The sodium chloride cotransporter (NCC) and epithelial sodium channel (ENaC) associate

The thiazide-sensitive sodium chloride cotransporter (NCC) and the epithelial sodium channel (ENaC) are two of the most important determinants of salt balance and thus systemic blood pressure. Abnormalities in either result in profound changes in blood pressure. There is one segment of the nephron where these two sodium transporters are coexpressed, the second part of the distal convoluted tubule. This is a key part of the aldosterone-sensitive distal nephron, the final regulator of salt handling in the kidney. Aldosterone is the key hormonal regulator for both of these proteins. Despite these shared regulators and coexpression in a key nephron segment, associations between these proteins have not been investigated. After confirming apical localization of these proteins, we demonstrated the presence of functional transport proteins and native association by blue native PAGE. Extensive coimmunoprecipitation experiments demonstrated a consistent interaction of NCC with α- and γ-ENaC. Mammalian two-hybrid studies demonstrated direct binding of NCC to ENaC subunits. Fluorescence resonance energy transfer and immunogold EM studies confirmed that these transport proteins are within appropriate proximity for direct binding. Additionally, we demonstrate that there are functional consequences of this interaction, with inhibition of NCC affecting the function of ENaC. This novel finding of an association between ENaC and NCC could alter our understanding of salt transport in the distal tubule.

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Abstract 076: Urinary Exosomal Epithelial Sodium Channel And Sodium-chloride Cotransporter Measurement In Humans

Hypertension ◽

10.1161/hyp.64.suppl_1.076 ◽

2014 ◽

Vol 64 (suppl_1) ◽

Author(s):

TAOPHEEQ A MUSTAPHA ◽

VICTOR NWAZUE ◽

KEVIN SCHEY ◽

RAJ SATISH ◽

JAMES M LUTHER

Keyword(s):

Sodium Chloride ◽

Sodium Channel ◽

Multiple Reaction Monitoring ◽

Epithelial Sodium Channel ◽

Sodium Balance ◽

Sodium Reabsorption ◽

Dependent Manner ◽

Creatinine Concentration ◽

Spot Urine ◽

Sodium Chloride Cotransporter

Sodium reabsorption in the distal nephron is tightly regulated in part by epithelial sodium channel (ENaC) and sodium chloride cotransporter (NCC), although non-invasive measure of these proteins in humans has not previously been feasible. We recently analyzed the urinary exosomal proteome and identified candidate targets for quantification of ENaC and NCC using targeted mass spectrometry. To test the hypothesis that urinary exosomal ENaC and NCC are altered during renin-angiotensin-aldosterone system activation, we activated the endogenous RAAS using a low sodium diet (LS) in two separate studies. We provided 8 subjects LS diet (10mmol/day for 7days) to assess urinary protein excretion at 7 days (study 1) and longitudinally over the course of 1 week (study 2). Daily 24-hour urine was collected to monitor sodium balance, and spot urine samples were obtained each morning on days 0, 2, 4, and 6 of LS diet. Urinary exosomal ENaC-α, ENaC-γ, and NCC peptides were analyzed using targeted multiple-reaction-monitoring analysis quantified with stable-isotope peptide standards, and results were normalized to urine creatinine concentration. In study 1, urinary ENaCγ increased after 8 days of LS diet (Figure A). In study 2, urinary exosomal ENaCγ (Figure B) and NCC peptides (Figure C) increased in a time-dependent manner during LS diet. These measures of urinary sodium channel expression may provide further insight into distal sodium reabsorption in human hypertension.

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Changes in plasma aldosterone and electrolytes levels, kidney epithelial sodium channel (ENaC) and blood pressure in normotensive WKY and hypertensive SHR rats following gonadectomy and chronic testosterone treatment

Steroids ◽

10.1016/j.steroids.2017.09.008 ◽

2017 ◽

Vol 128 ◽

pp. 128-135 ◽

Cited By ~ 3

Author(s):

Su Yi Loh ◽

Nelli Giribabu ◽

Naguib Salleh

Keyword(s):

Blood Pressure ◽

Sodium Channel ◽

Epithelial Sodium Channel ◽

Plasma Aldosterone ◽

Shr Rats ◽

Testosterone Treatment ◽

Epithelial Sodium Channel Enac

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Sub-chronic testosterone treatment increases the levels of epithelial sodium channel (ENaC)-α,βandγin the kidney of orchidectomized adult male Sprague–Dawley rats

PeerJ ◽

10.7717/peerj.2145 ◽

2016 ◽

Vol 4 ◽

pp. e2145 ◽

Cited By ~ 11

Author(s):

