T1731 Apical Membrane Intermediate Conductance K+ (KCNN4) Channels Provide Driving Force for CA2+-Activated Cl−Secretion in Rat Distal Colon

2009 ◽  
Vol 136 (5) ◽  
pp. A-568
Author(s):  
Nanda Kumar Navalpur Shanmugam ◽  
Vazhaikkurichi M. Rajendran
Keyword(s):  
Driving Force ◽  
Apical Membrane ◽  
Distal Colon ◽  
Cl Secretion ◽  
Rat Distal Colon

Distribution and regulation of apical Cl/anion exchanges in surface and crypt cells of rat distal colon

1999 ◽  
Vol 276 (1) ◽  
pp. G132-G137 ◽  
Author(s):  
Vazhaikkurichi M. Rajendran ◽  
Henry J. Binder
Keyword(s):  
Spatial Distribution ◽  
Concentration Gradient ◽  
Apical Membrane ◽  
Membrane Vesicles ◽  
Distal Colon ◽  
Ph Homeostasis ◽  
Rat Distal Colon ◽  
Na Depletion ◽  
Crypt Cells ◽  
Apical Membrane Vesicles

Na depletion inhibits electroneutral Na-Cl absorption in intact tissues and Na/H exchange in apical membrane vesicles (AMV) of rat distal colon. Two anion (Cl/HCO3 and Cl/OH) exchanges have been identified in AMV from surface cells of rat distal colon. To determine whether Cl/HCO3 and/or Cl/OH exchange is responsible for vectorial Cl movement, this study examined the spatial distribution and the effect of Na depletion on anion-dependent 36Cl uptake by AMV in rat distal colon. These studies demonstrate that HCO3 concentration gradient-driven36Cl uptake (i.e., Cl/HCO3 exchange) is 1) primarily present in AMV from surface cells and 2) markedly reduced by Na depletion. In contrast, OH concentration gradient-driven36Cl uptake (i.e., Cl/OH exchange) present in both surface and crypt cells is not affected by Na depletion. In Na-depleted animals HCO3 also stimulates36Cl via Cl/OH exchange with low affinity. These results suggest that Cl/HCO3 exchange is responsible for vectorial Cl absorption, whereas Cl/OH exchange is involved in cell volume and/or cell pH homeostasis.

Cl-HCO3 and Cl-OH exchanges mediate Cl uptake in apical membrane vesicles of rat distal colon

1993 ◽  
Vol 264 (5) ◽  
pp. G874-G879 ◽  
Author(s):  
V. M. Rajendran ◽  
H. J. Binder
Keyword(s):  
Apical Membrane ◽  
Kinetic Constant ◽  
Membrane Vesicles ◽  
Distal Colon ◽  
Ph Gradient ◽  
Disulfonic Acid ◽  
Ph Homeostasis ◽  
Rat Distal Colon ◽  
Hyperbolic Curve ◽  
Apical Membrane Vesicles

This study describes Cl-HCO3 and Cl-OH exchanges as the mechanism for Cl uptake by apical membrane vesicles (AMV) of rat distal colon. Although HCO3 gradient-stimulated 36Cl uptake was additionally stimulated by the additional presence of a pH gradient, pH gradient-stimulated 36Cl uptake was not further enhanced by a HCO3 gradient. HCO3 gradient-stimulated and OH gradient-stimulated 36Cl uptake was not inhibited by voltage clamping, with K and its ionophore valinomycin, but was inhibited by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, an anion exchange inhibitor, with an apparent inhibitory constant of 7.8 and 106.0 microM, respectively. Increasing intravesicular OH concentration in the absence of HCO3 (with fixed extravesicular Cl concentration) yielded a sigmoidal curve for 36Cl uptake. In contrast, increasing intravesicular OH concentration in the presence of equimolar intra- and extravesicular HCO3 (25 mM) yielded a saturable hyperbolic curve. Increasing extravesicular Cl concentration saturated both HCO3 gradient-stimulated and OH gradient-stimulated 36Cl uptake, with a kinetic constant for Cl of approximately 11.9 and 22.6 mM, respectively. We conclude that Cl uptake in AMV of rat distal colon occurs via two separate anion (Cl-HCO3 and Cl-OH) exchange processes. We speculate that one of these two anion exchanges may be responsible for transcellular Cl movement, while the other may be important in the regulation of intracellular pH homeostasis.

