Role of luminal anion and pH in distal tubule potassium secretion

J. B. O. Amorim1, M. A. Bailey2, R. Musa-Aziz3, G. Giebisch2, and G. Malnic3

1 Basic Science Department, Faculdade de Odontologia de São José dos Campos, and 3 Department of Physiology and Biophysics, Instituto de Ciências Biomédicas, Universidade de São Paulo, 05508-900 São Paulo, Brazil; and 2 Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06510


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Potassium secretory flux (JK) by the distal nephron is regulated by systemic and luminal factors. In the present investigation, JK was measured with a double-barreled K+ electrode during paired microperfusion of superficial segments of the rat distal nephron. We used control solutions (100 mM NaCl, pH 7.0) and experimental solutions in which Cl- had been replaced with a less permeant anion and/or pH had been increased to 8.0. JK increased when Cl- was replaced by either acetate (~37%), sulfate (~32%), or bicarbonate (~62%), and also when the pH of the control perfusate was increased (~26%). The majority (80%) of acetate-stimulated JK was Ba2+ sensitive, but furosemide (1 mM) further reduced secretion (~10% of total), suggesting that K+-Cl- cotransport was operative. Progressive reduction in luminal Cl- concentration from 100 to 20 to 2 mM caused increments in JK that were abolished by inhibitors of K+-Cl- cortransport, i.e., furosemide and [(dihydroindenyl)oxy]alkanoic acid. Increasing the pH of the luminal perfusion fluid also increased JK even in the presence of Ba2+, suggesting that this effect cannot be accounted for only by K+ channel modulation of K+ secretion in the distal nephron of the rat. Collectively, these data suggest a role for K+-Cl- cotransport in distal nephron K+ secretion.

distal tubule; collecting duct; potassium secretion; postassium-chloride cotransport


    INTRODUCTION
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

A LARGE NUMBER OF FACTORS modulate potassium (K+) secretion by the initial and cortical collecting tubules (10). Luminal modulators include changes in the delivery of Na+ (12, 26), the transepithelial potential difference (9), the concentration of Ca2+ (21), the presence of impermeant anions, and the concentration of Cl- relative to that of other anions (6).

K+ secretion rises sharply when impermeant anions are present in the lumen of the distal tubule (27). Earlier studies attributed this effect to the increase in the lumen-negative transepithelial potential difference generated by persistent Na+ reabsorption in the face of the decrease in anion conductance resulting from the replacement of permeable Cl- by a much less permeant species, such as sulfate (5, 11). In vivo voltage-clamp experiments confirm the transepithelial potential difference as an important modulator of K+ secretion in the distal tubule (9). Nevertheless, the stimulatory effect of sulfate on K+ secretion persists even when the lumen-negative potential difference is collapsed by amiloride (27) and is not affected by the inclusion of Ba2+ in the perfusate (6). Therefore, it is not possible to attribute the effect of anions on K+ secretion solely to changes in transepithelial potential difference, and it has been proposed that substitution of Cl- by sulfate may enhance electroneutral K+ secretion by creating a favorable transmembrane gradient for K+-Cl- cotransport from cell to lumen (6).

Similar conclusions can be reached from studies in humans. Infusions of Na sulfate have a greater kaliuretic effect than do those of NaCl (4), although this is only apparent when the urinary Cl- concentration is <15 mM. That the ability of the impermeant sulfate to modulate K+ excretion is critically dependent on urinary Cl- concentration provides strong, if circumstantial, evidence for K+-Cl- cotransport as a pathway for K+ secretion in the distal nephron. These studies further reveal that infusions of Na bicarbonate stimulate K+ excretion to a greater extent than do those of Na sulfate (4). In contrast to the actions of sulfate, the kaliuretic effect of bicarbonate was not dependent on a concomitantly low urinary Cl- concentration. These observations would suggest that bicarbonate ions wield an additional influence on K+ secretion and thus excretion.

