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

Vazhaikkurichi M. Rajendran and Henry J. Binder

Departments of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06520


    ABSTRACT
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Abstract
Introduction
Materials and methods
Results
Discussion
References

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-driven 36Cl 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-driven 36Cl 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 stimulates 36Cl 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.

aldosterone; cell functions; sodium depletion; vectorial ion movement; vesicle uptake


    INTRODUCTION
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Abstract
Introduction
Materials and methods
Results
Discussion
References

ELECTRONEUTRAL CHLORIDE absorption is both Na and HCO3 dependent and is the primary mechanism of Cl absorption in the rat distal colon (4). On the basis of studies in both intact tissue and apical membrane vesicles (AMV), the general consensus has been that electroneutral Na-Cl absorption is the result of the coupling of Na/H exchange and Cl/HCO3 exchange (3, 9-11, 14, 15).

Previous studies (15) of Cl/anion exchange concluded that Cl/HCO3 and Cl/OH exchanges are distinct and separate anion exchanges. This conclusion was based on several observations. First, in the absence of HCO3, OH concentration ([OH]) gradient-driven 36Cl uptake yielded a sigmoidal curve as a function of [OH] consistent with the participation of two Cl transport systems on these apical membranes (15). Second, in the presence of HCO3, [OH] gradient-driven 36Cl uptake was hyperbolic as a function of [OH] (15). Taken together, these two findings suggested that the [OH] gradient stimulates 36Cl uptake via both Cl/OH and Cl/HCO3 exchanges, while the HCO3 concentration ([HCO3]) gradient stimulates 36Cl uptake only via Cl/HCO3 exchange. Third, there was a 20-fold difference in the inhibitor constant (Ki) for DIDS for Cl/OH and Cl/HCO3 exchanges (15). Fourth, the Michaelis constant (Km) for Cl for these two Cl/anion exchanges also significantly differed (15).

Na depletion inhibits electroneutral Na-Cl absorption as well as stimulating electrogenic Na absorption in rat distal colon (4, 10, 12). Studies with AMV isolated from normal and Na-depleted animals demonstrated that Na depletion both inhibited [H] gradient-driven 22Na uptake (i.e., Na/H exchange) and induced amiloride-sensitive apical Na channels (17). Because preliminary studies (18) demonstrated that Na depletion partially inhibited Cl/anion exchange in AMV from rat distal colon, these present experiments were designed to determine whether Na depletion altered Cl/HCO3 and/or Cl/OH exchanges.

The present study was initiated to identify the colonic Cl/anion exchange activity responsible for vectorial Cl movement. In this study, the spatial distribution in surface and crypt cells and the Na depletion regulation of Cl/anion exchanges in AMV were established. The surface cell-specific localization and inhibition of Cl/HCO3 exchange by Na depletion suggests that Cl/HCO3 exchange is involved in vectorial Cl movement, while Cl/OH exchange may be responsible for cell volume and/or cell pH homeostasis in rat distal colon.


    MATERIALS AND METHODS
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Abstract
Introduction
Materials and methods
Results
Discussion
References

Surface and crypt cell isolation. Surface and crypt cells were isolated from the distal colon of normal and Na-depleted rats (200- to 250-g Sprague-Dawley rats) by divalent chelation techniques, as described previously (16). Normal animals were given Purina rat chow, while Na-free animals were fed an Na-free diet (20 g/day) for 6-7 days, as described previously, which results in elevated serum aldosterone levels (10, 12).1 In brief, everted colonic segments were incubated in isolation buffer that contained 112 mM NaCl, 5 mM KCl, 30 mM EDTA, 20 mM HEPES, and 0.5 mM dithiothreitol. Surface-to-crypt sequential fractions were isolated by shaking and incubating (6 times for 10 min) at 37°C. Surface and crypt cells were sedimented by centrifugation (Beckman GS-6KR; GH-3.8 rotor) at 3,000 g for 5 min and 5,000 g for 2 min, respectively. Surface and crypt cells were distinguished based on ouabain-insensitive and -sensitive K-ATPase activities, respectively (19).