Modulation of secretagogue-induced chloride secretion by intracellular bicarbonate

1994 ◽  
Vol 266 (5) ◽  
pp. G929-G934 ◽  
Author(s):  
P. C. Dagher ◽  
T. Z. Morton ◽  
C. S. Joo ◽  
A. Taglietta-Kohlbrecher ◽  
R. W. Egnor ◽  
...  
Keyword(s):  
Cell Line ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Tumor Cell Line ◽  
Distal Colon ◽  
Rat Colon ◽  
Sprague Dawley ◽  
T84 Cells ◽  
Cl Secretion ◽  
Rat Distal Colon

We have previously demonstrated inhibition of basal Cl- secretion by intracellular bicarbonate concentration ([HCO3-]i) in rat distal colon. We now examined whether secretagogue-induced Cl- secretion is inhibited by [HCO3-]i as well. Stripped segments of distal colon from male Sprague-Dawley rats and the colon tumor cell line T84 were studied. Flux measurements were performed in the Ussing chamber under short-circuit conditions. [HCO3-]i was calculated from intracellular pH (pHi) values that were estimated with the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein. Dibutyryl adenosine 3',5'-cyclic monophosphate (cAMP) and carbachol were used as secretagogues. In both distal colon and T84 cells, [HCO3-]i did not affect cAMP-induced Cl- secretion. However, carbachol-induced secretion was inhibited by [HCO3-]i; in rat colon, Cl- secretion decreased from 2.3 to 1.5 mueq.cm-2.h-1 when [HCO3-]i was increased from 15.0 to 28.4 mM (P < 0.05). In T84 cells, the change in short-circuit current decreased from 8.1 to 1.1 microA/cm2 over a range of [HCO3-]i from 0 to 15.6 mM (P < 0.001). We conclude that [HCO3-]i is an important modulator of carbachol-stimulated Cl- secretion in both rat distal colon and the T84 cell line. cAMP-mediated secretion is not affected by [HCO3-]i.

Characterization of apical membrane Cl-dependent Na/H exchange in crypt cells of rat distal colon

2001 ◽  
Vol 280 (3) ◽  
pp. G400-G405 ◽  
Author(s):  
Vazhaikkurichi M. Rajendran ◽  
John Geibel ◽  
Henry J. Binder
Keyword(s):  
Channel Blocker ◽  
Apical Membrane ◽  
Distal Colon ◽  
Proton Gradient ◽  
Disulfonic Acid ◽  
High Dose ◽  
Cl Channel ◽  
Rat Distal Colon ◽  
Na Uptake ◽  
Crypt Cells

A novel Cl-dependent Na/H exchange (Cl-NHE) has been identified in apical membranes of crypt cells of rat distal colon. The presence of Cl is required for both outward proton gradient-driven Na uptake in apical membrane vesicles (AMV) and Na-dependent intracellular pH recovery from an acid load in the crypt gland. The present study establishes that Cl-dependent outward proton gradient-driven 22Na uptake 1) is saturated with increasing extravesicular Na concentration with a Michaelis constant ( K m) for Na of ∼24.2 mM; 2) is saturated with increasing outward H concentration gradient with a hyperbolic curve and a K m for H of ∼1.5 μM; 3) is inhibited by the Na/H exchange (NHE) inhibitors amiloride, ethylisopropylamiloride, and HOE-694 with an inhibitory constant ( K i) of ∼480.2, 1.1, and 9.5 μM, respectively; 4) is inhibited by 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid, an anion exchange inhibitor at low concentration and a Cl channel blocker at high dose, and by 5-nitro-2(3-phenylpropylamino)benzoic acid, a Cl channel blocker, with a K i of ∼280.6 and 18.3 μM, respectively; and 5) substantially stimulated Cl-NHE activity by dietary Na depletion, which increases plasma aldosterone and inhibits NHE in surface cell AMV. These properties of Cl-NHE are distinct from those of NHE1, NHE2, and NHE3 isoforms that are present in colonic epithelial cells; thus these results suggest that the colonic crypt cell Cl-NHE is a novel NHE isoform.

HCO3ȡ secretion occurs via multiple and distinct apical membrane transport processes in rat distal colon

2003 ◽  
Vol 124 (4) ◽  
pp. A308
Author(s):  
Sadasivan Vidyasagar ◽  
Christian Barmeyer ◽  
Henry J. Binder ◽  
Vazhaikkurichi M. Rajendran
Keyword(s):  
Membrane Transport ◽  
Apical Membrane ◽  
Distal Colon ◽  
Transport Processes ◽  
Rat Distal Colon

