Coexpression Studies with Endothelin Receptor Subtypes Indicate the Existence of Intracellular Cross-Talk between ETA and ETB Receptors.

Human Girardi heart cells expressing endothelin ETB receptors (GHB cells) were transfected with human ETA cDNA, and coexpression of ETA and ETB in the ratio of 4:6 was demonstrated by Scatchard analysis. [125I]Endothelin (ET)-1 binding to ETA-transfected GH cells (GHAB cells) was displaced by an ETA antagonist, BQ-123, in a biphasic manner. An ETB agonist, BQ-3020, and an ETB antagonist, BQ-788, inhibited [125I]ET-1 binding to GHAB cells in a monophasic manner with low affinities (IC50=2,800 and 890 nM, respectively); IC50 values for ETB receptors seemed to be as weak as those for ETA receptors. However, BQ-3020 and BQ-788 had a high affinity for ETB receptors in a binding experiment using [125I]ET-1 in the presence of 1 μM BQ-123, where ETA receptors are masked (IC50 =0.49 and 0.89 nM, respectively). The ETB-mediated increase in intracellular calcium concentrations in GHAB cells was not affected by 0.1 μM BQ-788 alone but was inhibited significantly by the same concentration of BQ-788 in combination with 10 μM BQ-123. ET-1 suppressed forskolin-stimulated accumulation of cAMP through the activation of ETA and ETB in GHAB cells; 1 μM BQ-123 or BQ-788 inhibited the suppression by only 20%, whereas a mixture of BQ-123 and BQ-788 (1 μM each) completely inhibited the cAMP decrease. These findings suggest that the stimulation of ETA receptors with ET-1 results in a lowering of the affinity of BQ-3020 and BQ-788 for ETB receptors in GHAB cells. We conclude that there is intracellular cross-talk between ETA and ETB receptors in GHAB cells. [ABSTRACT FROM AUTHOR]