AMV preparation. AMV were prepared from surface and crypt cells by the Percoll gradient and differential centrifugation method of Stieger et al. (21), as described previously (16, 17). Purity of AMV was validated by a 9- to 10-fold enrichment of K-ATPase activity (6). Protein was assayed by the method of Lowry et al. (13).

Uptake studies. Uptake of 36Cl (NEN, Boston, MA) was performed by the rapid filtration technique, as described previously (15). Both [HCO3] and [OH] gradient-driven 36Cl uptake were linear for at least up to 15 s. As a result, 36Cl uptake was characterized at 12 s. Values are means ± SE of triplicate assays; SE <5% are not shown. All experiments were repeated with at least three different membrane preparations.


    RESULTS
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Abstract
Introduction
Materials and methods
Results
Discussion
References

Previous studies (15) of 36Cl uptake in AMV prepared from distal colon of normal rats established the presence of both Cl/HCO3 and Cl/OH exchanges and provided evidence that these two Cl/anion exchanges are separate and distinct transport processes. Because Na depletion inhibits active electroneutral Na-Cl absorption in in vitro ion flux studies performed under voltage-clamp conditions across isolated mucosa of rat distal colon (10, 12), studies of anion-driven 36Cl uptake were performed in AMV prepared from distal colon of both normal and Na-depleted rats. Figure 1 presents outward [HCO3] gradient (Fig. 1A) and outward [OH] (Fig. 1B) gradient-driven 36Cl uptake in AMV from distal colon of normal and Na-depleted rats. Similar to our earlier observations (15), both [HCO3] gradient and [OH] gradient stimulated 36Cl uptake in AMV from normal animals (Fig. 1). In AMV from Na-depleted animals, both [HCO3] and [OH] gradient-driven 36Cl uptakes were markedly reduced (Fig. 1). Although Na depletion inhibited both Cl/HCO3 and Cl/OH exchanges, the inhibition (70%) of Cl/HCO3 exchange was significantly greater than that (47%) of Cl/OH exchange. These observations are consistent with the possibility that Na depletion selectively inhibited Cl/HCO3 exchange, and, as a consequence, Cl uptake in the Na-depleted animals occurs primarily via Cl/OH and not via Cl/HCO3 exchange.


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Fig. 1.   Effect of Na depletion on Cl/HCO3 and Cl/OH exchanges. Apical membrane vesicles (AMV) from distal colon of normal (circles) and Na-depleted (squares) rats were preloaded with 50 mM HEPES-Tris (pH 7.5) containing 10 mM N-methyl-D-glucamine (NMG) gluconate and 150 mM of either KHCO3 (A) or potassium gluconate (B). A: HCO3 gradient. Uptake of 36Cl was measured for 0.2-90 min by incubating the KHCO3 AMV in medium containing 50 mM HEPES-Tris (pH 7.5), 10 mM 36Cl-NMG, and 150 mM of either potassium gluconate (filled symbols) or KHCO3 (open symbols). B: OH gradient. Uptake of 36Cl was measured for 0.2-90 min by incubating potassium gluconate AMV in medium containing 150 mM potassium gluconate, 10 mM 36Cl-NMG, and 50 mM of either MES-Tris (pH 5.5) (filled symbols) or HEPES-Tris (pH 7.5) (open symbols). HCO3-containing media were gassed with 17% CO2 balanced with O2 for 30 min, and the uptakes was measured under 17% CO2 atmosphere. All media contained 25 µM valinomycin and 0.8% ethanol.

To provide additional support for the differential inhibition of Cl/HCO3 and Cl/OH exchanges by Na depletion, we studied the effect of increasing outward alkaline pH gradient on 36Cl uptake in AMV from both normal and Na-depleted animals. In this study, intravesicular pH was increased, while extravesicular pH was maintained constant. As shown in Fig. 2A, outward pH gradients of 0.3 and 0.6 U stimulated 36Cl uptake by 0.25 and 0.37 nmol · mg protein-1 · 12 s-1 in AMV from normal animals, while in Na-depleted AMV 36Cl uptake was not stimulated at these pH gradients. In AMV from Na-depleted animals, the threshold of intravesicular pH for stimulation of 36Cl uptake was 6.4. Plotting these 36Cl uptakes as a function of [OH] gradient yielded a sigmoidal curve in AMV from normal animals (indicating the presence of two Cl transport processes) but a hyperbolic curve for 36Cl uptake in AMV from Na-depleted animals consistent with the presence of a single Cl transport system (Fig. 2B). These results suggest that one of the Cl/anion exchanges (i.e., Cl/HCO3 exchange) in AMV is not expressed (or is functionally inactive) in Na-depleted animals.2


