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• W2004501944 abstract "Final urinary acidification is achieved by electrogenic vacuolar H+-ATPases expressed in acid-secretory intercalated cells (ICs) in the connecting tubule (CNT) and the cortical (CCD) and initial medullary collecting duct (MCD), respectively. Electrogenic Na+ reabsorption via epithelial Na+ channels (ENaCs) in the apical membrane of the segment-specific CNT and collecting duct cells may promote H+-ATPases-mediated proton secretion by creating a more lumen-negative voltage. The exact localization where this supposed functional interaction takes place is unknown. We used several mouse models performing renal clearance experiments and assessed the furosemide-induced urinary acidification. Increasing Na+ delivery to the CNT and CCD by blocking Na+ reabsorption in the thick ascending limb with furosemide enhanced urinary acidification and net acid excretion. This effect of furosemide was abolished with amiloride or benzamil blocking ENaC action. In mice deficient for the IC-specific B1 subunit of the vacuolar H+-ATPase, furosemide led to only a small urinary acidification. In contrast, in mice with a kidney-specific inactivation of the alpha subunit of ENaC in the CCD and MCD, but not in the CNT, furosemide alone and in combination with hydrochlorothiazide induced normal urinary acidification. These results suggest that the B1 vacuolar H+-ATPase subunit is necessary for the furosemide-induced acute urinary acidification. Loss of ENaC channels in the CCD and MCD does not affect this acidification. Thus, functional expression of ENaC channels in the CNT is sufficient for furosemide-stimulated urinary acidification and identifies the CNT as a major segment in electrogenic urinary acidification. Final urinary acidification is achieved by electrogenic vacuolar H+-ATPases expressed in acid-secretory intercalated cells (ICs) in the connecting tubule (CNT) and the cortical (CCD) and initial medullary collecting duct (MCD), respectively. Electrogenic Na+ reabsorption via epithelial Na+ channels (ENaCs) in the apical membrane of the segment-specific CNT and collecting duct cells may promote H+-ATPases-mediated proton secretion by creating a more lumen-negative voltage. The exact localization where this supposed functional interaction takes place is unknown. We used several mouse models performing renal clearance experiments and assessed the furosemide-induced urinary acidification. Increasing Na+ delivery to the CNT and CCD by blocking Na+ reabsorption in the thick ascending limb with furosemide enhanced urinary acidification and net acid excretion. This effect of furosemide was abolished with amiloride or benzamil blocking ENaC action. In mice deficient for the IC-specific B1 subunit of the vacuolar H+-ATPase, furosemide led to only a small urinary acidification. In contrast, in mice with a kidney-specific inactivation of the alpha subunit of ENaC in the CCD and MCD, but not in the CNT, furosemide alone and in combination with hydrochlorothiazide induced normal urinary acidification. These results suggest that the B1 vacuolar H+-ATPase subunit is necessary for the furosemide-induced acute urinary acidification. Loss of ENaC channels in the CCD and MCD does not affect this acidification. Thus, functional expression of ENaC channels in the CNT is sufficient for furosemide-stimulated urinary acidification and identifies the CNT as a major segment in electrogenic urinary acidification. The kidneys play a central role in maintaining acid–base homeostasis by reabsorbing bicarbonate and excreting acid equivalents generated by metabolism. Final urinary acidification takes place in the connecting tubule (CNT) and along the different segments of the collecting duct (CD) namely the cortical (CCD), outer medullary, and initial inner medullary collecting duct (MCD). These parts of the nephron are composed of segment-specific cells reabsorbing Na+ and secreting K+ and of intercalated cells (ICs) involved in acid–base transport. Many studies have described functionally and morphologically at least two types of ICs: type A and type B.1.Schuster V.L. Function and regulation of collecting duct intercalated cells.Annu Rev Physiol. 