodulator in the human cerebral cortex.