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Fig. 2.   Effect of pH gradients. A: AMV from normal (open circle ) and Na-depleted (bullet ) rats were preloaded with 150 mM potassium gluconate, 5 mM NMG gluconate, and 50 mM of either MES-Tris (pH 5.5, 5.8, 6.14, 6.44, 6.74, 6.91, or 7.14) or HEPES-Tris (pH 7.26, 7.5, or 7.96). Uptake of 36Cl was measured for 12 s by incubating AMV in medium containing 150 mM potassium gluconate, 5 mM 36Cl-NMG, and 50 mM MES-Tris (pH 5.5). Uptake was also measured in presence of 1 mM DIDS. Absolute values presented were calculated by subtracting uptake in presence of DIDS from that in its absence. All media contained 25 µM valinomycin and 0.8% ethanol. B: uptake of 36Cl presented as function of pH in A is presented as function of OH concentration ([OH]) gradient for normal (open circle ) and Na-depleted (bullet ) animals.

Kinetic studies were therefore performed to determine whether HCO3 gradient-driven 36Cl uptake seen in AMV from Na-depleted animals (Fig. 1A) represents uptake via Cl/HCO3 and/or Cl/OH exchanges. As shown in Fig. 3, outward [HCO3] gradients of 2.5 and 5 mM stimulated 36Cl uptake in AMV from normal animals, but not in AMV from Na-depleted animals. However, 36Cl uptake in AMV from Na-depleted animals was stimulated by [HCO3] gradient at 7.5 mM. Lineweaver-Burk plot analyses of these data revealed an apparent Km for HCO3 of 6.8 ± 1.6 and 28.2 ± 3.3 mM for AMV from normal and aldosterone animals, respectively, and maximal velocity (Vmax) for Cl of 2.6 ± 0.6 and 0.8 ± 0.2 nmol · mg protein-1 · 12 s-1 for AMV from normal and aldosterone animals, respectively. Thus Na depletion is associated with a fivefold increase in Km for HCO3. The demonstrated Na depletion-mediated decrease in affinity for HCO3 shown in Fig. 3 is consistent with two alternate possibilities: 1) Na depletion had solely altered Cl/HCO3 exchange so that the affinity of Cl/HCO3 exchange for HCO3 was substantially decreased or 2) Na depletion had markedly reduced Cl/HCO3 exchange and that, in the absence of Cl/HCO3 exchange, HCO3 was transported via Cl/OH exchange, whose affinity for HCO3 is less than that of Cl/HCO3 exchange.


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Fig. 3.   Effect of HCO3 concentration ([HCO3]) gradients. AMV from normal (open circle ) and Na-depleted (bullet ) rat colon were preloaded with 50 mM HEPES-Tris (pH 7.5), 5 mM NMG gluconate, and varying concentrations of KHCO3 (0 or 4.5 mM 0.5% CO2; 7.5 mM 1% CO2; 13.6 mM 1.5% CO2; 31.8 mM 3.5% CO2; 45.4 mM 5% CO2; 68.1 mM 7.5% CO2; 90.8 mM 10% CO2; 154 mM 17% CO2) and potassium gluconate (in mM: 50, 145.5, 142.5, 136.4, 118.2, 104.6, 81.9, 59.2, or 0). Uptake of 36Cl was measured for 12 s by incubating AMV in incubation medium containing 150 mM potassium gluconate, 5 mM 36Cl-NMG, and 50 mM HEPES-Tris (pH 7.5). Vesicular [HCO3] were maintained by gassing with appropriate CO2-O2 mixture for 30 min. Uptake was measured under appropriate gas atmosphere. Uptake was also measured in presence of 1 mM DIDS. Absolute values presented were calculated by subtracting uptake obtained in presence of DIDS from that in its absence. All media contained 25 µM valinomycin and 0.8% ethanol.

