Halide transport patterns of apical anion exchange in rabbit cortical collecting duct intercalated cells

1996 ◽  
Vol 271 (4) ◽  
pp. F799-F805 ◽  
Author(s):  
C. Emmons
Keyword(s):  
Anion Exchange ◽  
Intracellular Ph ◽  
Anion Exchanger ◽  
Collecting Duct ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Intracellular Acidification ◽  
Transport Patterns ◽  
Exchange Activity

To investigate halide selectivity patterns of cortical collecting duct (CCD) intercalated cells (ICs), intracellular pH (pHi) was measured in perfused rabbit outer CCD ICs in Na(+)-free, HCO-3-buffered, 115 mM Cl solutions. Apical anion exchange activity was measured as the dpH/dt after lumen Cl- replacement with gluconate. The perfusate was randomly changed to equimolar Br-, F-, or I-. Addition of Br- to the lumen in place of gluconate caused a brisk acidification similar to that with Cl- return. The acidification rate with I- replacement was only approximately 25% of that seen with Cl- readdition, and F- substitution resulted in a small alkalinization. In a separate protocol, each halide was substituted for lumen Cl-. Replacement of lumen Cl- with Br- resulted in intracellular acidification, whereas F- substitution caused an increase in pHi similar to that with gluconate. A slower rate of alkalinization was seen with I-. Separate tubules were perfused and bathed in Cl(-)-free gluconate solutions for 10 min, and then either Cl- or Br- was returned to the lumen. Similar rates of acidification were found with either Cl- or Br- return. Taken together, the results show that, at a halide concentration of 115 mM, the halide selectivity transport pattern of the apical anion exchanger of CCD ICs is Cl- = Br- > I- > F-.

Transport characteristics of the apical anion exchanger of rabbit cortical collecting duct β-cells

1999 ◽  
Vol 276 (4) ◽  
pp. F635-F643 ◽  
Author(s):  
Cheryl Emmons
Keyword(s):  
Anion Exchange ◽  
Anion Exchanger ◽  
Collecting Duct ◽  
Similar Rate ◽  
Cortical Collecting Duct ◽  
Β Cells ◽  
Β Cell ◽  
Collecting Ducts ◽  
The Mean ◽  
Exchange Activity

To functionally characterize transport properties of the apical anion exchanger of rabbit β-intercalated cells, the mean change in anion exchange activity, dpHi/d t(where pHi is intracellular pH), was measured in response to lumen Cl− replacement with gluconate in perfused cortical collecting ducts (CCDs). β-Cell apical anion exchange was not affected by 15-min exposure to 0.2 mM lumen DIDS in the presence of 115 mM Cl−. In contrast, apical anion exchange was significantly inhibited by 0.1 mM lumen DIDS in the absence of Cl−. β-Cell apical anion exchange was unchanged by 15 mM maleic anhydride, 10 mM phenylglyoxal, 0.2 mM niflumic acid, 1 mM edecrin, 1 mM furosemide, 1 mM probenecid, or 0.1 mM diphenylamine-2-carboxylate. However, β-cell apical anion exchange was inhibited by α-cyano-4-hydroxycinnamic acid, with an IC50 of 2.4 mM. Substitution of either sulfate or gluconate for lumen Cl− resulted in a similar rate of alkalinization. Conversely, pHi was unchanged by substitution of sulfate for lumen gluconate, confirming the lack of transport of sulfate on the β-cell apical anion exchanger. Taken together, the results demonstrate a distinct “fingerprint” of the rabbit CCD β-cell apical anion exchanger that is unlike that of other known anion exchangers.

