CHARACTERIZATION OF THE BINDING OF ENDOTHELIN ET(B) SELECTIVE LIGANDSIN HUMAN AND RAT-HEART

1 We determined competition binding characteristics of endothelin ET(B ) receptor selective ligands in human left ventricle and compared thes e values to those obtained with rat left ventricle. Sarafotoxin S6c, E T-3, BQ788 and IRL2500 competed against [I-125]-BQ3020 (ET(B) selectiv e radioligand; Molenaar et al., 1992) with high affinity and against [ I-125]-PD151242 (ET(A) selective radioligand) with low affinity in hum an left ventricle, confirming the ET(B) selectivity of these compounds . 2 ET-3 competed with moderate selectivity for ET(B) over ET(A) recep tors in human left ventricle and with slightly higher selectivity in r at left ventricle (460 and 1,400 fold, respectively). There was a smal l difference in the affinity of ET(A) receptors for ET-3 (K-D ET(A) in human left ventricle = 0.07 +/- 0.02 mu M; K-D ET(A) in rat left vent ricle = 0.27 +/- 0.08 mu M; P = 0.05) but no difference in the affinit y of ET(B) receptors for this ligand (K-D ET(B) in human left ventricl e = 0.15 +/- 0.06 nM; K-D ET(B) in rat left ventricle = 0.19 +/- 0.03 nM). 3 The selectivity of sarafotoxin S6c for ET(B) over ET(A) recepto rs in human left ventricle was 5,900 fold compared with 59,400 fold in rat left ventricle. The affinity of ET(A) receptors for sarafotoxin S 6c was higher in human than in rat left ventricle (K-D ET(A) = 2.00 +/ - 0.20 mu M and 3.50 +/- 0.26 mu M, respectively; P = 0.03), while the affinity of ET(B) receptors for this ligand was higher in rat left ve ntricle (K-D ET(B) = 0.06 +/- 0.02 nM) than in human left ventricle (K -D ET(B) = 0.34 +/- 0.13 nM) (P = 0.02). The affinity of ET(B) recepto rs for sarafotoxin S6c in rat left ventricle determined in the absence or presence of GTP was the same indicating that differing affinity st ates of ET(B) receptors in human and rat left ventricle do not account for the variation observed between species. 4 There was no difference in the affinity of ET(A) receptors for BQ788 (K-D ET(A) = 1.01 +/- 0. 20 mu M and K-D ET(A) = 1.39 +/- 0.35 mu M) or for the novel ET(B) sel ective antagonist, IRL2500 (K-D ET(A) = 30.0 +/- 20.8 mu M and K-D ET( A) = 55.6 +/- 9.93 mu M) in human and rat left ventricle, respectively . ET(B) receptors had a significantly higher affinity for B4788 (K-D E T(B) = 9.8 +/- 1.3 nM and K-D ET(B) = 31.0 +/- 5.4 nM; P = 0.02) and I RL2500 (K-D ET(B) = 78.2 +/- 9.7 nM and K-D ET(B) = 300.0 +/- 75.1 nM; P = 0.03) in human and rat left ventricle, respectively. The syntheti cally synthesized ET(B) selective antagonist RES-701-1 (0.1-3 mu M) fa iled to inhibit [I-125]-ET-1 binding in either tissue. 5 In conclusion , we have compared equilibrium dissociation constants for a number of ET(B) selective compounds in human and rat heart. The affinity of ET(B ) receptors for sarafotoxin S6c, B4788 and IRL2500 differed in human a nd rat left ventricle. No difference in affinity was detected for ET-3 binding at ET(B) receptors. Sarafotoxin S6c binding was unaffected by GTP indicating that the different receptor affinities in human and ra t heart cannot be explained by differing ET(B) receptor affinity state s. This study highlights the need to consider differences in binding c haracteristics that may arise from the use of tissues obtained from di fferent species.