title: The Role of Epithelial Sodium Channel ENaC and the Apical Cl-/HCO3- Exchanger Pendrin in Compensatory Salt Reabsorption in the Setting of Na-Cl Cotransporter (NCC) Inactivation.
abstract: BACKGROUND
  The absence of NCC does not cause significant salt wasting in NCC deficient mice under basal conditions. We hypothesized that ENaC and pendrin play important roles in compensatory salt absorption in the setting of NCC inactivation, and their inhibition and/or downregulation can cause significant salt wasting in NCC KO mice.
  
  
  METHODS
  WT and NCC KO mice were treated with a daily injection of either amiloride, an inhibitor of ENaC, or acetazolamide (ACTZ), a blocker of salt and bicarbonate reabsorption in the proximal tubule and an inhibitor of carbonic anhydrases in proximal tubule and intercalated cells, or a combination of acetazolamide plus amiloride for defined durations. Animals were subjected to daily balance studies. At the end of treatment, kidneys were harvested and examined. Blood samples were collected for electrolytes and acid base analysis.
  
  
  RESULTS
  Amiloride injection significantly increased the urine output (UO) in NCC KO mice (from 1.3 ml/day before to 2.5 ml/day after amiloride, p<0.03, n = 4) but caused only a slight change in UO in WT mice (p>0.05). The increase in UO in NCC KO mice was associated with a significant increase in sodium excretion (from 0.25 mmol/24 hrs at baseline to 0.35 mmol/24 hrs after amiloride injection, p<0.05, n = 4). Daily treatment with ACTZ for 6 days resulted in >80% reduction of kidney pendrin expression in both WT and NCC KO mice. However, ACTZ treatment noticeably increased urine output and salt excretion only in NCC KO mice (with urine output increasing from a baseline of 1.1 ml/day to 2.3 ml/day and sodium excretion increasing from 0.22 mmole/day before to 0.31 mmole/day after ACTZ) in NCC KO mice; both parameters were significantly higher than in WT mice. Western blot analysis demonstrated significant enhancement in ENaC expression in medulla and cortex of NCC KO and WT mice in response to ACTZ injection for 6 days, and treatment with amiloride in ACTZ-pretreated mice caused a robust increase in salt excretion in both NCC KO and WT mice. Pendrin KO mice did not display a significant increase in urine output or salt excretion after treatment with amiloride or ACTZ.
  
  
  CONCLUSIONS
  1. ENaC plays an important role in salt reabsorption in NCC KO mice. 2. NCC contributes to compensatory salt reabsorption in the setting of carbonic anhydrase inhibition, which is associated with increased delivery of salt from the proximal tubule and the down regulation of pendrin. 3. ENaC is upregulated by ACTZ treatment and its inhibition by amiloride causes significant diuresis in NCC KO and WT mice. Despite being considered mild agents individually, we propose that the combination of acetazolamide and amiloride in the setting of NCC inhibition (i.e., hydrochlorothiazide) will be a powerful diuretic regimen.
authors.affiliation: Center on Genetics of Transport and Epithelial Biology, University of Cincinnati, Cincinnati, Ohio, United States of America.|Department of Medicine, University of Cincinnati, Cincinnati, Ohio, United States of America.
authors.forename: Mina
authors.initials: M
authors.lastname: Patel-Chamberlin
authors.affiliation: Center on Genetics of Transport and Epithelial Biology, University of Cincinnati, Cincinnati, Ohio, United States of America.|Department of Medicine, University of Cincinnati, Cincinnati, Ohio, United States of America.
authors.forename: Mujan
authors.initials: M
authors.lastname: Varasteh Kia
authors.affiliation: Center on Genetics of Transport and Epithelial Biology, University of Cincinnati, Cincinnati, Ohio, United States of America.
authors.forename: Jie
authors.initials: J
authors.lastname: Xu
authors.affiliation: Center on Genetics of Transport and Epithelial Biology, University of Cincinnati, Cincinnati, Ohio, United States of America.|Research Services, Veterans Affairs Medical Center, Cincinnati, Ohio, United States of America.
authors.forename: Sharon
authors.initials: S
authors.lastname: Barone
authors.affiliation: Center on Genetics of Transport and Epithelial Biology, University of Cincinnati, Cincinnati, Ohio, United States of America.|Research Services, Veterans Affairs Medical Center, Cincinnati, Ohio, United States of America.
authors.forename: Kamyar
authors.initials: K
authors.lastname: Zahedi
authors.affiliation: Center on Genetics of Transport and Epithelial Biology, University of Cincinnati, Cincinnati, Ohio, United States of America.|Research Services, Veterans Affairs Medical Center, Cincinnati, Ohio, United States of America.|Department of Medicine, University of Cincinnati, Cincinnati, Ohio, United States of America.
authors.forename: Manoocher
authors.initials: M
authors.lastname: Soleimani
mesh_terms: D000086:Acetazolamide / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D027321:Anion Transport Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D062686:Epithelial Sodium Channel Blockers / Q000494:pharmacology; D053503:Epithelial Sodium Channels / Q000235:genetics / Q000378:metabolism*; D007687:Kidney Tubules, Proximal / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D012965:Sodium Chloride / Q000652:urine*; D064486:Solute Carrier Family 12, Member 3 / Q000235:genetics / Q000378:metabolism; D000076988:Sulfate Transporters
chemical_list: D027321:Anion Transport Proteins; D062686:Epithelial Sodium Channel Blockers; D053503:Epithelial Sodium Channels; C578875:Slc12a3 protein, mouse; C494849:Slc26a4 protein, mouse; D064486:Solute Carrier Family 12, Member 3; D000076988:Sulfate Transporters; D012965:Sodium Chloride; D000584:Amiloride; D000086:Acetazolamide