Stimulation of Cl- self exchange by intracellular HCO3- in rabbit cortical collecting duct

1989 ◽  
Vol 257 (1) ◽  
pp. C94-C101 ◽  
Author(s):  
K. Matsuzaki ◽  
J. B. Stokes ◽  
V. L. Schuster
Keyword(s):  
Anion Exchange ◽  
Intracellular Ph ◽  
Rate Coefficient ◽  
Collecting Duct ◽  
Cortical Collecting Duct ◽  
High K ◽  
Shift Experiment ◽  
Ethanesulfonic Acid ◽  
The Individual ◽  
Stimulation Of

In rabbit cortical collecting duct, Cl- self exchange accounts for most of the transepithelial Cl- tracer rate coefficient, KCl (nm/s); a small fraction is effected by Cl--HCO3- exchange and Cl- diffusion. We previously reported that changing from a CO2-free N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) bath to a 5% CO2-25 mM HCO3- bath stimulates Cl- self exchange. Here, we examine in further detail the individual components of the CO2-HCO3- system that stimulate KCl. Addition of 0.5% CO2 to a HEPES bath (final pH = 7.24) stimulated KCl by 70 +/- 19 nm/s, a delta KCl comparable to that induced by 1% CO2 (pH 7.12), 6% CO2 (pH 6.6), or 6% CO2-25 mM HCO3- (pH 7.4). The roles of intracellular pH (pHi) and HCO3- concentration were examined by clamping pHi using high K+ and nigericin. Increasing pHi from 6.9 to 7.6 in solutions without exogenous CO2 or HCO3- increased KCl by 71 +/- 17 nm/s. These results suggest that pHi might regulate anion exchange. However, during such a pHi-shift experiment, metabolically derived CO2 produces a concomitant change in intracellular HCO3- concentration [( HCO3-]i). To determine whether an increase in [HCO3-]i could stimulate Cl- self exchange, we replaced HEPES with 6% CO2-5 mM HCO3- isohydrically (pHi clamped at 6.9). With this increase in [HCO3-]i at constant pHi, KCl increased by 51 +/- 10 nm/s. These maneuvers had negligible effects on Cl- diffusion and Cl--HCO3- exchange. These experiments demonstrate that increases in cell [HCO3-] (or perhaps CO2) can stimulate transepithelial anion exchange.(ABSTRACT TRUNCATED AT 250 WORDS)

Cellular actions of cAMP on HCO3(-)-secreting cells of rabbit CCD: dependence on in vivo acid-base status

1994 ◽  
Vol 266 (4) ◽  
pp. F528-F535 ◽  
Author(s):  
C. Emmons ◽  
J. B. Stokes
Keyword(s):  
Anion Exchanger ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Disulfonic Acid ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Acid Base ◽  
Acid Base Status

HCO3- secretion by cortical collecting duct (CCD) occurs via beta-intercalated cells. In vitro CCD HCO3- secretion is modulated by both the in vivo acid-base status of the animal and by adenosine 3',5'-cyclic monophosphate (cAMP). To investigate the mechanism of cAMP-induced HCO3- secretion, we measured intracellular pH (pHi) of individual beta-intercalated cells of CCDs dissected from alkali-loaded rabbits perfused in vitro. beta-Intercalated cells were identified by demonstrating the presence of an apical anion exchanger (cell alkalinization in response to removal of lumen Cl-). After 180 min of perfusion to permit decrease of endogenous cAMP, acute addition of 0.1 mM 8-bromo-cAMP or 1 microM isoproterenol to the bath caused a transient cellular alkalinization (> 0.20 pH units). In the symmetrical absence of either Na+, HCO3-, or Cl-, cAMP produced no change in pHi. Basolateral dihydrogen 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (0.1 mM) for 15 min before cAMP addition also prevented this alkalinization. In contrast to the response of cells from alkali-loaded rabbits, addition of basolateral cAMP to CCDs dissected from normal rabbits resulted in an acidification of beta-intercalated cells (approximately 0.20 pH units). The present studies demonstrate the importance of the in vivo acid-base status of the animal in the regulation of CCD HCO3- secretion by beta-intercalated cells. The results identify the possible existence of a previously unrecognized Na(+)-dependent Cl-/HCO3- exchanger on the basolateral membrane of beta-intercalated cells in alkali-loaded rabbits.

