Receptor proteins and biological effects of C-type natriuretic peptides in the renal glomerulus of the rat.

Binding studies on rat glomeruli using 125I-labeled Tyr0-C-type natriuretic peptide-(1-22) [125I-Tyr0-CNP-(1-22)] and 125I-labeled alpha-atrial natriuretic peptide (alpha-125I-ANP), and the unlabeled ligands CNP-(1-22), alpha-ANP, and des-Gln18,Ser19,Gly20,Leu21,Gly22-ANP-(4-2 3)-NH2 (C-ANP) suggest that receptor-like sites that bind both alpha-ANP and C-ANP fall into two categories, one with high [dissociation constant (Kd) approximately 10(-9)M] and one with low (Kd approximately 10(-5)M) affinity for CNP-(1-22). Covalent attachment of 125I-Tyr0-CNP-(1-22) and alpha-125I-ANP to these sites identifies two membrane proteins with corresponding properties. The first, which can be labeled by both radioligands, is a disulfide-bridged approximately 140-kDa protein that is reduced by dithiothreitol to approximately 67 kDa. This protein binds C-ANP and has Kd approximately 10(-10) M for CNP-(1-22). The second protein, which is labeled only by alpha-125I-ANP, also binds C-ANP, but has Kd approximately 10(-5)M for CNP-(1-22). This approximately 77-kDa protein may also have a disulfide-bridged, high-molecular-mass form of approximately 140 kDa in the absence of dithiothreitol. Studies of glomerular function show that alpha-125I-ANP is internalized whereas 125I-Tyr0-CNP-(1-22) is not. C-ANP abolishes the specific internalization of alpha-125I-ANP. CNP-(1-22) inhibits internalization of 400 pM alpha-125I-ANP weakly, only approximately 60% being inhibited by 10 microM CNP-(1-22). This implies that the approximately 77-kDa protein, with its low affinity for CNP-(1-22), mediates internalization. Furthermore, CNP-(1-22), as well as alpha-ANP and C-ANP, inhibits glomerular levels of adenosine 3',5'-cyclic monophosphate (cAMP), and CNP-(1-22) does so with a high affinity, which corresponds to its affinity for the approximately 67-kDa protein. The results suggest that the approximately 67-kDa receptor is distinct from the natriuretic peptide clearance receptor and may control cAMP levels.