1993; 55: 267-288Crossref PubMed Scopus (171) Google Scholar, 2.Kim J. Kim Y.H. Cha J.H. et al.Intercalated cell subtypes in connecting tubule and cortical collecting duct of rat and mouse.J Am Soc Nephrol. 1999; 10: 1-12PubMed Google Scholar, 3.Alper S.L. Natale J. Gluck S. et al.Subtypes of intercalated cells in rat kidney collecting duct defined by antibodies against erythroid band 3 and renal vacuolar H+-ATPase.Proc Natl Acad Sci USA. 1989; 86: 5429-5433Crossref PubMed Scopus (315) Google Scholar Type A ICs secrete protons via an apically expressed vacuolar H+-ATPase.4.Wagner C.A. Finberg K.E. Breton S. et al.Renal vacuolar H+-ATPase.Physiol Rev. 2004; 84: 1263-1314Crossref PubMed Scopus (336) Google Scholar This proton secretion is functionally coupled to the basolateral anion exchanger AE1 releasing bicarbonate into blood. Type B ICs reverse this process, thereby secreting bicarbonate into urine and absorbing protons.1.Schuster V.L. Function and regulation of collecting duct intercalated cells.Annu Rev Physiol. 1993; 55: 267-288Crossref PubMed Scopus (171) Google Scholar,5.Wagner C.A. Geibel J.P. Acid-base transport in the collecting duct.J Nephrol. 2002; 5 (suppl): S112-S127Google Scholar The role of a third subtype, non-A/non-B ICs, is not fully clarified yet. Proton secretion through vacuolar H+-ATPases in the CNT and CCD is electrogenic and is thought to be indirectly coupled to Na+ reabsorption.4.Wagner C.A. Finberg K.E. Breton S. et al.Renal vacuolar H+-ATPase.Physiol Rev. 2004; 84: 1263-1314Crossref PubMed Scopus (336) Google Scholar,6.Koeppen B.M. Helman S.I. Acidification of luminal fluid by the rabbit cortical collecting tubule perfused in vitro.Am J Physiol. 1982; 242: F521-F531PubMed Google Scholar Na+ reabsorption through the amiloride-sensitive epithelial Na+ channel (ENaC) expressed in neighboring segment-specific cells creates a more lumen-negative potential which has been hypothetized to enhance H+ secretion by H+-ATPases.7.Weinstein A.M. A mathematical model of rat collecting duct III. Paradigms for distal acidification defects.Am J Physiol Renal Physiol. 2002; 283: F1267-F1280Crossref PubMed Scopus (12) Google Scholar, 8.Weinstein A.M. A mathematical model of rat collecting duct I. Flow effects on transport and urinary acidification.Am J Physiol Renal Physiol. 2002; 283: F1237-F1251Crossref PubMed Scopus (32) Google Scholar, 9.Batlle D.C. Segmental characterization of defects in collecting tubule acidification.Kidney Int. 1986; 30: 546-554Abstract Full Text PDF PubMed Scopus (100) Google Scholar, 10.Chang H. Fujita T. A numerical model of acid–base transport in rat distal tubule.Am J Physiol Renal Physiol. 2001; 281: F222-F243PubMed Google Scholar In the MCD, ENaC expression is much lower than in the CNT and CCD,11.Loffing J. Kaissling B. Sodium and calcium transport pathways along the mammalian distal nephron: from rabbit to human.Am J Physiol Renal Physiol. 2003; 284: F628-F643Crossref PubMed Scopus (141) Google Scholar the lumen potential is more positive and H+ secretion is independent from Na+ absorption.12.Jacobsen H.J. Furuya H. Breyer M.D. Mechanism and regulation of proton transport in the outer medullary collecting duct.Kidney Int. 1991; 40: S51-S56Google Scholar The ENaC consists of three subunits termed α, β, γ.13.Kellenberger S. Schild L. Epithelial sodium channel/degenerin family of ion channels: a variety of functions for a shared structure.Physiol Rev. 2002; 82: 735-767Crossref PubMed Scopus (808) Google Scholar,14.Rossier B.C. Pradervand S. Schild L. et al.Epithelial sodium channel and the control of sodium balance: interaction between genetic and environmental factors.Annu Rev Physiol. 