Renal denervation improves sodium excretion in rats with chronic heart failure: effects on expression of renal ENaC and AQP2

Abstract

Previously we have shown that increased expression of renal epithelial sodium channels (ENaC) may contribute to the renal sodium and water retention observed during chronic heart failure (CHF). The goal of this study was to examine whether renal denervation (RDN) changed the expressions of renal sodium transporters ENaC, sodium-hydrogen exchanger-3 proteins (NHE3), and water channel aquaporin 2 (AQP2) in rats with CHF. CHF was produced by left coronary artery ligation in rats. Four weeks after ligation surgery, surgical bilateral RDN was performed. The expression of ENaC, NHE3, and AQP2 in both renal cortex and medulla were measured. As a functional test for ENaC activation, diuretic and natriuretic responses to ENaC inhibitor benzamil were monitored in four groups of rats (Sham, Sham+RDN, CHF, CHF+RDN). Western blot analysis indicated that RDN (1 wk later) significantly reduced protein levels of α-ENaC, β-ENaC, γ-ENaC, and AQP2 in the renal cortex of CHF rats. RDN had no significant effects on the protein expression of kidney NHE3 in both Sham and CHF rats. Immunofluorescence studies of kidney sections confirmed the reduced signaling of ENaC and AQP2 in the CHF+RDN rats compared with the CHF rats. There were increases in diuretic and natriuretic responses to ENaC inhibitor benzamil in rats with CHF. RDN reduced the diuretic and natriuretic responses to benzamil in CHF rats. These findings suggest a critical role for renal nerves in the enhanced expression of ENaC and AQP2 and subsequent pathophysiology of renal sodium and water retention associated with CHF.NEW & NOTEWORTHY This is the first study to show in a comprehensive way that renal denervation initiated after a period of chronic heart failure reduces the expression of epithelial sodium channels and aquaporin 2 leading to reduced epithelial sodium channel function and sodium retention.

Keywords: heart failure; renal function; renal nerve; sympathetic nerve activity.

Fig. 1.

Representative Western blots showing steady-state levels of full-length ENaC subunits α-ENaC (A), β-ENaC (B), and γ-ENaC (C) protein expression in the cortex and medulla of kidney tissue from Sham, CHF, Sham+RDN, and CHF+RDN rats. Mean values of ENaC expression in the kidneys from each group (bottom). *P < 0.05 vs. Sham, #P < 0.05 vs. without RDN. Images of the bands detected from 40 to 200 kDa using α-ENaC and γ-ENaC antibodies in a Sham (left lane) and CHF rat (right lane) (D). It shows the full-length α-ENaC (97 kDa) and γ-ENaC (80 kDa) bands as well as cleaved forms of α-ENaC and γ-ENaC indicated by the brackets on the left of the blots. Mean values of cleaved forms of α-ENaC and γ-ENaC expression in the kidneys (bottom). CHF, chronic heart failure; ENaC, epithelial sodium channels; RDN, renal denervation.

Fig. 2.

A: representative Western blots showing levels of NHE3 protein expression in the cortex and medulla of kidney tissue from Sham, CHF, Sham+RDN, and CHF+RDN rats. B: mean values of NHE3 expression in the kidneys from each group. *P < 0.05 vs. Sham. CHF, chronic heart failure; NHE3, sodium-hydrogen exchanger-3; RDN, renal denervation.

Fig. 3.

A: representative Western blots showing steady-state levels of AQP2 protein expression in the cortex and medulla of kidney tissue from Sham, CHF, Sham+RDN, and CHF+RDN rats. B: mean values of AQP2 expression in the kidneys from each group. *P < 0.05 vs. Sham, #P < 0.05 vs. without RDN. AQP2, aquaporin 2; CHF, chronic heart failure; RDN, renal denervation.

Fig. 4.

A: immunofluorescent microscopy of renal sections for α-ENaC, β-ENaC, and γ-ENaC in the cortex of kidney from Sham, CHF, Sham+RDN, and CHF+RDN rats. (magnification, ×400). B: mean values of ENaC staining intensity in the kidneys from each group. *P < 0.05 vs. Sham, #P < 0.05 vs. without RDN. CHF, chronic heart failure; ENaC, epithelial sodium channels; RDN, renal denervation.

Fig. 5.

A: immunofluorescent microscopy of renal sections for NHE3 and AQP2 in the cortex of kidney from Sham, CHF, Sham+RDN, and CHF+RDN rats (magnification, ×400). B: mean values of NHE3 and AQP2 staining intensity in the kidneys from each group. *P < 0.05 vs. Sham, #P < 0.05 vs. without RDN. AQP2, aquaporin 2; CHF, chronic heart failure; NHE3, sodium-hydrogen exchanger-3; RDN, renal denervation.

Fig. 6.

A: urine flow in response to ENaC inhibitor benzamil injection in Sham, CHF, Sham+RDN, and CHF+RDN rats. B: cumulative urine flow at baseline and 50 min after benzamil injection in each group. *P < 0.05 vs. respective Sham group; #P < 0.05 vs. without RDN. CHF, chronic heart failure; ENaC, epithelial sodium channels; gkw, grams of kidney weight; RDN, renal denervation.

Fig. 7.

A: sodium excretion in response to ENaC inhibitor benzamil injection in Sham, CHF, Sham+RDN, and CHF+RDN rats. B: cumulative sodium excretion at baseline and 50 min after benzamil injection in each group. *P < 0.05 vs. respective Sham group; #P < 0.05 vs. without RDN. CHF, chronic heart failure; ENaC, epithelial sodium channels; gkw, gram of kidney weight; RDN, renal denervation.