Transcriptional down-regulation of the rabbit pulmonary artery endothelin B receptor during phenotypic modulation

1 We confirmed that endothelium-independent contraction of the rabbit pulmo nary artery (RPA) is mediated through both an endothelin A (ETAR) and endot helin B (ETB2R) receptor. 2 The response of endothelium-denuded RPA rings to endothelin-1 (ET-1, pD(2 )=7.84 +/- 0.03) was only partially inhibited by BQ123 (10 mu M), an ETAR a ntagonist. 3 Pretreatment with 1 nM sarafotoxin S6c (S6c), an ETBR agonist, desensitiz ed the ETB2R and significantly attenuated the response to ET-3 (pD(2) = 7.4 0 +/- 0.02 before, <6.50 after S6c). 4 Pretreatment with S6c had little effect an the response to ET-1, but BQ12 3 (10 mu M) caused a parallel shift to the right of the residual ETAR-media ted response to ET-1 (pD(2) = 7.84 +/- 0.03 before S6c, 7.93 +/- 0.03 after S6c, 6.81 +/- 0.05 after BQ123). 5 Binding of radiolabelled ET-1 to early passage cultures of RPA vascular s mooth muscle cells (VSMC) displayed two patterns of competitive displacemen t characteristic of the ETAR (BQ123 pIC(50) = 8.73 +/- 0.05) or ETB2R (S6c pIC(50) = 10.15). 6 Competitive displacement experiments using membranes from late passage VS MC confirmed only the presence of the ETAR (ET-1 pIC(50) = 9.3, BQ123 pIC(5 0) = 8.0, S6c pIC(50) < 6.0). 7 The ETAR was functionally active and coupled to rises in intracellular ca lcium which exhibited prolonged homologous desensitization. 8 Using a reverse transcriptase polymerase chain reaction for the rabbit ET B2R, we demonstrated the absence of mRNA expression in phenotypically modif ied VSMC. 9 We conclude that the ETB2R expressed by VSMC which mediates contraction o f RPA is rapidly down-regulated at the transcriptional level during phenoty pic modulation in vitro.