title: Human endothelin receptors characterized using reverse transcriptase-polymerase chain reaction, in situ hybridization, and subtype-selective ligands BQ123 and BQ3020: evidence for expression of ETB receptors in human vascular smooth muscle.
abstract: Our aim was to characterize and determine the function of endothelin (ET) receptor subtypes in human vascular tissue. Reverse transcriptase-polymerase chain reaction with nested oligonucleotide primers detected the presence of mRNA encoding both ETA and ETB receptors in the media from aorta and pulmonary and coronary arteries. In situ hybridization confirmed the presence of mRNA for both subtypes in the media of coronary arteries. Saturation binding assays using 125I-ET-1 found a single population of high-affinity ET receptors (n = three patients, +/- SEM) in aorta (Kd = 0.507 +/- 0.020 nM; Bmax = 9 +/- 4 fmol/mg protein) and pulmonary (Kd = 0.845 +/- 0.245 nM; Bmax = 15 +/- 10 fmol/mg protein) and coronary arteries (Kd = 0.141 +/- 0.020 nM; Bmax = 71 +/- 21 fmol/mg protein). Using media from coronary arteries, the ETA-selective ligand BQ123 (cyclo[D-Asp-L-Pro-D-Val-L-Leu-D-Trp]) and the ETB-selective ligand BQ3020 (Ala11,15-Ac-ET-1[6-21]) both produced biphasic competition binding curves against 125I-ET-1, confirming the presence of high- and low-affinity sites corresponding to the two subtypes: BQ123 (KdETA = 0.85 +/- 0.03 nM; KdETB = 7.58 +/- 2.27 microM; ETA/ETB, 87%:13%) and BQ3020 (KdETA = 0.22 +/- 0.04 microM; KdETB = 0.77 +/- 0.34 nM; ETA/ETB, 62%:38%). BQ123 (0.1 microM) caused a significant parallel rightward shift of ET-1-induced vasoconstriction of coronary arteries in vitro, but BQ3020 and Ala1,3,11,15-ET-1 failed to show any agonist activity when tested at concentrations of < or = 3 microM in three vessels.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Clinical Pharmacology Unit, University of Cambridge, Addenbrooke's Hospital, England.
authors.forename: A P
authors.initials: AP
authors.lastname: Davenport
authors.forename: G
authors.initials: G
authors.lastname: O'Reilly
authors.forename: P
authors.initials: P
authors.lastname: Molenaar
authors.forename: J J
authors.initials: JJ
authors.lastname: Maguire
authors.forename: R E
authors.initials: RE
authors.lastname: Kuc
authors.forename: A
authors.initials: A
authors.lastname: Sharkey
authors.forename: C R
authors.initials: CR
authors.lastname: Bacon
authors.forename: A
authors.initials: A
authors.lastname: Ferro
mesh_terms: D000328:Adult; D000595:Amino Acid Sequence; D001345:Autoradiography; D001483:Base Sequence; D003331:Coronary Vessels / Q000187:drug effects / Q000378:metabolism; D004587:Electrophoresis, Agar Gel; D065128:Endothelin Receptor Antagonists; D016232:Endothelins / Q000494:pharmacology; D006801:Humans; D017403:In Situ Hybridization; D007457:Iodine Radioisotopes; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D011651:Pulmonary Artery / Q000187:drug effects / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry; D017466:Receptors, Endothelin / Q000737:chemistry* / Q000187:drug effects
chemical_list: D065128:Endothelin Receptor Antagonists; D016232:Endothelins; D007457:Iodine Radioisotopes; D010446:Peptide Fragments; D010456:Peptides, Cyclic; D012333:RNA, Messenger; D017466:Receptors, Endothelin; C075771:BQ 3020; D012194:RNA-Directed DNA Polymerase; C072247:cyclo(Trp-Asp-Pro-Val-Leu)