Rat vascular tissue contains a neutral endopeptidase capable of degrading atrial natriuretic peptide.
Neutral endopeptidase (NEP, EC 3.4.24.11), purified from renal brush border, cleaves the Cys7-Phe8 amide bond of rat atrial natriuretic peptide (ANP), to generate the inactive metabolite (ANP cleaved at the Cys7-Phe8 bond; x-ANP). To determine if NEP contributes to the inactivation of circulating ANP, we investigated the degradation of rat ANP (rANP, 1-28) in the vasculature. Formation of x-ANP from exogenous ANP was studied in a mesenteric arterial preparation by perfusion in a single pass system in the presence and absence of the NEP inhibitors, thiorphan or phosphoramidon. In addition, a purified membrane fraction was prepared from mesenteric arterial homogenate and compared with an equivalent renal membrane fraction. Formation of x-ANP was quantified by a specific immunoassay (ELISA). Renal and vascular membranes shared the same pH optima for x-ANP formation (pH 7.5), although x-ANP generation was considerably greater in renal vs. vascular membranes (31.6 and 0.4 nmol min-1 mg-1 of protein, respectively). Both preparations were inhibited in a similar, dose-dependent manner by phosphoramidon, thiorphan or a polyclonal antibody to NEP. In perfused mesenteric arteries, 1.6 +/- 0.8 pmol of x-ANP were generated from 1 microgram of ANP; this formation was inhibited 51% by 10 microM phosphoramidon or thiorphan. Plasma levels of x-ANP after bolus i.v. administration of rANP in rats, were inhibited (70-80%) by thiorphan at comparable doses to those used in perfused mesenteric arteries. These studies indicate that ANP is degraded in the vasculature by NEP or an "NEP-like" enzyme(s).
Tamburini
P P
PP
Unigene Laboratories, Inc., Fairfield, New Jersey.
Koehn
J A
JA
Gilligan
J P
JP
Charles
D
D
Palmesino
R A
RA
Sharif
R
R
McMartin
C
C
Erion
M D
MD
Miller
M J
MJ
D000818:Animals; D009320:Atrial Natriuretic Factor / Q000378:metabolism*; D001808:Blood Vessels / Q000201:enzymology*; D010450:Endopeptidases / Q000032:analysis / Q000502:physiology*; D005260:Female; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008297:Male; D011817:Rabbits; D051381:Rats; D011919:Rats, Inbred Strains; D015244:Thiorphan / Q000494:pharmacology
D009320:Atrial Natriuretic Factor; D015244:Thiorphan; D010450:Endopeptidases