2002; 64: 877-897Crossref PubMed Scopus (297) Google Scholar Loss-of-function mutations in either the human α, β, or γ subunits of ENaC cause pseudohypoaldosteronism type 1 characterized by severe neonatal salt wasting, hyperkalemia, and metabolic acidosis.15.Lifton R.P. Gharavi A.G. Geller D.S. Molecular mechanisms of human hypertension.Cell. 2001; 104: 545-556Abstract Full Text Full Text PDF PubMed Scopus (1306) Google Scholar The α subunit plays an essential role in the trafficking of the channel to the cell surface as well as forming part of the pore.13.Kellenberger S. Schild L. Epithelial sodium channel/degenerin family of ion channels: a variety of functions for a shared structure.Physiol Rev. 2002; 82: 735-767Crossref PubMed Scopus (808) Google Scholar Apart from the kidney, ENaC is also expressed in distal colon and in upper and lower airways where it also mediates Na+ reabsorption.14.Rossier B.C. Pradervand S. Schild L. et al.Epithelial sodium channel and the control of sodium balance: interaction between genetic and environmental factors.Annu Rev Physiol. 2002; 64: 877-897Crossref PubMed Scopus (297) Google Scholar Mice with complete inactivation of αENaC develop acute postnatal respiratory distress and die within 40 h of birth from failure to clear their lungs from liquid.16.Hummler E. Barker P. Gatzy J. et al.Early death due to defective neonatal lung liquid clearance in alpha-ENaC-deficient mice.Nat Genet. 1996; 12: 325-328Crossref PubMed Scopus (732) Google Scholar Recently, a novel mouse model has been generated with specific inactivation of αENaC in the entire CD but not in CNT,17.Rubera I. Loffing J. Palmer L.G. et al.Collecting duct-specific gene inactivation of alphaENaC in the mouse kidney does not impair sodium and potassium balance.J Clin Invest. 2003; 112: 554-565Crossref PubMed Scopus (170) Google Scholar providing a tool for studying sodium balance as well as disturbances of proton secretion secondary to defective Na+ absorption. The vacuolar H+-ATPase is composed of at least 13 subunits of which several cell- and tissue-specific isoforms exist.4.Wagner C.A. Finberg K.E. Breton S. et al.Renal vacuolar H+-ATPase.Physiol Rev. 2004; 84: 1263-1314Crossref PubMed Scopus (336) Google Scholar, 18.Nelson N. Harvey W.R. Vacuolar and plasma membrane proton-adenosinetriphosphatases.Physiol Rev. 1999; 79: 361-385Crossref PubMed Scopus (360) Google Scholar, 19.Nishi T. Forgac M. The vacuolar (H+)-ATPases – nature's most versatile proton pumps.Nat Rev Mol Cell Biol. 2002; 3: 94-103Crossref PubMed Scopus (934) Google Scholar The B subunit forms part of the peripheral domain V1 and two isoforms of this subunit, B1 and B2, have been identified. Whereas the B2 isoform (ATP6V1B2) is almost ubiquitously expressed and appears to serve in most cells a house-keeping function, the B1 isoform (ATP6V1B1) has a more limited tissue distribution: specialized cells of the epididymis,20.Finberg K.E. Wagner C.A. Stehberger P.A. et al.Molecular cloning and characterization of Atp6v1b1, the murine vacuolar H+-ATPase B1-subunit.Gene. 2003; 318: 25-34Crossref PubMed Scopus (28) Google Scholar,21.Breton S. Smith P.J. Lui B. Brown D. Acidification of the male reproductive tract by a proton pumping (H+)-ATPase.Nat Med. 1996; 2: 470-472Crossref PubMed Scopus (217) Google Scholar the vas deferens,21.Breton S. Smith P.J. Lui B. Brown D. Acidification of the male reproductive tract by a proton pumping (H+)-ATPase.Nat Med. 1996; 2: 470-472Crossref PubMed Scopus (217) Google Scholar the ciliary body of the eye,22.Wax M.B. Saito I. Tenkova T. et al.Vacuolar H+-ATPase in ocular ciliary epithelium.Proc Natl Acad Sci USA. 1997; 94: 6752-6757Crossref PubMed Scopus (42) Google Scholar the inner ear,23.Karet F.E. Finberg K.E. Nelson R.D. et al.Mutations in the gene encoding B1 subunit of H+-ATPase cause renal tubular acidosis with sensorineural deafness.Nat Genet. 