To distinguish between these two possibilities additional kinetic experiments were performed. As shown in Fig. 4, increasing extravesicular Cl concentration saturated the [HCO3] gradient-driven 36Cl uptake in AMV from both normal and Na-depleted animals. Lineweaver-Burk plot analyses of these studies yielded a Km of ~8.9 ± 1.1 mM and a Vmax of 2.4 ± 0.6 nmol · mg protein-1 · 12 s-1 for Cl of [HCO3] gradient-driven 36Cl uptake for normal animals and a Km of 24.3 ± 1.8 mM and a Vmax of 0.8 ± 0.2 nmol · mg protein-1 · 12 s-1 for Na-depleted animals. These results demonstrate that the Km for Cl for Cl/HCO3 exchange is increased almost threefold in Na-depleted animals compared with normal animals. Similarly, DIDS inhibition kinetics were also determined for Cl/HCO3 exchange in normal and Na-depleted animals. Figure 5 reveals that Na depletion increased Ki for DIDS of ClHCO3 by 15-fold (Ki for DIDS of normal vs. Na depletion: 5.9 ± 2.9 vs. 91.1 ± 3.6 µM). Both the Km for Cl (24.3 ± 1.8 mM) and the Ki for DIDS (91.1 ± 3.6 µM) for [HCO3] gradient-driven 36Cl uptake in AMV of Na-depleted animals are almost identical to those [Km of 22.6 mM for Cl and Ki of 106.0 µM for DIDS (15)] previously established for Cl/OH exchange in normal animals (15).


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Fig. 4.   Effect of Cl concentrations ([Cl]) on HCO3 gradient-driven 36Cl uptake. AMV from normal (open circle ) and Na-depleted (bullet ) distal colon were preloaded with 150 mM KHCO3, 5 mM NMG gluconate, and 50 mM HEPES-Tris (pH 7.5). Uptake was measured for 12 s by incubating AMV in incubation medium containing 50 mM HEPES-Tris (pH 7.5), 36Cl-NMG, and varying concentrations of KCl. Isosmolarity was maintained by varying potassium gluconate concentrations. Uptake was also measured in presence of 1 mM DIDS. Absolute values presented were calculated by subtracting uptake obtained in presence of DIDS from that in its absence. Vesicles were gassed for 30 min, and uptakes were performed under 17% CO2 atmosphere. All media contained 25 µM valinomycin and 0.8% ethanol.


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Fig. 5.   Effect of DIDS on HCO3 gradient-driven 36Cl uptake. AMV from normal (open circle ) and Na-depleted (bullet ) distal colon were preloaded with 150 mM KHCO3, 5 mM NMG gluconate, and 50 mM HEPES-Tris (pH 7.5). Uptake was measured for 12 s by incubating AMV in medium containing 150 mM potassium gluconate, 5 mM 36Cl-NMG, 50 mM HEPES-Tris (pH 7.5), and varying concentrations of DIDS. AMV were gassed for 30 min, and uptake was performed under 17% CO2. Absolute values were calculated by subtracting uptake obtained in the absence of HCO3 gradients. All media contained 25 µM valinomycin and 0.8% ethanol.

These results suggest that HCO3 stimulates 36Cl uptake in AMV from Na-depleted animals via Cl/OH exchange, which has a relatively low affinity for HCO3 and is consistent with the concept that these two Cl/anion exchanges are not only distinct and separate transport processes but are also differentially regulated by Na depletion. The demonstration that Na depletion inhibits Cl/HCO3 exchange (Fig. 1A) and Na-dependent Cl absorption (10, 12), but not Cl/OH exchange (Fig. 1B), supports the prior speculation that Cl/HCO3 exchange is responsible for transepithelial Cl movement, while Cl/OH exchange is linked to one or more Cl-dependent epithelial cell functions (i.e., intracellular pH regulation and cell volume regulation) (15).

