Localization and quantitation of active renin in monkey kidney by radioinhibitor binding and in vitro autoradiography.
We developed an in vitro autoradiographic method to localize and quantify active renin in primate tissues. Active renin in monkey kidney sections was labeled with the primate specific renin inhibitor, 3H-CGP29287, and quantitated with autoradiography and computerized densitometry. Microscopic emulsion autoradiography was carried out to clarify the detailed localization of the binding. Non-specific binding to aspartyl proteases other than renin was blocked using 1 mumol/L of N-acetyl-pepstatin. To assess the usefulness of this procedure, binding of 3H-CGP29287 was examined both by film and emulsion autoradiography in the kidneys of monkeys (Macaca fuscata) that were given chronically either an angiotensin converting enzyme inhibitor (trandolapril), an angiotensin II receptor antagonist (E4177), or vehicle. 3H-CGP29287 was found to bind very selectively to the juxtaglomerular apparatus (JGA) under control conditions. In monkeys treated with trandolapril or E4177, 3H-CGP29287 binding was increased in proportion to the increase in renal renin concentration determined enzymatically; in these kidneys, emulsion autoradiography revealed radioinhibitor binding extending far from the JGA. The potency of a series of unlabeled renin inhibitor in competing for 3H-CGP29287 binding in the autoradiographic system closely paralleled their potencies, as determined in inhibiting renin by an enzymatic assay. This technique permits specific labeling of the catalytic site of renin in the monkey kidney sections.
Song
K
K
Department of Pharmacology, Osaka Medical College, Japan.
Miyazaki
M
M
Okunishi
H
H
Ishii
K
K
Takai
S
S
Shiota
N
N
Kim
S
S
Mendelsohn
F A
FA
D000806:Angiotensin-Converting Enzyme Inhibitors / Q000494:pharmacology; D000818:Animals; D001345:Autoradiography / Q000379:methods*; D001665:Binding Sites; D015773:Enalaprilat / Q000494:pharmacology; D005260:Female; D007093:Imidazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D007606:Juxtaglomerular Apparatus / Q000378:metabolism; D007668:Kidney / Q000378:metabolism*; D007672:Kidney Cortex / Q000378:metabolism; D007679:Kidney Medulla / Q000378:metabolism; D008251:Macaca; D008297:Male; D009842:Oligopeptides / Q000378:metabolism*; D010436:Pepstatins / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D011869:Radioligand Assay; D012083:Renin / Q000378:metabolism*
D000806:Angiotensin-Converting Enzyme Inhibitors; D007093:Imidazoles; D007211:Indoles; D009842:Oligopeptides; D010436:Pepstatins; D011725:Pyridines; C014195:Streptomyces pepsin inhibitor; C083778:3-((2'-carboxybiphenyl-4-yl)methyl)-2-cyclopropyl-7-methyl-3H-imidazo(4,5-b)pyridine; C052035:trandolapril; C045501:CGP 29287; D012083:Renin; D015773:Enalaprilat