1999; 21: 84-90Crossref PubMed Scopus (539) Google Scholar and all subtypes of ICs of the kidney.20.Finberg K.E. Wagner C.A. Stehberger P.A. et al.Molecular cloning and characterization of Atp6v1b1, the murine vacuolar H+-ATPase B1-subunit.Gene. 2003; 318: 25-34Crossref PubMed Scopus (28) Google Scholar,24.Nelson R.D. Guo X.L. Masood K. et al.Selectively amplified expression of an isoform of the vacuolar H+-ATPase 56-kilodalton subunit in renal intercalated cells.Proc Natl Acad Sci USA. 1992; 89: 3541-3545Crossref PubMed Scopus (181) Google Scholar Mutations in the gene encoding the B1 subunit result in distal renal tubular acidosis (dRTA) in man characterized by the inability of the distal nephron to appropriately acidify the urine.23.Karet F.E. Finberg K.E. Nelson R.D. et al.Mutations in the gene encoding B1 subunit of H+-ATPase cause renal tubular acidosis with sensorineural deafness.Nat Genet. 1999; 21: 84-90Crossref PubMed Scopus (539) Google Scholar A mouse model deficient for the Atp6v1b1 gene has recently been generated with impaired urinary acidification.25.Finberg K.E. Wagner C.A. Bailey M.A. et al.The B1 subunit of the H+ATPase is required for maximal urinary acidification.Proc Nat Acad Sci USA. 2005; 102: 13616-13621Crossref PubMed Scopus (106) Google Scholar Vacuolar H+-ATPase activity is almost completely absent from the CCD. Interestingly, enhanced luminal appearance of the B2 subunit has been noted in B1-deficient ICs, suggesting that the B2 isoform could compensate for the loss of B1.25.Finberg K.E. Wagner C.A. Bailey M.A. et al.The B1 subunit of the H+ATPase is required for maximal urinary acidification.Proc Nat Acad Sci USA. 2005; 102: 13616-13621Crossref PubMed Scopus (106) Google Scholar Nevertheless, Atp6v1b1-deficient mice develop a more severe metabolic acidosis with inappropriately alkaline urine when challenged with an oral acid-load (NH4Cl) characteristic of dRTA. Different subtypes of dRTA have been proposed and classified based on clinical tests.26.Arruda J.A. Kurtzman N.A. Mechanisms and classification of deranged distal urinary acidification.Am J Physiol. 1980; 239: F515-F523PubMed Google Scholar,27.DuBose Jr, T.D. Alpern R.J. Renal tubular acidosis.in: Scriver C.R. Beaudet A.L. Sly W.S. Valle D. The Metabolic and Molecular Bases of Inherited Disease. 8th edn. McGraw-Hill, New York2001: 4983-5021Google Scholar These tests include oral NH4Cl- or parenteral Na2SO4-loading and application of furosemide, which all lead to an acute urinary acidification in healthy subjects but not in patients with specific subtypes of dRTA. Furosemide inhibits the luminal Na+/K+/2Cl− cotransporter in the thick ascending limb, thereby increasing the delivered fraction of Na+ to the subsequent nephron segments and stimulating Na+ reabsorption through ENaC. Patients lacking an appropriate urinary acidification after furosemide application have been classified as suffering from the ‘voltage-defective’ form of dRTA.9.Batlle D.C. Segmental characterization of defects in collecting tubule acidification.Kidney Int. 1986; 30: 546-554Abstract Full Text PDF PubMed Scopus (100) Google Scholar In order to test for and localize the possible functional interaction between Na+ reabsorption and H+ secretion in the different parts of the distal nephron, we performed clearance studies in mice treated acutely with furosemide, hydrochlorothiazide, and amiloride, and measured urinary acidification and net acid excretion (NAE). Immunostaining of mouse kidney with antibodies against calbindin D28k, B1 subunit of the vacuolar H+-ATPase, and the β subunit of the ENaC revealed that the B1 subunit of the H+-ATPase is exclusively expressed in CNTs and CDs which were identified on account