Because absorptive processes are primarily present in surface cells, the spatial distribution of Cl/HCO3 and Cl/OH exchanges in surface and crypt cells was also examined. Figure 6A presents the results of outward [HCO3] gradient 36Cl uptake in AMV from surface and crypt cells. Compared with Cl/HCO3 exchange in AMV from surface cells, there was a significant reduction in outward [HCO3] gradient-driven 36Cl uptake in AMV prepared from crypt cells. DIDS-sensitive outward [HCO3] gradient-driven 36Cl uptake in crypt AMV was 2.4% of that in surface AMV. In contrast, an outward [OH] gradient-driven 36Cl uptake in AMV prepared from crypt cells was 50% of that in AMV prepared from surface cells (Fig. 6B). Thus the distribution of these two apical membrane anion exchanges in surface and crypt cells differ: Cl/HCO3 exchange is primarily present only in surface cells, while Cl/OH exchange is localized to apical membranes of both surface and crypt cells. These observations provide additional support for the previous speculation (15) that Cl/HCO3 and Cl/OH exchanges are separate and distinct transport processes.


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Fig. 6.   Distribution of Cl/HCO3 (A) and Cl/OH (B) exchanges along surface-to-crypt cell axis. AMV prepared from surface and crypt cells of normal rat distal colon were preloaded with 50 mM HEPES-Tris (pH 7.5), 5 mM NMG gluconate, and 150 mM of either KHCO3 (A) or potassium gluconate (B). Uptake was measured for 12 s by incubating AMV in medium containing 150 mM potassium gluconate, 5 mM 36Cl-NMG, 25 µM valinomycin, and 50 mM of either HEPES-Tris (pH 7.5) (A) or MES-Tris (pH 5.5) (B). AMV with HCO3 were gassed for 30 min, and uptake was measured under 17% CO2 atmosphere. Uptake was also measured in presence of 1 mM DIDS. Absolute values presented represent DIDS-sensitive uptake that was derived by subtracting uptake in presence of DIDS from that in the absence of DIDS.


    DISCUSSION
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Abstract
Introduction
Materials and methods
Results
Discussion
References

The mechanism(s) for the absorption of Na and Cl in the mammalian colon has been well studied in several species, including human tissue, during the past two or more decades (4, 7, 8, 11, 12). The physiology of Cl absorption has been extensively investigated in the rat distal colon by several different complementary methods that have included in vivo luminal perfusion, in vitro ion fluxes across isolated mucosa under voltage-clamp conditions, and uptake measurements by colonocyte AMV (4, 11). Studies (4, 7, 8, 12) performed across isolated colonic mucosa under voltage-clamp conditions have established the presence of active Cl absorption that is electroneutral, both Na and HCO3 dependent, and is inhibited by both aldosterone and increases in mucosal cAMP content.

Prior studies of Cl/anion exchange in AMV from normal animals indicated the presence of distinct and separate Cl/HCO3 and Cl/OH exchanges (15). The present results provide additional and compelling evidence that two distinct apical membrane Cl/anion exchanges are present in the rat distal colon and are differentially regulated by Na depletion. First, Cl/HCO3 exchange is present in AMV from surface cells but not in AMV from crypt cells (Fig. 6). Second, Cl/HCO3 exchange was substantially inhibited by Na depletion (Fig. 1A), an observation that parallels our prior demonstration that Na depletion inhibits Na/H exchange (17). Although Cl/OH exchange is also partially inhibited by Na depletion (Fig. 1B), data presented in Figs. 2 and 3 provide evidence that the fraction of Cl/OH exchange that appears inhibited by Na depletion represents [OH] gradient-driven 36Cl uptake via Cl/HCO3 exchange (Fig. 1B). As a result, the inhibition of active electroneutral Na-Cl absorption by Na depletion in the rat distal colon is secondary to the parallel inhibition of both Na/H and Cl/HCO3 exchanges.