We have used stationary microperfusion of superficial segments of the rat distal tubule to investigate the role of luminal anions and pH, either separately or in combination, in modulating K+ secretion. We observed that K+ secretion was greater under conditions in which Cl- was replaced by either sulfate or acetate. This increment in K+ secretion could be abolished by known inhibitors of K+-Cl- cotransport, suggesting that the effect of both sulfate and acetate was indirect, attributable to low luminal Cl- concentration. Replacement of Cl- with bicarbonate induced a further increment in K+ excretion. Our experiments suggest that modulation of K+-Cl- cotransport by luminal fluid pH plays a key role in this action of bicarbonate on K+ secretion.


    METHODS
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Male Wistar rats (wt range 180-320 g), kept on a standard rat chow (plasma K+ concentration = 4.0 ± 0.7 mM), were anesthetized with Inactin (100 mg/kg ip) and prepared surgically for micropuncture experiments. During the experiments, body temperature was maintained at 37°C, and isotonic saline, containing 3% mannitol, was infused intravenously at a rate of 4 ml/h.

Stationary microperfusion. The technique employed in these experiments (1) is illustrated in Fig. 1. A proximal tubule was impaled by a double-barreled micropipette, and FD&C green control solution was injected from barrel P to localize distal tubule segments with one or more surface loops (Fig. 1A). Once the K+-selective microelectrode (see below) had been inserted into the last surface loop of the distal segment (Fig. 1B), a large column of Sudan-black-colored castor oil was injected into the proximal tubule through barrel C to prevent downstream flow of native tubular fluid. The oil column was subsequently split by injection of the control solution at a rate sufficient to lower distal tubular K+ concentration, as recorded by the electrode, to the level of the perfusion fluid. After control measurements, the experimental solution was injected through a single-barreled micropipette (P) inserted into either a late proximal loop or the early distal tubule (Fig. 1C), and recordings were made again. In a typical experiment, each nephron was perfused two to three times with both an experimental and control solution, allowing paired measurement of K+ secretion. In each animal, one to three nephrons were perfused in the manner described.


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Fig. 1.   Schematic drawing of the stationary microperfusion method. A: a superficial segment of a proximal convoluted tubule is punctured with a double-barreled micropipette containing Sudan-black colored castor oil (C) and control perfusate (P). B: a late superficial segment of the distal tubule is punctured with a double-barreled microelectrode, consisting of a potassium-selective barrel (IE) and a reference barrel (Ref) connected to a high-impedance electrometer (E). C: a single-barreled micropipette containing the experimental solution (S) is inserted into a late segment of the proximal tubule or, as shown, the early distal tubule.

Solutions and chemicals. The composition of the solutions used in the present study are presented in Table 1. It should be noted that the control solution (100 mM NaCl) was bicarbonate free and used at either pH 7.0 or 8.0. In all of the other solutions, NaCl, reduced to 20 or 2 mM, was replaced by sodium salts of acetate, sulfate, or bicarbonate. Acetate was used because it exerted a similar effect to that of sulfate and has a soluble barium salt. Raffinose pentahydrate (Riedel-de Haën, Hannover, Germany) was added to each solution to establish isosmolality with plasma. Under these equilibrium conditions, some Na+ reabsorption may persist but fluid reabsorption is minimized so that increases in luminal K+ concentration relate directly to K+ secretion. Furosemide and [(dihydroindenyl)oxy]alkanoic acid (DIOA), both established inhibitors of K+-Cl- cotransport (7, 29), were dissolved in dimethylsulfoxide before dilution; vehicle alone (0.5%) was included in the control solutions. Hexamethyldisilazane and K ionophore cocktail A were both obtained from Fluka (Buchs, Switzerland). All other reagents were from Sigma.

                              
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Table 1.   Perfusion solutions

Ion-selective microelectrodes. Double-barreled microelectrodes were pulled from assymetric borosilicate glass to a tip diameter of ~1 µm. The smaller, reference barrel was backfilled with a solution containing 0.24 M NaCl and 0.76 M Na acetate, colored by FD&C green. The total cation/anion mobility of this solution was similar because the mobility in a solution of Cl- is larger than that of Na+, whereas that of acetate is smaller. The animal was grounded (tail), and measurements of transepithelial potential difference were made through the reference barrel.