Expression of isoforms of the Na+/H+exchanger in M-1 mouse cortical collecting duct cells

2002 ◽  
Vol 282 (4) ◽  
pp. F649-F654 ◽  
Author(s):  
C. Hill ◽  
A. N. Giesberts ◽  
S. J. White
Keyword(s):  
Intracellular Ph ◽  
Apical Membrane ◽  
Collecting Duct ◽  
Extracellular Nucleotides ◽  
Cortical Collecting Duct ◽  
Intracellular Acidification ◽  
Basolateral Membranes ◽  
Nephron Segments ◽  
Collecting Duct Cells ◽  
Nhe Isoforms

Na+/H+ exchanger (NHE) proteins perform a variety of functions in the kidney and are differentially distributed among nephron segments. The purpose of this study was to identify NHE isoforms in murine M-1 cells as a model of cortical collecting duct principal cells. It was found that mRNAs corresponding to NHE1, NHE2, and NHE4 are expressed in M-1 cells. NHE-dependent regulation of intracellular pH (pHi) was investigated in the absence of extracellular HCO[Formula: see text]. Application of a 20 mM NH4Cl pulse resulted in a reversible intracellular acidification from which recovery was partially inhibited by application of 1 mM amiloride to either the apical or the basolateral membranes and was abolished when amiloride was applied to both sides of the monolayers, which suggests that NHEs are expressed in both the apical and the basolateral cell membranes of M-1 cells. The purinergic agonists ATP and benzoylbenzoyl-ATP caused a reduction of pHi when applied to the apical membrane, which suggests pHi may be influenced by extracellular nucleotides in the luminal fluid of the cortical collecting duct.

Distribution of Cl-/HCO3- exchange and intercalated cells in rabbit cortical collecting duct

1994 ◽  
Vol 267 (6) ◽  
pp. F952-F964 ◽  
Author(s):  
I. D. Weiner ◽  
A. E. Weill ◽  
A. R. New
Keyword(s):  
B Cells ◽  
B Cell ◽  
Collecting Duct ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Base Exchange ◽  
Intercalated Cell ◽  
A Cells ◽  
A Cell ◽  
Exchange Activity

At least two cortical collecting duct (CCD) intercalated cell populations mediate HCO3- secretion and reabsorption. The present study examined the membrane location of intercalated cell Cl-/base exchange activity and the axial distribution of CCD intercalated cells. CCD were studied using in vitro microperfusion in CO2/HCO3(-)-containing solutions; intracellular pH was measured using 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. The A-type intercalated cell (A cell) and B-type intercalated cell (B cell) were identified functionally by the absence and presence of apical Cl-/HCO3- exchange activity, respectively. When a 0 mM Cl-, 0 mM HCO3- luminal solution was used, removal of Cl- from the peritubular solution caused intracellular alkalinization in all B cells. The alkalinization required neither extracellular Na+ nor changes in membrane potential. Peritubular 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) (10(-4) M) inhibited A cell but not B cell basolateral Cl-/base exchange activity. In comparison to studies performed with a 0 mM Cl- 0 mM HCO3- luminal solution, the use of a 0 mM Cl-, 25 mM HCO3- luminal solution inhibited both the identification and the magnitude of B cell basolateral Cl-/base exchange activity. When CCD from the inner and outer cortex were separately studied, only 7% of outer CCD intercalated cells were A cells, whereas 93% were B cells. In contrast, in the inner CCD, 58% of intercalated cells were A cells and 42% were B cells. Under stop-flow conditions, outer CCD alkalinized the luminal fluid, whereas inner CCD acidified the luminal fluid. These results indicate that all CCD intercalated cells possess basolateral Cl-/base exchange activity; however, A cell and B cell basolateral Cl-/base exchange activity differs, at least in terms of sensitivity to DIDS. Furthermore, there is axial heterogeneity in both intercalated cell type and function.

scholarly journals ENaC inhibition stimulates HCl secretion in the mouse cortical collecting duct. II. Bafilomycin-sensitive H+ secretion

