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.

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.

Apical membrane localization of ouabain-sensitive K(+)-activated ATPase activities in rat distal colon

1991 ◽  
Vol 261 (6) ◽  
pp. G1005-G1011 ◽  
Author(s):  
J. R. Del Castillo ◽  
V. M. Rajendran ◽  
H. J. Binder
Keyword(s):  
Atpase Activity ◽  
Parietal Cell ◽  
Colonic Mucosa ◽  
Apical Membrane ◽  
Distal Colon ◽  
Proximal Colon ◽  
Gastric Parietal Cell ◽  
Fold Enrichment ◽  
Rat Distal Colon ◽  
Apical Membranes

This study sought to establish the presence of K(+)-activated adenosinetriphosphatase (ATPase) activity in the colonic mucosa of the rat distal colon. K(+)-activated ATPase activity was present in apical membranes but not in basolateral membranes. K(+)-activated ATPase activity in apical membranes represented an approximate 10-fold enrichment compared with that in the homogenate. Na(+)-K(+)-activated ATPase activity was also present in homogenate but was enriched less than fourfold in apical membranes. K(+)-activated ATPase activity in apical membranes had both ouabain-sensitive and ouabain-insensitive components. In contrast, Na(+)-K(+)-activated ATPase activity was completely inhibited by ouabain. Similar half-maximal concentrations for K+ and pH activation curves were found for both ouabain-sensitive and ouabain-insensitive fractions. In addition to K+, the ouabain-sensitive fraction of K(+)-activated ATPase activity was stimulated by Rb+, NH+4, and Cs+, whereas the ouabain-insensitive fraction was activated only by Rb+. K(+)-activated ATPase activity was significantly inhibited by vanadate but not by N-ethylmaleimide or omeprazole. In the proximal colon, in contrast to the distal colon, active K+ absorption is not present, and K(+)-activated ATPase is approximately 20% of that in the distal colon. These studies demonstrate that K(+)-activated ATPase is present in apical membranes of rat distal colon and permit the speculation that this enzyme represents a unique and distinct ATPase (compared with either Na(+)-K(+)-ATPase or gastric parietal cell K(+)-ATPase) and is likely linked closely to the active K+ absorptive process present in this epithelium.

Role of short-chain fatty acids in colonic HCO3secretion

2005 ◽  
Vol 288 (6) ◽  
pp. G1217-G1226 ◽  
Author(s):  
Sadasivan Vidyasagar ◽  
Christian Barmeyer ◽  
John Geibel ◽  
Henry J. Binder ◽  
Vazhaikkurichi M. Rajendran
Keyword(s):  
Colonic Mucosa ◽  
Apical Membrane ◽  
Membrane Vesicles ◽  
Anion Channel ◽  
Short Chain Fatty Acids ◽  
Short Chain ◽  
Channel Activity ◽  
Colonic Crypts ◽  
Ph Stat

Luminal isobutyrate, a relatively poor metabolized short-chain fatty acid (SCFA), induces HCO3secretion via a Cl-independent, DIDS-insensitive, carrier-mediated process as well as inhibiting both Cl-dependent and cAMP-induced HCO3secretion. The mechanism(s) responsible for these processes have not been well characterized. HCO3secretion was measured in isolated colonic mucosa mounted in Lucite chambers using pH stat technique and during microperfusion of isolated colonic crypts.14C-labeled butyrate,14C-labeled isobutyrate, and36Cl uptake were also determined by apical membrane vesicles (AMV) isolated from surface and/or crypt cells. Butyrate stimulation of Cl-independent, DIDS-insensitive 5-nitro-3-(3-phenylpropyl-amino)benzoic acid-insensitive HCO3secretion is greater than that by isobutyrate, suggesting that both SCFA transport and metabolism are critical for HCO3secretion. Both lumen and serosal 25 mM butyrate inhibit cAMP-induced HCO3secretion to a comparable degree (98 vs. 90%). In contrast, Cl-dependent HCO3secretion is downregulated by lumen 25 mM butyrate considerably more than by serosal butyrate (98 vs. 37%). Butyrate did not induce HCO3secretion in isolated microperfused crypts, whereas an outward-directed HCO3gradient-driven induced14C-butyrate uptake by surface but not crypt cell AMV. Both36Cl/HCO3exchange and potential-dependent36Cl movement in AMV were inhibited by 96–98% by 20 mM butyrate. We conclude that 1) SCFA-dependent HCO3secretion is the result of SCFA transport across the apical membrane via a SCFA/HCO3exchange more than intracellular SCFA metabolism; 2) SCFA-dependent HCO3secretion is most likely a result of an apical membrane SCFA/HCO3exchange in surface epithelial cells; 3) SCFA downregulates Cl-dependent and cAMP-induced HCO3secretion secondary to SCFA inhibition of apical membrane Cl/HCO3exchange and anion channel activity, respectively.

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.

Corticosteroid alteration of active electrolyte transport in rat distal colon

1983 ◽  
Vol 245 (5) ◽  
pp. G668-G675 ◽  
Author(s):  
E. S. Foster ◽  
T. W. Zimmerman ◽  
J. P. Hayslett ◽  
H. J. Binder
Keyword(s):  
Colonic Mucosa ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Distal Colon ◽  
Transport Processes ◽  
Electrolyte Transport ◽  
Sodium Absorption ◽  
Rat Distal Colon ◽  
Potassium Absorption ◽  
Potassium Secretion

To determine the effect of corticosteroids on active transport processes, unidirectional fluxes of 22Na, 36Cl, and 42K were measured under short-circuit conditions across isolated stripped distal colonic mucosa of the rat in control, secondary hyperaldosterone, and dexamethasone-treated animals. In controls net sodium and chloride fluxes (JNanet and JClnet) and short-circuit current (Isc) were 6.6 +/- 2.2, 7.6 +/- 1.6, and 1.3 +/- 0.2 mu eq X h-1 X cm-2, respectively. Although aldosterone increased Isc to 7.3 +/- 0.5 mu eq X h-1 X cm-2, JNanet (6.9 +/- 0.7 mu eq X h-1 X cm-2) was not altered and JClnet was reduced to 0 compared with controls. Dexamethasone also stimulated Isc but did not inhibit JClnet. In Cl-free Ringer both aldosterone and dexamethasone produced significant and equal increases in JNanet and Isc. Theophylline abolished JNanet in control animals but not in the aldosterone group. Aldosterone reversed net potassium absorption (0.58 +/- 0.11 mu eq X h-1 X cm-2) to net potassium secretion (-0.94 +/- 0.08 mu eq X h-1 X cm-2). Dexamethasone reduced net potassium movement to 0 (-0.04 +/- 0.12 mu eq X h-1 X cm-2). These studies demonstrate that 1) corticosteroids stimulate electrogenic sodium absorption and 2) aldosterone, but not dexamethasone, inhibits neutral NaCl absorption and stimulates active potassium secretion. The effects of mineralocorticoids and glucocorticoids on electrolyte transport are not identical and may be mediated by separate and distinct mechanisms.

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.

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