The larger barrel was exposed for 30 min at room temperature to hexamethyldisilazane before the tip was filled with K+ ionophore. Silanized electrodes were kept in a desiccator until the day of the experiment, when they were filled with the electrolyte solutions (1). The electrode was calibrated before and after each impalement by superfusion onto the kidney surface of standards (kept at 37°C). Standards of 3, 10, and 30 mM KCl were used, and 100 mM NaCl was added to each standard to compensate for the fact that the K+ ionophore has an inherent sensitivity to sodium. The mean voltage difference per decade change in K+ concentration was similar to values previously reported by us (1). The voltage difference between the reference and ion-selective barrels, representing luminal K+ activity, was recorded using a differential, high-impedance electrometer (model 223, WPI, New Haven, CT) and sampled every second by an analog-digital converter (Lynx, São Paulo, Brazil).

Analysis and statistics. For each perfusion, luminal K+ activity increased from the initial value of 0.5 mM, given as [K+]o, to a stationary K+ concentration ([K+]s) as a result of K+ secretion into the nephron. These data were analyzed by a Visual Basic program that fit an exponential to the approach of luminal K+ activities to the stationary level, i.e., by plotting the increase in luminal K+ activity against time. The half-time (t1/2) of the approach of K+ activity to the stationary level was calculated from this exponential, and secretory K+ fluxes (JK) were obtained by the following relationship, where r is tubule radius and other terms are as defined above
J<SUB>K</SUB> (nmol·cm<SUP>−2</SUP>·s<SUP>−1</SUP>) = <FR><NU>ln 2</NU><DE><IT>t</IT><SUB><SUB>½</SUB></SUB></DE></FR>·([K<SUP>+</SUP>]<SUB>s</SUB> − [K<SUP>+</SUP>]<SUB>o</SUB>)·<FR><NU><IT>r</IT></NU><DE>2</DE></FR>
A value for each variable was obtained from individual perfusions: the mean of repeated perfusions under each condition was calculated to give a pair of values for each tubule, and n corresponds to the number of tubules perfused. Statistical comparisons were made using Student's paired t-test or, when nonpaired groups were compared, analysis of variance and the Bonferroni contrast test. The probability of 0.05 was taken as the limit of statistical significance.


    RESULTS
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ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

A representative graph plotting the approach of the luminal K+ concentration to the stationary value (i.e., Ks - luminal K+ concentration) against time is shown in Fig. 2. Figure 2A details the secretory response under control conditions (100 mM NaCl). Similar data obtained with a solution in which Cl- was largely replaced by sulfate is depicted in Fig. 2B. Data were obtained from the same superficial distal tubule, and paired data sets were acquired for each set of experimental conditions. These curves were used to calculate t1/2, an index of the rate of K+ secretion. According to the equation cited in METHODS, JK depends on [K+]s and on the rate of the approach to [K+]s from the initial perfusion value of 0.5 mM (i.e., [K+]o), given as ln2/t1/2. From an inspection of Table 2, it is apparent that paired values for [K+]s were similar for Cl- replacement with acetate, sulfate, or bicarbonate. Modulation of K+ secretion by acute changes in luminal components thus reflects increases or decreases in t1/2. It is apparent that the rate at which the steady-state luminal K+ concentration is established is greater, and t1/2 is smaller, when Cl- is replaced by a less permeant anionic species.


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Fig. 2.   Approach of luminal K+ concentration ([K+]) to steady-state level ([K+]s) from the initial perfusate concentration of 0.5 mM ([K+]o). The change in [K+], which equals [K+]s - [K+] at time t, is plotted as a function of time (s). triangle , Experimental points; solid lines, exponentials fitted to data. A: control perfusion (100 mM NaCl). B: perfusion of the same nephron segment using a solution in which 80 mM Cl- has been replaced by sulfate.