of their characteristic localization in the cortical labyrinth and the medullary rays, respectively, and by their strong labelling with antibodies against the beta subunit of the epithelial sodium channel (ENaC) and the calcium-binding protein calbindin D28k (Figure 1). Higher magnification showed that the B1 subunit is only expressed in a subtype of epithelial cells lining CNT and CD that do not express βENaC or calbindinD28k. This staining pattern is consistent with an exclusive localization of the B1 subunit in ICs28.Loffing J. Loffing-Cueni D. Valderrabano V. et al.Distribution of transcellular calcium and sodium transport pathways along mouse distal nephron.Am J Physiol Renal Physiol. 2001; 281: F1021-F1027Crossref PubMed Scopus (263) Google Scholar,29.Biner H.L. Arpin-Bott M.P. Loffing J. et al.Human cortical distal nephron: distribution of electrolyte and water transport pathways.J Am Soc Nephrol. 2002; 13: 836-847PubMed Google Scholar which are intermingled between the ENaC (and calbindin) positive CNT and CD (principal) cells. In a first group of control animals, the effect of acute furosemide and subsequent amiloride application on urinary acidification was tested. As shown in Figure 2a, the initial urinary pH was similar in all groups before the application of furosemide. A bolus of furosemide (2 μg/g of body weight (BW)) after 60 min lowered the urinary pH with being significantly more acidic after 90 min (furosemide: pH 6.16±0.08 versus control: pH 6.49±0.07), after 120 min (furosemide: pH 5.72±0.10 versus control: pH 6.39±0.08), and 150 min (furosemide: 5.63±0.084 versus control: pH 6.16±0.05). The furosemide-induced urinary acidification was completely abolished by the subsequent administration of amiloride or benzamil, inhibitors of ENaC activity: pH 6.09±0.09 in the furosemide+amiloride group versus 5.72±0.10 in furosemide-alone group after 120 min as well as after 150 min: pH 6.19±0.12 for furosemide+amiloride versus pH 5.63±0.08 for furosemide alone. The urine production and excretion remained stable in the control group during the entire experiment, whereas it increased in furosemide-treated animals from 1.9±0.3 μl/g after 60 min to 5.2±0.8 μl/g after 90 min. A similar effect was observed in mice treated subsequently with amiloride, resulting in an increase in urine output from 3.0±0.6 μl/g BW after 60 min to 6.5±0.7 μl/g BW after 90 min. (Figure 2b). The analysis of urinary electrolyte excretion revealed that the fractional excretion (FE in %) of sodium and chloride increased in mice treated with furosemide plus amiloride (Figure 2d and e) with a maximum after 120 min (FE of sodium 1.67±0.31% for furosemide+amiloride versus 0.25±0.08% in control and FE of chloride 2.29±0.38% in furosemide+amiloride versus 0.70±0.24% in control). As summarized in Table 1 and Figure 3, furosemide treatment resulted in a slightly decreased blood potassium concentration (4.8±0.3 mmol/l in furosemide versus 5.7±0.2 mmol/l in control) and augmented blood bicarbonate concentration (21.0±0.7 with furosemide versus 19.7±1.0 mmol/l under control).Table 1Summary of blood pH, gas, and electrolytes of wild-type mice left untreated (control) and wild-type and Atp6V1b1-deficient mice treated with furosemide and amiloride at the end of the experimental periodAtp6v1b1 +/+ controlAtp6v1b1 +/+ furosemideAtp6v1b1 +/+ furosemide amilorideAtp6v1b1 -/- furosemidepH7.22±0.027.24±0.027.24±0.027.25±0.04pCO2 (mmHg)50.6±3.950.8±2.047.7±2.153.8±7.4HCO3− (mmol/l)19.7±1.021.0±0.719.8±0.421.8±1.6*Marks significant differences P<0.05,K+ (mmol/l)5.7±0.24.8±0.3**P<0.001.5.5±0.24.3±0.1**P<0.001.Na+ (mmol/l)145.5±0.8147.7±0.9145.4±0.5147.6±1.4Cl− (mmol/l)121.8±1.5120.9±0.8121.1±0.8119.6±2.4Serum creatinine (mg/dl)0.14±0.020.11±0.010.13±0.020.12±0.01Weight (g)27.5±1.529.6±1.125.4±1.124.6±1.0CrCl/BW 30 min (μl/min/g)18.3±4.233.4±6.115.6±4.928.5±6.0CrCl/BW 90 min (μl/min/g)18.7±4.744.6±8.5*Marks significant differences P<0.05,19.0±3.426.6±5.6CrCl/BW 120 min (μl/min/g)13.8±3.933.2±4.6**P<0.001.22.1±7.020.3±9.2NH3/NH4+ (mM)/creatinine (mg/dl), 30 min1.04±0.050.94±0.081.47±0.521.18±0.15NH3/NH4+ (mM)/creatinine (mg/dl), 150 min1.40±0.0.182.60±0.082.04±0.082.56±0.26NAE (mM/creatinine (mg/dl), 30 min1.21±1.111.03±0.071.63±0.641.29±0.20NAE (mM/creatinine (mg/dl), 150 min2.91±0.286.16±0.595.02±0.334.40±0.44*Marks significant differences P<0.05,BW, body weight; CrCl, creatinine clearance; NAE, net acid excretion.