C Type Natriuretic Peptide Receptor Activation Inhibits Sodium Channel Activity in Human Aortic Endothelial Cells by Activating the Diacylglycerol-Protein Kinase C Pathway

Abstract

The C-type natriuretic peptide receptor (NPRC) is expressed in many cell types and binds all natriuretic peptides with high affinity. Ligand binding results in the activation or inhibition of various intracellular signaling pathways. Although NPRC ligand binding has been shown to regulate various ion channels, the regulation of endothelial sodium channel (EnNaC) activity by NPRC activation has not been studied. The objective of this study was to investigate mechanisms of EnNaC regulation associated with NPRC activation in human aortic endothelial cells (hAoEC). EnNaC protein expression and activity was attenuated after treating hAoEC with the NPRC agonist cANF compared to vehicle, as demonstrated by Western blotting and patch clamping studies, respectively. NPRC knockdown studies using siRNA’s corroborated the specificity of EnNaC regulation by NPRC activation mediated by ligand binding. The concentration of multiple diacylglycerols (DAG) and the activity of protein kinase C (PKC) was augmented after treating hAoEC with cANF compared to vehicle, suggesting EnNaC activity is down-regulated upon NPRC ligand binding in a DAG-PKC dependent manner. The reciprocal cross-talk between NPRC activation and EnNaC inhibition represents a feedback mechanism that presumably is involved in the regulation of endothelial function and aortic stiffness.

Activity and function of the endothelial sodium channel is regulated by the effector domain of MARCKS like protein 1 in mouse aortic endothelial cells

“…EnNaC) with a conductance of 16pS. In a previous study we reported the expression of a 25pS channel in human aortic endothelial cells (7). As expected, single channel characteristics of various ion channels could be different among species.…”

“…Our group previously showed the function of EnNaC is negatively regulated by activation of the NPRC and inhibition of NPRC results in an increase in EnNaC activity (7).…”

“…Natriuretic peptides have been shown to inhibit ENaC activity in the kidney. A previous study showed activation of the clearance receptor for the natriuretic peptides (NPR-C) is coupled to the inhibition of EnNaC activity in human aortic endothelial cells (7). Downregulation of NPR-C gene expression and protein function has been observed in multiple disease models.…”

Deletion of the Npr3 gene increases severity of acute lung injury in obese mice

“…27 NPR-C, which is encoded by the Npr3 gene, has no guanylate cyclase domain 28 and was originally thought to function primarily as a clearance receptor, although it has also been shown to have potential signaling mechanisms via activation of Gcoupled proteins or phosphatidylinositol-4,5-biphosphate (PIP 2 ). 29,30 The receptors that mediate the protective effect of ANP on ALI are not known, however, studies by our lab suggest that both NPR-A and NPR-C may play a role. Previously, we have shown that ANP attenuates LPS-induced pulmonary edema even in the absence of NPR-A.…”

“…Ligand binding results in activation of the guanylyl cyclase and synthesis of cGMP, which is thought to mediate most of the biological properties of the natriuretic peptides 27 . NPR‐C, which is encoded by the Npr3 gene, has no guanylate cyclase domain 28 and was originally thought to function primarily as a clearance receptor, although it has also been shown to have potential signaling mechanisms via activation of G‐coupled proteins or phosphatidylinositol‐4,5‐biphosphate (PIP 2 ) 29,30 . The receptors that mediate the protective effect of ANP on ALI are not known, however, studies by our lab suggest that both NPR‐A and NPR‐C may play a role.…”