                              
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Table 2.   Stationary potassium concentration and approach to half-time during paired perfusion of the late distal tubule

Calculation of JK from [K+]s - [K+]o and t1/2 yields the data presented in Figs. 3-6. A significant increase in JK was observed when either sulfate or acetate replaced luminal Cl-. When the principal anion was bicarbonate, a further increment was recorded (Fig. 3). As shown in Table 3, perfusion with sulfate-containing solutions resulted in only a minor increase in the lumen-negative transepithelial potential difference, whereas neither bicarbonate nor acetate altered this difference. These data indicate that the role of changes in transepithelial potential difference in mediating K+ secretion is minor.


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Fig. 3.   Net K+ secretory flux (JK) calculated from [K+]s and t1/2, during paired perfusion of the distal tubule with control solutions (Cl-) and solutions in which Cl- has been replaced by acetate, sulfate, or bicarbonate, as detailed in Table 1. Values are means ± SE; n = 9, 14, and 18 tubules, respectively. *P < 0.05, **P < 0.01, paired t-test. The effect of bicarbonate was significantly greater than that of either acetate or sulfate (P < 0.05, unpaired t-test).



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Fig. 4.   Net JK during paired perfusion of Cl- and acetate and of acetate plus barium (Ba) and acetate plus barium plus furosemide (furo). Ba (3 mM) reduced acetate-stimulated JK by ~80% (P < 0.01). On addition of furosemide (1 mM) to this perfusate, the flux decreased further to ~11% of control values (P < 0.01). Values are means ± SE; n = 14 tubules for each paired condition. *P < 0.05, **P < 0.01, compared with Cl-.



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Fig. 5.   Top: JK increases as luminal Cl- concentration decreases from 100 (P < 0.05) to 2 mM (P < 0.01), ANOVA. Bottom: stimulatory effect of reducing luminal Cl- concentration is abolished by either furosemide (1 mM; P < 0.01) or [(dihydroindenyl)oxy]alkanoic acid (DIOA; 100 µM; P < 0.01), inhibitors of K-Cl cotransport. DIOA (100 µM) has no significant effect during perfusions with 100 mM Cl-.



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Fig. 6.   K+ secretion in the distal tubule during luminal perfusion with the control perfusate (100 mM NaCl) at pH 7.0 and 8.0. Values are means ± SE; n = 11 (pH) and 18 (bicarbonate) tubules. Increasing the pH of the luminal fluid significantly increased JK (*P < 0.05 vs. pH 7.0), although this effect was smaller than that of bicarbonate at the same pH (**P < 0.05 vs. bicarbonate).


                              
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Table 3.   Transepithelial potential difference in the late distal tubule during perfusion with solutions of differing principal anions

To assess the role of channels in mediating K+ secretion, Ba2+ was added to an acetate-containing perfusate. Ba2+ markedly reduced JK (Fig. 4), leaving a residual K+ secretion of 0.18 nmol · cm-2 · s-1. The remaining JK was of a similar magnitude to the increment in K+ secretion resulting from replacement of Cl- with acetate (0.23 nmol · cm-2 · s-1), suggesting that the effect of acetate was not mediated through ion channels. Addition of furosemide (1 mM) to the solution further reduced K+ secretion (Fig. 4). The remainder was still significantly greater than zero (P < 0.05) and probably represents passive paracellular flux of K+ driven along the favorable electrochemical gradient (-42.0 ± 1.7 mV).

Taken together, these observations strongly suggest that electroneutral K+-Cl- cotransport is operative after the establishment of a favorable cell-to-lumen Cl- concentration gradient. This hypothesis is supported by the observation that increasing the driving force for K+-Cl- cotransport by reducing luminal Cl- concentration from 20 to 2 mM evoked a further increase in K+ secretion (Fig. 5, top). With conditions employed herein, a reduction in luminal Cl- is associated with an increase in luminal sulfate, and we cannot preclude a direct effect of the replacement anion. Nevertheless, it is noteworthy that either furosemide or DIOA (a more selective inhibitor of K+-Cl- cotransport) was able to reduce anion/low-Cl--stimulated K+ secretion to levels observed with 100 mM Cl- (Fig. 5, bottom). In addition, in the presence of 100 mM Cl- in the lumen, 100 µM DIOA did not affect K+ secretion significantly. These experiments thus imply that the magnitude of the cell-to-luminal Cl- concentration gradient rather than the nature of the luminal anion accounts for the augmentation of K+ secretion.