* Marks significant differences P<0.05,** P<0.001. Open table in a new tab Figure 3Effect of furosemide and amiloride on blood pH and electrolytes. (a–d) Furosemide treatment resulted in decreased blood potassium concentration as expected from the increased urinary excretion. Blood bicarbonate levels showed a tendency to be higher which did not reach statistical significance. **P<0.001 versus control.View Large Image Figure ViewerDownload (PPT) BW, body weight; CrCl, creatinine clearance; NAE, net acid excretion. Thus, treatment of mice with furosemide led to the expected increase in urinary output with an increase in fractional sodium and potassium excretion accompanied by a strong urinary acidification as described previously for rats and humans.9.Batlle D.C. Segmental characterization of defects in collecting tubule acidification.Kidney Int. 1986; 30: 546-554Abstract Full Text PDF PubMed Scopus (100) Google Scholar The stimulation of urinary acidification was abolished by the ENaC inhibitors amiloride and benzamil consistent with a role of ENaC in the furosemide-induced urinary acidification. A similar effect of amiloride and the structurally unrelated ENaC blocker triamterene on the furosemide-induced urinary acidification has also been reported in rats.30.Hropot M. Fowler N. Karlmark B. et al.Tubular action of diuretics: distal effects on electrolyte transport and acidification.Kidney Int. 1985; 28: 477-489Abstract Full Text PDF PubMed Scopus (121) Google Scholar To demonstrate that the furosemide-induced urinary acidification depended on the activity of vacuolar H+-ATPases localized in ICs, we used a mouse model deficient for the IC-specific B1 subunit (Atp6v1b1 -/-). Atp6v1b1 -/- furosemide-treated mice exhibited a markedly higher basal urine pH than the Atp6v1b1 +/+ furosemide-treated mice (pH 7.19±0.16 versus 6.44±0.10) and only a mild urinary acidification upon furosemide administration could be observed: pH 6.86±0.15 versus 5.72±0.10 after 120 min (Figure 4a). The residual urinary acidification observed in the B1-deficient mice may be owing to a partial compensation by the B2 isoform which is more luminally localized in ICs of Atp6v1b1 -/- mice.25.Finberg K.E. Wagner C.A. Bailey M.A. et al.The B1 subunit of the H+ATPase is required for maximal urinary acidification.Proc Nat Acad Sci USA. 2005; 102: 13616-13621Crossref PubMed Scopus (106) Google Scholar To assess the effect of the treatments on NAE, we measured total NH3/NH4+ and total phosphate excretion in urine and estimated NAE from these data as the urine samples were too small to measure titratable acidity. Both total NH3/NH4+ and total phosphate excretion increased in response to furosemide as described previously in rats.30.Hropot M. Fowler N. Karlmark B. et al.Tubular action of diuretics: distal effects on electrolyte transport and acidification.Kidney Int. 1985; 28: 477-489Abstract Full Text PDF PubMed Scopus (121) Google Scholar Application of furosemide increased NAE from 2.9±0.3 mmol/l/mg/dl creatinine in control animals to 6.2±0.6 mmol/l/mg/dl creatinine (Figure 4b). In mice given amiloride, NAE showed a tendency to be lower (P=0.08). In contrast, in benzamil-treated mice and in the B1-deficient mice, NAE was significantly lower (benzamil: 3.7±0.6 mmol/l/mg/dl creatinine, B1 KO: 4.4±0.4 mmol/l/mg/dl