Characterization of receptors for the atrial natriuretic factor in rat renal microvessels


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Abstract

Objective:To characterize [125l]-atrial natriuretic factor [ANF-(99-126)l binding sites in the renal preglomerular microvasculature of Sprague-Dawley rats.Methods:Renal preglomerular microvessels were isolated by infusion of a magnetized iron oxide solution into the renal arteries and detachment from non-vascular tissue by a magnetic field. In order to characterize [125l]-ANF-(99-126) binding sites, saturation and competitive binding experiments were performed. To evaluate the proportions of ANF receptor subtypes (ANF-R1, ANF-R2), competition curves were charted in the presence of 10-6mol/l C-ANF-(4-23), a specific ligand of ANF-R2 (ANP-C).Results:[25I]-ANF binding to vascular membranes was saturable and of high affinity. Equilibrium saturation binding curves suggested the presence of one group of high-affinity receptors [equilibrium dissociation constant (Kd) 22±6pmol/l; binding capacity (Bmax) 118±6fmol/mg protein]. In competitive inhibition experiments, no significant differences were found in binding capacity between experiments performed either in the presence or in the absence of an excess (1 u,mol/l) of C-ANF (94 ± 27 versus 151±35fmol/mg protein, respectively), suggesting that most receptors in the renal vasculature are of the subtype ANF-R1. Incubation of renal microvessels with ANF-(99-126) stimulated cyclic GMP production in a dose-related manner. In parallel studies, the proportion of ANF-R1 (ANP-A, -B) and ANF-R2 (ANP-C) receptors in glomeruli, calculated from competitive inhibition experiments, was 86 ± 2 and 14 ±2%, respectively (P<0.005).Conclusions:These results indicate that rat renal preglomerular microvessels contain a high proportion of guanylate cyclase-coupled ANF-R1 (ANP-A, -B) and a low density of ANF-R2 (ANP-C) receptors. This difference in the proportion of ANF receptor subtypes, compared to that reported in glomeruli and other vascular beds, may have physiological significance.

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