In a series of further experiments, we investigated the effect of changes in luminal pH on K+ secretion because the effects of bicarbonate could be attributed to alkalinization of the tubular fluid. Increasing pH of the control (100 mM NaCl) solution from 7.0 to 8.0 resulted in a significant stimulation of K+ secretion (Fig. 6). However, this increment was significantly less than that resulting from perfusion of the lumen with a bicarbonate solution, also at pH 8.0. It is notable that the increment in JK resulting from alkalinzation of the tubular fluid was similar in magnitude to the difference between the respective stimulations resulting from bicarbonate and from sulfate. To further investigate the mechanism by which alkalinization of the luminal perfusate enhanced K+ secretion, we performed the experiments depicted in Fig. 7. Increasing the pH of the acetate-containing perfusate from 7.0 to 8.2 resulted in a marked increase in K+ secretion, despite the presence of Ba2+ in both solutions.


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Fig. 7.   Increasing the pH of the acetate perfusate from 7.0 to 8.2 stimulates JK despite the continued presence of Ba (3 mM). Values are means ± SE; n = 16 tubules; **=P < 0.01.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

K+ secretion in the distal nephron proceeds by two distinct processes: active uptake of K+ in exchange for Na+ by the Na+-K+-ATPase across the basolateral membrane provides the driving force for translocation across the apical membrane along a favorable electrochemical gradient. The present investigation concerns the mechanisms by which changes in the composition of the tubular fluid, independently of maneuvers that alter the peritubular fluid, affect K+ secretion. We have specifically focused on the mechanisms by which changes in the concentration of distal tubular bicarbonate and other anions modulate K+ transport.

Previous studies have firmly established several luminal factors known to alter K+ secretion in the distal convoluted tubule and initial and final collecting ducts. These factors include the delivery of Na+ (12), fluid (13), and Ca2+ (21), the pH of fluid (3), and the nature of the principal anionic species (2, 26, 27). Studies in humans have defined the important role of bicarbonate as a powerful stimulus of K+ secretion (4, 15, 16) and have shown that its kaliuretic potency exceeds that of other poorly reabsorbable anions such as sulfate.

In the present study, K+ secretion in the initial collecting duct was measured under equilibrium conditions in which Na+ reabsorption was not associated with significant fluid movement. The main findings were 1) replacement of Cl- with either sulfate or acetate increased the rate of K+ secretion; 2) replacement of Cl- with bicarbonate elicited a further increment in K+ secretion; and 3) increasing the pH of the control perfusate stimulated K+ secretion.

The effect of sulfate and acetate. It has been suggested that impermeant anions increase the luminal negativity of the transepithelial potential difference, thereby imposing a more favorable electrochemical gradient for K+ secretion (5, 11). That the transepithelial potential difference regulates K+ secretion is beyond doubt (9). The importance of this variable in the present study, however, must be questioned because sulfate induced only a small increase in the lumen-negative transepithelial potential difference and both acetate and bicarbonate were without effect. Our data imply that a change in potential difference would contribute in only a minor way to the anion-stimulated K+ secretion and are in accord with earlier studies which found that sulfate can stimulate K+ secretion even when the negative transepithelial potential difference is abolished (27).

K+ channels and K+-Cl- cotransport. It is reasonable to conclude that a large fraction of K+ secretion in our experiments was mediated by diffusion across the apical membrane through K+ channels because addition to the tubular fluid of Ba2+, an effective blocker of both the small-conductance and the maxi-K channel (10), reduced JK by ~80% (Figs. 4 and 7). However, the persistence of a residual JK suggests additional secretory pathways. It is possible that K+-Cl- cotransport participated in distal nephron K+ secretion as first proposed on the basis of experiments demonstrating that K+ secretion was stimulated by low concentrations of luminal Cl- (27), even in the presence of Ba2+ (6). Ba2+-insensitive K+ secretion has also been reported in the isolated cortical collecting duct (20, 24) but only in animals treated with deoxycorticosterone acetate.