2015 ◽  
Vol 309 (3) ◽  
pp. F259-F268 ◽  
Author(s):  
Masayoshi Nanami ◽  
Vladimir Pech ◽  
Yoskaly Lazo-Fernandez ◽  
Alan M. Weinstein ◽  
Susan M. Wall
Keyword(s):  
Intracellular Ph ◽  
Collecting Duct ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Transport Pathway ◽  
Anion Secretion ◽  
Intercalated Cell ◽  
Atpase Inhibitors ◽  
Hcl Secretion ◽  
Transepithelial Voltage

Epithelial Na+ channel (ENaC) blockade stimulates stilbene-sensitive conductive Cl− secretion in the mouse cortical collecting duct (CCD). This study's purpose was to determine the co-ion that accompanies benzamil- and DIDS-sensitive Cl− flux. Thus transepithelial voltage, VT, as well as total CO2 (tCO2) and Cl− flux were measured in CCDs from aldosterone-treated mice consuming a NaCl-replete diet. We reasoned that if stilbene inhibitors (DIDS) reduce conductive anion secretion they should reduce the lumen-negative VT. However, during ENaC blockade (benzamil, 3 μM), DIDS (100 μM) application to the perfusate reduced net H+ secretion, which increased the lumen-negative VT. Conversely, ENaC blockade alone stimulated H+ secretion, which reduced the lumen-negative VT. Application of an ENaC inhibitor to the perfusate reduced the lumen-negative VT, increased intercalated cell intracellular pH, and reduced net tCO2 secretion. However, benzamil did not change tCO2 flux during apical H+-ATPase blockade (bafilomycin, 5 nM). The increment in H+ secretion observed with benzamil application contributes to the fall in VT observed with application of this diuretic. As such, ENaC blockade reduces the lumen-negative VT by inhibiting conductive Na+ absorption and by stimulating H+ secretion by type A intercalated cells. In conclusion, 1) in CCDs from aldosterone-treated mice, benzamil application stimulates HCl secretion mediated by the apical H+-ATPase and a yet to be identified conductive Cl− transport pathway; 2) benzamil-induced HCl secretion is reversed with the application of stilbene inhibitors or H+-ATPase inhibitors to the perfusate; and 3) benzamil reduces VT not only by inhibiting conductive Na+ absorption, but also by stimulating H+ secretion.

Regulation of B-type intercalated cell apical anion exchange activity by CO2/ HCO 3 −

1998 ◽  
Vol 274 (6) ◽  
pp. F1086-F1094 ◽  
Author(s):  
Amy E. Milton ◽  
I. David Weiner
Keyword(s):  
Carbonic Anhydrase ◽  
B Cell ◽  
Anion Exchange ◽  
Collecting Duct ◽  
Cortical Collecting Duct ◽  
Base Exchange ◽  
Text Formation ◽  
Intracellular Alkalinization ◽  
Exchange Activity

The cortical collecting duct (CCD) B cell possesses an apical anion exchanger dissimilar to AE1, AE2, and AE3. The purpose of these studies was to characterize this transporter more fully by examining its regulation by CO2 and[Formula: see text]. We measured intracellular pH (pHi) in single intercalated cells of in vitro microperfused CCD using the fluorescent, pH-sensitive dye, 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). In the absence of extracellular CO2/[Formula: see text], luminal Cl− removal caused reversible intracellular alkalinization, identifying this transporter as a Cl−/base exchanger able to transport bases other than [Formula: see text]. Adding extracellular CO2/[Formula: see text]decreased B cell pHi while simultaneously increasing Cl−/base exchange activity. Since intracellular acidification inhibits AE1, AE2, and AE3, we examined mechanisms other than pHiby which the stimulation occurred. These studies showed that B cell apical anion exchange activity was CO2 stimulated and carbonic anhydrase dependent. Moreover, the stimulation was independent of luminal bicarbonate, luminal pH or pHi, and changes in buffer capacity. We conclude that the B cell possesses an apical Cl−/base exchanger whose activity is regulated by CO2-stimulated, carbonic anhydrase-dependent cytoplasmic [Formula: see text]formation.