creatinine). Atp6v1b1 -/- furosemide-treated mice also had a strikingly higher urine output than the Atp6v1b1 +/+ furosemide-treated mice after 60 min before the furosemide administration: 3.2±0.83 μl/g BW versus 1.9±0.34 μl/g BW (Figure 4c). Furosemide administration led to a more profound diuresis in Atp6v1b1 -/- mice: 8.9±1.0 μl/g BW versus 5.2±0.7 μl/g BW after 90 min (Figure 4c). In addition, Atp6v1b1 -/- mice had a significantly lower basal FE of potassium than Atp6v1b1 +/+ mice (1.89±0.24 versus 6.09±0.60%) (Figure 4d), basal FE of sodium (0.06±0.01 versus 0.13±0.01%) (Figure 4e), and basal FE of chloride (0.04±0.01 versus 0.14±0.01%) (Figure 4f) pointing to a defect in urine concentration. Furosemide administration further revealed a significant difference in the FE of potassium, whereas the FE of sodium and chloride did not differ between Atp6v1b1 -/- and Atp6v1b1 +/+ mice (Figure 4e and f). As summarized in Table 1 and Figure 5, analysis of blood electrolytes revealed differences in potassium and bicarbonate levels. Scnn1aloxloxCre mice lack the alpha subunit of the ENaC in the CCD and MCD but not in the connecting segment.17.Rubera I. Loffing J. Palmer L.G. et al.Collecting duct-specific gene inactivation of alphaENaC in the mouse kidney does not impair sodium and potassium balance.J Clin Invest. 2003; 112: 554-565Crossref PubMed Scopus (170) Google Scholar We used these mice to test for the furosemide-stimulated urinary acidification. Basal urine output, urine pH, and electrolyte content were not significantly different between control mice (Scnn1aloxlox) and mice with the specific ablation of αENaC expression (Scnn1aloxloxCre) (compare Tables 1 and 2). Furosemide administration caused urinary acidification in both genotypes to a similar extent: Scnn1aloxlox and Scnn1aloxloxCre mice: pH 6.00±0.10 versus 5.89±0.09 after 120 min (Figure 6a). Also, estimated NAE at the time point 150 min was similar in both groups of mice (Figure 6b; Scnn1aloxlox 5.4±0.9 mmol/l/mg/dl creatinine versus Scnn1aloxloxCre 4.2±0.6 mmol/l/mg/dl creatinine, P=0.31). No difference between Scnn1aloxlox and Scnn1aloxloxCre mice in urine output could be detected (7.9±1.9 μl/g BW versus 8.2±1.1 μl/g BW after 90 min.) (Figure 6c). Furosemide also failed to unmask a possible difference between Scnn1aloxlox and Scnn1aloxloxCre mice in the FE of sodium: 1.87±0.42% versus 2.13±0.54% after 120 min (Figure 6e and Table 2). However, blood electrolyte analysis revealed a significantly lower blood bicarbonate concentration (23.5±0.7 mmol/l in Scnn1aloxlox versus 21.3±0.7 mmol/l in Scnn1aloxloxCre (Table 2 and Figure 7b). Thus, after application of furosemide, no significant difference was found in the diuretic response, urinary acidification, and electrolyte excretion suggesting that preserved expression of αENaC in the CNT is sufficient to maintain the furosemide-induced urinary acidification.Table 2Summary of blood pH, gas, and electrolytes of Scnn1aloxlox and Scnn1aloxloxCre mice deficient for the alpha ENaC subunit treated with furosemide and hydrochlorothiazideScnn1aloxlox furosemideScnn1aloxloxCre furosemideScnn1aloxlox furosemide HCTScnn1aloxloxCre furosemide HCTpH7.28±0.027.28±0.037.33±0.037.25±0.03PCO2 (mmHg)51.4±2.247.3±3.143.9±4.955.4±2.1*Marks significant differences P<0.05, **P<0.001.HCO3− (mmol/l)23.5±0.721.3±0.7*Marks significant differences P<0.05, **P<0.001.21.9±0.923.3±1.3K+ (mmol/l)4.4±0.24.4±0.34.5±0.24.9±0.2Na+ (mmol/l)147.8±0.9147.8±0.7147.6±0.8148.0±0.5Cl− (mmol/l)119.9±1.8120.2±1.3114.2±1.5114.6±1.6Serum creatinine (mg/dl)0.15±0.020.11±0.010.14±0.030.13±0.02Weight (g)25.4±1.924.9±1.233.6±1.729.9±1.9CrCl/BW 30 min (μl/min/g)14.4±3.421.7±3.324.15±6.329.0±4.4CrCl/BW 90 min (μl/min/g)21.5±3.720.8±2.721.6±3.022.4±4.1CrCl/BW 120 min (μl/min/g)10.5±2.013.9±2.212.8±1.713.4±2.4NH3/NH4+ (mM)/creatinine (mg/dl), 30 min1.25±0.301.47±0.73NH3/NH4+ (mM)/creatinine (mg/dl), 150 min2.24±0.361.82±0.24NAE (mM/creatinine (mg/dl), 30 min1.46±0.321.93±0.70NAE (mM/creatinine (mg/dl), 150 min5.41±0.904.25±0.60BW, body weight; CrCl, creatinine clearance; NAE, net acid excretion.