The present study confirmed the presence of a Ba2+-insensitive component of K+ secretion in the distal nephron. This component was almost completely inhibited by both furosemide and DIOA (Figs. 4 and 5). Similarly, maneuvers designed to increase the cell-to-lumen Cl- concentration gradient evoked a gradient-dependent increase in JK, an increase that was eradicated by K+-Cl- cotransport inhibition (Fig. 5). Although the specificity of K+-Cl- cotransport inhibitors is limited, the most likely interpretation of these findings is that K+-Cl- cotransport contributes to the stimulation of JK by acetate, sulfate, and bicarbonate. Ba2+-insensitive K+ secretion in the late distal tubule of control rats had previously been demonstrated (1) and supports the possibility that K+-Cl- cotransport is operative under normal conditions. In support of this hypothesis, addition of either furosemide or DIOA to low- Cl- perfusates reduced K+ secretion to a level that was slightly below that found with 100 mM Cl-. However, we observed that addition of DIOA to 100 mM Cl--containing perfusates did not cause significant alteration of K+ transport, suggesting that K+-Cl- cotransport is only operative when the cell-to-lumen gradient is favorable (Fig. 5). The observation of a component of Ba2+-insensitive K+ secretion at higher luminal or urinary Cl- concentrations (4, 15, 20, 24) suggests the possibility of K+ secretion via a paracellular transport route.

Effect of bicarbonate and pH. K+-Cl- cotransport would be expected to contribute to the stimulation of JK resulting from the replacement of Cl- with bicarbonate (Fig. 3) for the reasons cited above. Nevertheless, bicarbonate has additional effects associated with the alkaline pH of such solutions. There is a strong correlation between the pH of the tubular fluid and K+ secretion in the distal nephron (17, 18), and reducing the perfusate pH from 7.4 to 6.8 impairs K+ secretion in the rabbit cortical collecting duct (3). In the present study, we found that increasing the pH of the control solution (100 mM NaCl) stimulated distal K+ secretion. Although the effect of cytoplasmic pH on K+ channel activity has been repeatedly demonstrated (8, 18), several observations make it very unlikely that increasing luminal pH may enhance secretion through the small-conductance K+ channel of the principal cell (ROMK). First, ROMK channels are insensitive to changes in extracellular pH (8, 10, 33). Second, intracellular pH of the principal cell was shown to be unaffected by changes in luminal pH due to the apparent lack of an acid-base transporter in the apical membrane (30). Sensitivity of ROMK to pH seems thus to be limited to the cytosole (8, 18). Finally, addition of Ba2+ to the acetate perfusate did not ablate the stimulatory effect on JK of increasing pH from 7.0 to 8.2. This suggests that luminal pH augments K+ efflux through a Ba2+-insensitive pathway. Two possibilities should be considered. The first is that changes in luminal pH activate K+-Cl- cotransport. Because the cell-to-lumen Cl- concentration gradient remained constant during experiments, extracellular pH would have to exert a direct modulating action on K+-Cl- cotransport. An alternative hypothesis invokes a role for the TASK2 K+ channel, which is activated by extracellular alkalinization over the physiological range yet is relatively insensitive to Ba2+ block (22). In situ hybridization studies localize TASK2 expression to the cortical distal tubules and collecting ducts, but a clearly defined role for these channels is lacking.

