Angiotensin II receptor isoforms in the rat adrenal gland: studies with the selective subtype antagonists DuP 753 and CGP42112A.
The angiotensin II (Ang II)-binding sites in rat adrenal gland membranes were characterized using 125I-radiolabelled Ang II. While Scatchard analysis identified a single population of Ang II receptor sites, isoelectric focusing (IEF) on polyacrylamide gels revealed four peaks of specific Ang II binding which migrated to isoelectric points (pI values) 6.8, 6.7, 6.5 and 6.3. In binding assays in the presence of an excess of the Ang II receptor AT1 subtype antagonist DuP 753, a monophasic dose-dependent displacement of 125I-labelled Ang II binding by the Ang II receptor AT2 subtype antagonist CGP42112A was observed, and vice versa. In this system, reduction of disulphide bridges using 1 mmol dithiothreitol (DTT)/l markedly increased the number of binding sites in the adrenal zona glomerulosa without affecting receptor affinity. Using IEF, it was found that both DuP 753 and CGP42112A were able to reduce specific binding of each of the four peaks to some extent. However, the predominant effect of DuP 753 was to reduce the labelling of the isoform at pI 6.7 substantially, while CGP42112A significantly inhibited the specific 125I-labelled Ang II binding to the pI 6.3 isoform. When DuP 753 and CGP42112A were used together, specific binding of 125I-labelled Ang II to the isoforms of pI values 6.8, 6.7 and 6.3 was completely eliminated. These data suggest that the four peaks of specific binding found may be composed of different isoforms of both AT1 and AT2 receptor subtypes and that the Ang II receptor isoforms which migrated to pI 6.7 and pI 6.3 are predominantly composed of AT1 and AT2 receptor subtypes respectively.(ABSTRACT TRUNCATED AT 250 WORDS)
Montiel
M
M
Department of Biochemistry and Molecular Biology, University of Malaga, Spain.
Barker
S
S
Vinson
G P
GP
Jiménez
E
E
D000311:Adrenal Glands / Q000378:metabolism*; D000804:Angiotensin II / Q000037:antagonists & inhibitors / Q000378:metabolism*; D057911:Angiotensin Receptor Antagonists; D000818:Animals; D001345:Autoradiography; D001667:Binding, Competitive; D001713:Biphenyl Compounds / Q000494:pharmacology*; D007093:Imidazoles / Q000494:pharmacology*; D007457:Iodine Radioisotopes; D007525:Isoelectric Focusing; D007700:Kinetics; D019808:Losartan; D008297:Male; D009842:Oligopeptides / Q000494:pharmacology*; D051381:Rats; D017208:Rats, Wistar; D011945:Receptors, Angiotensin / Q000302:isolation & purification / Q000378:metabolism*; D013777:Tetrazoles / Q000494:pharmacology*
D057911:Angiotensin Receptor Antagonists; D001713:Biphenyl Compounds; D007093:Imidazoles; D007457:Iodine Radioisotopes; D009842:Oligopeptides; D011945:Receptors, Angiotensin; D013777:Tetrazoles; D000804:Angiotensin II; C060894:CGP 42112A; D019808:Losartan