Su Yi Loh ◽

Nelli Giribabu ◽

Naguib Salleh

Keyword(s):

Blood Pressure ◽

Sodium Channel ◽

Adult Male ◽

Epithelial Sodium Channel ◽

Physiological Parameter ◽

Mrna Levels ◽

Sprague Dawley ◽

Collecting Ducts ◽

Distal Tubules ◽

Epithelial Sodium Channel Enac

Testosterone has been reported to cause blood pressure to increase. However mechanisms that underlie the effect of this hormone on this physiological parameter are currently not well understood. The aims of this study were to investigate effects of testosterone on expression ofα,βandγ-epithelial sodium channel (ENaC) proteins and messenger RNAs (mRNAs) in kidneys, the channel known to be involved in Na+reabsorption, which subsequently can affect the blood pressure.Methods.Adult male Sprague–Dawley (SD) rats were orchidectomized fourteen days prior to receiving seven days treatment with testosterone propionate (125 µg/kg/day or 250 µg/kg/day) with or without flutamide (androgen receptor blocker) or finasteride (5α-reductase inhibitor). Following sacrifice, the kidneys were removed and were subjected forα,βandγ-ENaC protein and mRNA expression analyses by Western blotting and Real-time PCR (qPCR) respectively. The distribution ofα,βandγ-ENaC proteins in kidneys were observed by immunofluorescence.Results.Theα,βandγ-ENaC proteins and mRNA levels in kidneys were enhanced in rats which received testosterone-only treatment. In these rats,α,βandγ-ENaC proteins were distributed in the distal tubules and collecting ducts of the nephrons. Co-treatment with flutamide or finasteride resulted in the levels ofα,βandγ-ENaC proteins and mRNAs in kidneys to decrease. In conclusions, increases inα,βandγ-ENaC protein and mRNA levels in kidneys mainly in the distal tubules and collecting ducts under testosterone influence might lead to enhance Na+reabsorption which subsequently might cause an increase in blood pressure.

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Regulation of blood pressure, the epithelial sodium channel (ENaC), and other key renal sodium transporters by chronic insulin infusion in rats

AJP Renal Physiology ◽

10.1152/ajprenal.00108.2005 ◽

2006 ◽

Vol 290 (5) ◽

pp. F1055-F1064 ◽

Cited By ~ 59

Author(s):

Jian Song ◽

Xinqun Hu ◽

Shahla Riazi ◽

Swasti Tiwari ◽

James B. Wade ◽

...

Keyword(s):

Blood Pressure ◽

Sodium Channel ◽

Insulin Infusion ◽

Collecting Duct ◽

Epithelial Sodium Channel ◽

Kidney Cortex ◽

Sodium Reabsorption ◽

Cortical Collecting Duct ◽

Sprague Dawley ◽

Epithelial Sodium Channel Enac

Hyperinsulinemia is associated with hypertension. Dysregulation of renal distal tubule sodium reabsorption may play a role. We evaluated the regulation of the epithelial sodium channel (ENaC) and the thiazide-sensitive Na-Cl cotransporter (NCC) during chronic hyperinsulinemia in rats and correlated these changes to blood pressure as determined by radiotelemetry. Male Sprague-Dawley rats (∼270 g) underwent one of the following three treatments for 4 wk ( n = 6/group): 1) control; 2) insulin-infused plus 20% dextrose in drinking water; or 3) glucose water-drinking (20% dextrose in water). Mean arterial pressures were increased by insulin and glucose (mmHg at 3 wk): 98 ± 1 (control), 107 ± 2 (insulin), and 109 ± 3 (glucose), P < 0.01. Insulin (but not glucose) increased natriuretic response to benzamil (ENaC inhibitor) and hydrochlorothiazide (NCC inhibitor) on average by 125 and 60%, respectively, relative to control rats, suggesting increased activity of these reabsorptive pathways. Neither insulin nor glucose affected the renal protein abundances of NCC or the ENaC subunits (α, β, and γ) in kidney cortex, outer medulla, or inner medulla in a major way, as determined by immunoblotting. However, insulin and to some extent glucose increased apical localization of these subunits in cortical collecting duct principal cells, as determined by immunoperoxidase labeling. In addition, insulin decreased cortical “with no lysine” kinase (WNK4) abundance (by 16% relative to control), which may have increased NCC activity. Overall, insulin infusion increased blood pressure, and NCC and ENaC activity in rats. Increased apical targeting of ENaC and decreased WNK4 expression may be involved.

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