The previous studies (15) also concluded that both OH and HCO3 ions were transported by Cl/HCO3 exchange, whereas Cl/OH exchange only transported the OH ion. However, those experiments did not directly address whether HCO3 was also transported by Cl/OH exchange. These present studies also established that the affinity of Cl/HCO3 exchange for HCO3 was less than that of Cl/OH for OH. The data presented in Figs. 2 and 3 with AMV from Na-depleted animals revealed that at very low [OH] and [HCO3] 36Cl uptake was absent (within the experimental limitations of these studies), but that at higher [OH] and [HCO3] a low rate of 36Cl uptake was observed. These results are consistent with the transportation of both OH and HCO3 ions by Cl/HCO3 exchange and with inactivation of Cl/HCO3 exchange by Na depletion (see Figs. 1A and 6). At higher [OH] and [HCO3] 36Cl uptake is mediated by Cl/OH exchange, which has a lower affinity for these anions than Cl/HCO3 exchange. Thus 36Cl uptake represents both Cl/OH and Cl/HCO3 exchanges in normal animals, while in Na-depleted rats 36Cl uptake is a result only of Cl/OH exchange that has an affinity for both OH and HCO3 ions.

Several observations provide the basis for the speculation that Cl/HCO3 exchange is the mechanism of vectorial, transepithelial Cl absorption. First, absorptive processes are primarily, though not exclusively, located in colonic surface cells, while secretory processes are in crypt cells. Figure 6 demonstrates that Cl/HCO3 exchange is primarily present in AMV prepared from surface cells and not in AMV from crypt cells. In contrast, Cl/OH exchange is present in AMV from both surface and crypt cells (Fig. 6). Second, in addition to Na depletion's induction of Na channels in apical membrane of the distal colon, Na depletion inhibits both electroneutral Na-Cl absorption and [H] gradient-driven 22Na uptake (i.e., Na/H exchange) (12, 17). Figure 1A demonstrates that Na depletion substantially inhibits Cl/HCO3 exchange. Thus these observations indicate that Cl/HCO3 exchange is present in surface cells and is most likely closely associated with transepithelial Cl movement. Conversely, Cl/OH exchange is present in both surface and crypt cells (Fig. 6) and we propose that Cl/OH exchange is not regulated by Na depletion and not associated with transepithelial Cl movement but rather with the regulation of cell volume and/or intracellular pH.

To date, three different anion exchange genes (AE1, AE2, and AE3) have been cloned from different species (1). However, understanding of the membrane-specific localization of AE isoforms is presently incomplete in polarized epithelia. The expression of AE1 isoform protein has been localized to basolateral membranes of acid-secreting intercalated cells of cortical and medullary collecting ducts (1). Although studies in native tissue clearly indicate the presence of anion exchange function in apical membranes (3, 4), expression of AE isoform proteins has not been reported in either small or large intestine. To date, AE2 isoform-specific protein has been expressed in basolateral membranes of rat stomach and intestine and human intestine (2, 6, 20).

In conclusion, the present study establishes the distribution and Na depletion regulation of Cl/anion exchanges in the AMV of rat distal colon. The pattern of surface cells to crypt cells spatial distribution and Na depletion regulation indicate Cl/HCO3 exchange is responsible for transepithelial Cl movement in apical membranes of rat distal colon. In contrast, Cl/OH exchange that is present in AMV of both surface and crypt cells and not regulated by Na depletion may be responsible for cell volume and/or cell pH homeostasis in rat distal colon.


    ACKNOWLEDGEMENTS

This study was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grant DK-14669-24.


    FOOTNOTES

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

1 In previous studies, dietary Na depletion and aldosterone infusion via minipumps produced identical changes in Na, Cl, and K transport in rat distal colon (22-24). In addition, serum aldosterone levels were similar in both the dietary Na-depleted animals and those infused with aldosterone via minipumps (12). Thus, in the present study, aldosterone is at times used to refer to Na-depleted animals.

2 The presence of a sigmoidal relationship between [OH] and 36Cl uptake in normal animals in Fig. 2B relies heavily on the experimental data points at low [OH]. As a result, the following alternate interpretation of this data is also possible: 1) that hyperbolic relationships are present in both groups, and 2) that Na depletion does not alter Km for OH but reduces Vmax by ~50%, which is a reflection of inhibition of Cl/HCO3 exchange.

Address for reprint requests: V. M. Rajendran, Dept. of Internal Medicine, Yale Univ., New Haven, CT 06520.

Received 25 June 1998; accepted in final form 9 October 1998.


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Top
Abstract
Introduction
Materials and methods
Results
Discussion
References

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Am J Physiol Gastroint Liver Physiol 276(1):G132-G137
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