NH4Cl activates AE2 anion exchanger in Xenopus oocytes at acidic pHi

1997 ◽  
Vol 272 (4) ◽  
pp. C1232-C1240 ◽  
Author(s):  
B. D. Humphreys ◽  
M. N. Chernova ◽  
L. Jiang ◽  
Y. Zhang ◽  
S. L. Alper
Keyword(s):  
Intracellular Ph ◽  
Anion Exchanger ◽  
Sodium Acetate ◽  
Xenopus Oocytes ◽  
Renal Medulla ◽  
Disulfonic Acid ◽  
Acidic Ph ◽  
Intracellular Acidification ◽  
Extracellular Acidification ◽  
Exchange Activity

In the course of experiments to define regulation by intracellular pH (pHi) of the AE2 anion exchanger expressed in Xenopus oocytes, we discovered an unexpected regulation of AE2 by NH4+. Intracellular acidification produced by extracellular acidification or produced by equimolar substitution of NaCl with sodium acetate each inhibited AE2 activity. In contrast, intracellular acidification by equimolar substitution of NaCl with NH4Cl activated AE2-associated, trans-anion-dependent, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid-sensitive 36Cl- influx and efflux. Regulation by NH4+ was isoform specific, since neither erythroid nor kidney AE1 was activated. AE2 activation was maximal at <5 mM NH4Cl; was not mimicked by extracellular KCl, chloroquine, or polyamines; and was insensitive to amiloride, bumetanide, barium, and gadolinium. Whether NH4Cl acts directly on AE2 or on another target remains to be determined. Activation of AE2 by NH4+ may serve to sustain Cl-/HCO3- exchange activity in the presence of acidic pH in renal medulla, colon, abscesses, and other AE2-expressing acidic locales exposed to elevated NH4+ concentration.

scholarly journals Potassium depletion increases proton pump (H+-ATPase) activity in intercalated cells of cortical collecting duct

2000 ◽  
Vol 279 (1) ◽  
pp. F195-F202 ◽  
Author(s):  
Randi B. Silver ◽  
Sylvie Breton ◽  
Dennis Brown
Keyword(s):  
Plasma Membrane ◽  
Atpase Activity ◽  
Collecting Duct ◽  
Proton Pumps ◽  
Intercalated Cells ◽  
Cortical Collecting Duct ◽  
Collecting Ducts ◽  
Acid Load ◽  
Collecting Tubules ◽  
Atpase Staining

Intercalated cells (ICs) from kidney collecting ducts contain proton-transporting ATPases (H+-ATPases) whose plasma membrane expression is regulated under a variety of conditions. It has been shown that net proton secretion occurs in the distal nephron from chronically K+-depleted rats and that upregulation of tubular H+- ATPase is involved in this process. However, regulation of this protein at the level of individual cells has not so far been examined. In the present study, H+-ATPase activity was determined in individually identified ICs from control and chronically K+-depleted rats (9–14 days on a low-K+ diet) by monitoring K+- and Na+-independent H+ extrusion rates after an acute acid load. Split-open rat cortical collecting tubules were loaded with the intracellular pH (pHi) indicator 2′,7′-bis(2-carboxyethyl)-5(6)-carboxyfluorescein, and pHiwas determined by using ratiometric fluorescence imaging. The rate of pHi recovery in ICs in response to an acute acid load, a measure of plasma membrane H+-ATPase activity, was increased after K+ depletion to almost three times that of controls. Furthermore, the lag time before the start of pHirecovery after the cells were maximally acidified fell from 93.5 ± 13.7 s in controls to 24.5 ± 2.1 s in K+-depleted rats. In all ICs tested, Na+- and K+-independent pHi recovery was abolished in the presence of bafilomycin (100 nM), an inhibitor of the H+-ATPase. Analysis of the cell-to-cell variability in the rate of pHi recovery reveals a change in the distribution of membrane-bound proton pumps in the IC population of cortical collecting duct from K+-depleted rats. Immunocytochemical analysis of collecting ducts from control and K+-depleted rats showed that K+-depletion increased the number of ICs with tight apical H+ATPase staining and decreased the number of cells with diffuse or basolateral H+-ATPase staining. Taken together, these data indicate that chronic K+ depletion induces a marked increase in plasma membrane H+ATPase activity in individual ICs.

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