* Marks significant differences P<0.05, **P<0.001. Open table in a new tab Figure 7Effect of furosemide on blood pH and electrolytes in Scnn1aloxlox and Scnn1aloxloxCre mice. Scnn1aloxloxCre mice had a significantly lower bicarbonate concentration. Other parameters were not different. *P<0.05 versus Scnn1aloxlox.View Large Image Figure ViewerDownload (PPT) BW, body weight; CrCl, creatinine clearance; NAE, net acid excretion. In order to inhibit a possible compensatory increase in Na+ absorption in the distal tubule via the thiazide-sensitive Na+/Cl− cotransporter, furosemide was applied together with hydrochlorothiazide. Figure 8a shows that the urinary pH of Scnn1aloxloxCre mice did not differ significantly from the urine pH of Scnn1aloxlox mice before furosemide and hydrochlorothiazide (furo+HCT) administration. After administration of furosemide together with hydrochlorothiazide (furo+HCT), Scnn1aloxloxCre mice decreased their urine pH to the same extent as the Scnn1aloxlox mice: pH 5.77±0.12 versus pH 5.66±0.10 after 120 min. Urine output, fractional electrolyte excretion and systemic electrolyte, and blood gas status were not distinguishable between both mouse lines (Figures 8 and 9 and Table 2).Figure 9Blood pH and electrolytes in furosemide- and hydrochlorothiazide-treated Scnn1aloxlox (n=5) and Scnn1aloxloxCre (n=7) mice. No significant difference between Scnn1aloxloxand Scnn1aloxloxCre mice in blood pH and electrolytes could be observed.View Large Image Figure ViewerDownload (PPT) In the present study, we investigated the functional interaction between renal Na+ reabsorption through ENaC and H+ secretion by vacuolar H+-ATPases in the connecting segment and CCD. Both ENaC and vacuolar H+-ATPases containing the B1 subunit isoform are expressed together in the same nephron segments, namely CNT and CCD in neighboring cells. H+ secretion by vacuolar H+-ATPases is electrogenic and thought to be indirectly coupled to Na+ reabsorption.31.Hamm L.L. Alpern R.J. Cellular mechanisms of renal tubular acidification.in: Seldin D.W. Giebisch G. The Kidney: Physiology and Pathophysiology. 3rd edn. Lippincott Williams & Wilkins, Philadelphia2000: 1935-1979Google S" @default.
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• W2004501944 title "The connecting tubule is the main site of the furosemide-induced urinary acidification by the vacuolar H+-ATPase" @default.
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• W2004501944 cites W1820962178 @default.
• W2004501944 cites W1906008749 @default.
• W2004501944 cites W1971353989 @default.
• W2004501944 cites W1993274286 @default.
• W2004501944 cites W2000335872 @default.
• W2004501944 cites W2007985127 @default.
• W2004501944 cites W2012329129 @default.
• W2004501944 cites W2019216539 @default.
• W2004501944 cites W2042311423 @default.
• W2004501944 cites W2046585193 @default.
• W2004501944 cites W2075574259 @default.
• W2004501944 cites W2081516285 @default.
• W2004501944 cites W2082624234 @default.
• W2004501944 cites W2082968664 @default.
• W2004501944 cites W2104599806 @default.
• W2004501944 cites W2111907151 @default.
• W2004501944 cites W2122800480 @default.
• W2004501944 cites W2124663880 @default.
• W2004501944 cites W2126636958 @default.
• W2004501944 cites W2130632820 @default.
• W2004501944 cites W2160843443 @default.
• W2004501944 cites W2163948482 @default.
• W2004501944 cites W2165743932 @default.
• W2004501944 cites W2172109574 @default.
• W2004501944 cites W2174130172 @default.
• W2004501944 cites W2186994815 @default.
• W2004501944 cites W2327777489 @default.
• W2004501944 cites W75942344 @default.
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