Three distinct mechanisms of HCO3−secretion in rat distal colon

2004 ◽  
Vol 287 (3) ◽  
pp. C612-C621 ◽  
Author(s):  
Sadasivan Vidyasagar ◽  
Vazhaikkurichi M. Rajendran ◽  
Henry J. Binder
Keyword(s):  
Colonic Mucosa ◽  
Short Chain Fatty Acid ◽  
Channel Blocker ◽  
Apical Membrane ◽  
Anion Channel ◽  
Distal Colon ◽  
Chain Fatty Acid ◽  
Direct Examination ◽  
Rat Distal Colon ◽  
Ph Stat

HCO3−secretion has long been recognized in the mammalian colon, but it has not been well characterized. Although most studies of colonic HCO3−secretion have revealed evidence of lumen Cl−dependence, suggesting a role for apical membrane Cl−/HCO3−exchange, direct examination of HCO3−secretion in isolated crypt from rat distal colon did not identify Cl−-dependent HCO3−secretion but did reveal cAMP-induced, Cl−-independent HCO3−secretion. Studies were therefore initiated to determine the characteristics of HCO3−secretion in isolated colonic mucosa to identify HCO3−secretion in both surface and crypt cells. HCO3−secretion was measured in rat distal colonic mucosa stripped of muscular and serosal layers by using a pH stat technique. Basal HCO3−secretion (5.6 ± 0.03 μeq·h−1·cm−2) was abolished by removal of either lumen Cl−or bath HCO3−; this Cl−-dependent HCO3−secretion was also inhibited by 100 μM DIDS (0.5 ± 0.03 μeq·h−1·cm−2) but not by 5-nitro-3-(3-phenylpropyl-amino)benzoic acid (NPPB), a Cl−channel blocker. 8-Bromo-cAMP induced Cl−-independent HCO3−secretion (and also inhibited Cl−-dependent HCO3−secretion), which was inhibited by NPPB and by glibenclamide, a CFTR blocker, but not by DIDS. Isobutyrate, a poorly metabolized short-chain fatty acid (SCFA), also induced a Cl−-independent, DIDS-insensitive, saturable HCO3−secretion that was not inhibited by NPPB. Three distinct HCO3−secretory mechanisms were identified: 1) Cl−-dependent secretion associated with apical membrane Cl−/HCO3−exchange, 2) cAMP-induced secretion that was a result of an apical membrane anion channel, and 3) SCFA-dependent secretion associated with an apical membrane SCFA/HCO3−exchange.

Apical NHE isoforms differentially regulate butyrate-stimulated Na absorption in rat distal colon

2003 ◽  
Vol 285 (5) ◽  
pp. C1246-C1254 ◽  
Author(s):  
Selvi Krishnan ◽  
Vazhaikkurichi M. Rajendran ◽  
Henry J. Binder
Keyword(s):  
Voltage Clamp ◽  
Apical Membrane ◽  
Distal Colon ◽  
Dibutyryl Camp ◽  
Differential Effects ◽  
Rat Distal Colon ◽  
Nhe Isoforms ◽  
Specific Inhibitors ◽  
Stimulation Of

Bicarbonate and butyrate stimulate electroneutral Na absorption via apical membrane Na-H exchange (NHE) in rat distal colon. cAMP downregulates NHE-3 isoform and inhibits HCO3-dependent, but not butyrate-dependent, Na absorption. This study sought to determine whether 1) the apical membrane NHE-2 and NHE-3 isoforms differentially mediated HCO3- and butyrate-dependent Na absorption, and 2) cAMP had different effects on NHE-2 and NHE-3 isoforms. The effect of specific inhibitors of NHE-2 and NHE-3 isoforms (50 μM HOE 694 and 2 μM S3226, respectively) on unidirectional 22Na transepithelial fluxes performed across isolated mucosa from rat distal colon under voltage-clamp conditions was examined. HCO3 stimulation of Na absorption was inhibited by EIPA, a nonspecific inhibitor of all NHE isoforms, by S3226 and dibutyryl cAMP but not by HOE 694. In contrast, butyrate stimulation of Na absorption was not altered by dibutyryl cAMP and was not inhibited by HOE 694 in the absence of dibutyryl cAMP, but in the presence of dibutyryl cAMP was HOE694 sensitive. In contrast, S3226 inhibited butyrate-stimulated Na absorption in the absence of dibutyryl cAMP, but not in its presence. We conclude that 1) HCO3-stimulated Na absorption is mediated solely by NHE-3 isoform, whereas butyrate-stimulated Na absorption is mediated by either NHE-3 or NHE-2 isoform, and 2) dibutyryl cAMP selectively inhibits NHE-3 isoform but stimulates NHE-2 isoform. Dibutyryl cAMP does not inhibit butyrate-stimulated Na absorption as a result of its differential effects on NHE-2 and NHE-3 isoforms.

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