Identification and characterization of endothelin receptor subtype B in rat retina.
The presence of immunoreactive (IR) endothelin (ET)-1 and ET-1 receptors in rat retina has been studied by radioimmunoassay and receptor assay, respectively. The specific binding of 125I-ET-1 to rat retinal particulate preparations was saturable. Apparent equilibrium conditions were established within 120-140 min. Scatchard analysis of binding data indicated a single class of high-affinity binding sites with a KD of 35 +/- 11 pM and a Bmax of 168 +/- 60 fmol/mg of protein. 125I-ET-1 binding to retinal particulate preparations was not inhibited by 1 microM 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 125I-ET-1 to retinal particulate preparations with disuccinimidyl suberate resulted in the labeling 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 neurotransmitter or neuromodulator in the retina.
de Juan
J A
JA
Centro de Diabetes e Hipertensión, Hospital Universitario San Carlos, Madrid, Spain.
Moya
F J
FJ
Garcia de Lacoba
M
M
Fernandez-Cruz
A
A
Fernandez-Durango
R
R
D000345:Affinity Labels; D000818:Animals; D001667:Binding, Competitive; D003432:Cross-Linking Reagents; D007700:Kinetics; D008297:Male; D011863:Radioimmunoassay; D051381:Rats; D017208:Rats, Wistar; D017466:Receptors, Endothelin / Q000145:classification / Q000378:metabolism*; D012160:Retina / Q000378:metabolism*
D000345:Affinity Labels; D003432:Cross-Linking Reagents; D017466:Receptors, Endothelin