Taken together, our data suggest an important role of K+-Cl- cotransport in mediating anion effects on K+ secretion. Because distal K+ secretion has been attributed mostly to the principal cell of cortical collecting duct, we have incorporated K+-Cl- cotransport into this cell. This suggestion is shown in our model of K+ secretion in the distal nephron (Fig. 8A). The molecular nature of this transport is unclear, although three of the four known isoforms of the K+-Cl- cotransporter are present in the kidney: KCC3 and KCC4 have been found in renal proximal tubules, particularly in the basolateral membrane (19). KCC1 is expressed in the apical membrane of the thick ascending limb; more recent evidence localizes this transporter also to distal tubule and collecting duct (Mount DB, personal communication). Nevertheless, alternative explanations for our findings should be considered (Fig. 8B). Stimulation of JK by both the high luminal pH and bicarbonate could also involve known transport mechanisms in B-intercalated cells. In these cells, coupling of the apically located Cl-/bicarbonate exchanger (14, 23, 25, 28, 32) with H+-K+-ATPase has been suggested to mediate Cl- reabsorption (32). Under special circumstances, such coordinated activity could also be involved in KCl secretion. We consider the inversion of active KCl reabsorption into secretion highly unlikely. However, alkalinization of B-intercalated cells by reversal of Cl-/ bicarbonate exchange could reduce H+-K+-ATPase activity and thus diminish K+ reabsorption. In parallel with continued K+ secretion via principal cells, enhanced net secretion of K+ would result. However, this hypothesis must be considered uncertain because the activity of H+-K+-ATPase appears to be significant only when these cells are acidified (31). This issue requires further investigation, especially in view of the fact that intercalated cells have been found to display significant heterogeneity with respect to expression of acid-base transporters (28, 31).


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Fig. 8.   Possible models of collecting duct K+ transport. Top: K+ secretion in the principal cell involves active uptake of K+ in exchange for Na+ by Na+-K+-ATPase across the basolateral membrane, which provides the driving force for translocation across the apical membrane along a favorable electrochemical gradient. Luminal K+ transport involves K+ channels and K-Cl cotransport. Luminal modulators of K+ secretion include the transepithelial potential difference, the concentration of luminal Cl- and bicarbonate, and pH. pH affects K+ channels from the cytoplasmic side, and K-Cl cotransport from the lumen. Bottom: alternative model involving B-type intercalated cell with apical H+-K+-ATPase and Cl-/HCO<UP><SUB>3</SUB><SUP>−</SUP></UP> exchange. Modified according to Zhou et al. (32). CCD, cortical collecting duct.

In conclusion, our studies show that elevation of bicarbonate in distal tubules is uniquely potent in stimulating K+ secretion. The data suggest an important role of K+-Cl- cotransport or coordinated changes in H+-K+-ATPase activity coupled to HCO<UP><SUB>3</SUB><SUP>−</SUP></UP>/Cl- exchange. The present findings also suggest that from a physiological point of view the proposed transport mechanism (K+-Cl- cotransport) would be operative in low-Cl- urines, found in patients with low-salt diets, and particularly in conditions of increased bicarbonate excretion such as metabolic alkalosis.


    ACKNOWLEDGEMENTS

M. A. Bailey was funded by the Wellcome Trust. G. Giebisch was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grant DK-17433. R. Musa-Aziz and G. Malnic were supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo and by Conselho Nacional de Desenvolvimento Cientifico e Tecnológico (Pronex).


    FOOTNOTES

Address for reprint requests and other correspondence: G. Malnic, Dept. Fisiologia e Biofisica, Inst. Ciencias Biomedicas, USP, Av. Prof. Lineu Prestes, 1524, 05508-900 São Paulo, Brazil (E-mail: gemalnic{at}usp.br).

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. Section 1734 solely to indicate this fact.

First published October 22, 2002;10.1152/ajprenal.00236.2002

Received 25 June 2002; accepted in final form 15 October 2002.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Amorim, JB, and Malnic G. V1 receptors in luminal action of vasopressin on distal K+ secretion. Am J Physiol Renal Physiol 278: F809-F816, 2000[Abstract/Free Full Text].

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4.   Carlisle, EJ, Donnelly SM, Ethier JH, Quaggin SE, Kaiser UB, Vasuvattakul S, Kamel KS, and Halperin ML. Modulation of the secretion of potassium by accompanying anions in humans. Kidney Int 39: 1206-1212, 1991[ISI][Medline].

5.   Clapp, JR, Rector FC, and Seldin DW. Effect of unreabsorbed anion on proximal and distal transtubular potentials in the rat. Am J Physiol 202: 781-786, 1962[Abstract/Free Full Text].

6.   Ellison, DH, Velazquez H, and Wright FS. Stimulation of distal potassium secretion by low lumen chloride in the presence of barium. Am J Physiol Renal Fluid Electrolyte Physiol 248: F638-F649, 1985[Abstract/Free Full Text].

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