Role of Aldosterone in the Regulation of Sodium and Chloride Transport in the Distal Colon of Sodium-Depleted Rats

Transcription

1 GASTROENTEROLOGY 196;91:17-33 Role of Aldosterone in the Regulation of Sodium and Chloride Transport in the Distal Colon of Sodium-Depleted Rats JONATHAN HALEVY, MARY E. BUDINGER, JOHN P. HAYSLETT, and HENRY J. BINDER Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut Dietary sodium depletion with elevated aldosterone levels induces electrogenic, amiloride-sensitive sodium absorption and inhibits electroneutral sodium chloride absorption in the rat distal colon. To assess the role of aldosterone in the production of these changes, unidirectional Na and 36Cl fluxes were performed under voltage clamp conditions across isolated distal colonic mucosa of rats given continuous aldosterone infusions for up to 1 days. Aldosterone infusion for 7-1 days produced identical changes in both electrogenic sodium absorption and electroneutral sodium chloride absorption compared with dietary sodium-depleted animals. In contrast, aldosterone at 4, 4, and 7 h produced varying changes in ion transport: electrogenic sodium absorption progressively increased, whereas electroneutral sodium chloride absorption was initially augmented but then inhibited. Aldosterone induced active potassium secretion, demonstrated by a reversed short-circuit current after addition of amiloride, in all experimental groups. These results demonstrate that the changes in ion transport observed in sodium-depleted animals are produced by aldosterone, and that aldosterone not only stimulates electrogenic sodium absorption and potassium secretion but in a time-dependent manner both stimulates and inhibits electroneutral sodium chloride absorption. Received February 11, 196. Accepted May 1, 196. Address requests for reprints to : Dr. Henry J. Binder, Department of Internal Medicine, 333 Cedar Street, New Haven, Connecticut Mary E. Budinger's present address is: SUNY-Stony Brook School of Medicine, Stony Brook, New York. Dr. Halevy's present address is: Beilinson Hospital, Tel Aviv, Israel. This study was supported in part by United States Public Health Service grants AM and AM1061 from the National Institute of Arthritis, Diabetes, Digestive and Kidney Diseases, and a grant from the John A. Hartford Foundation, Inc. 196 by the American Gastroenterological Association /6/$3.50 Sodium depletion induced by the administration of a sodium-deficient diet results in' several changes in colonic and renal fluid and electrolyte function (1-3). Studies performed in vivo have revealed that dietary sodium deprivation increases net sodium absorption and net potassium secretion in the rat distal large intestine (-5). In detailed studies performed in the colon both sodium-potassium-stimulated adenosine triphosphatase activity and the surface area of the basolateral membrane are greater in sodium-depleted rats compared with control animals (5,6). Examination of ion transport in vitro under voltage clamp conditions has established that sodium depletion augments electrogenic sodium absorption, induces amiloride sensitivity, and stimulates electrogenic potassium secretion while inhibiting electroneutral sodium chloride absorption (7,). As sodium depletion is associated with secondary hyperaldosteronism, it is likely that these changes in sodium and potassium transport result directly from the action of aldosterone on colonic epithelium. However, it is possible that the changes observed in sodium-depleted animals are in part due to factors altered by sodium depletion, such as volume depletion, angiotensin, or other as yet unidentified factors. To assess the importance of aldosterone in the production of the changes in electrolyte transport observed in sodium deprivation, colonic ion transport was determined in the distal colon of rats given continuous infusion of aldosterone administered by minipump for varying periods of time, up to 1 days, and was compared with those in which sodium depletion had been produced by the administration of a sodium-free diet. These experiments establish that ion transport is identical in the distal large intestine in sodium-depleted animals and in those Abbreviations used in this paper: G, conductance; I,c' short-circuit current; Jnel Cl, net chloride absorption; Jnel Na, net s odium absorption.

2 1 HALEVY ET AL. GASTROENTEROLOGY Vol. 91, No.5 Table 1. Unidirectional and Net Sodium and Chloride Fluxes in Control, Sodium-Depleted, and Long-Term Aldosterone Animals Q Sodium fluxes (J1.Eq/h. cm ) Chloride fluxes (f.1.eq/h. cm ) Isc G Jms Jsm Jnet Jms Jsnl Jnet (f.1.eq/h. cm ) (ms/cm) Control Before amiloride 9.5 ± ± ± ± ± ± ± ± 0.6 After amiloride.5 ± ± ± ± ± ± ± ± 0.7 Probability value b <0.01 NS NS NS NS NS NS NS Sodium depletion Before amiloride 10.9 ± ± ± ± ± ± ± ± 0.9 After amiloride 5.4 ± ± ± ± ± ± ± ± 0. Probability value b <0.005 NS <0.005 NS NS <0.05 <0.005 <0.005 Aldosterone long-term Before amiloride 11. ± ± ±: ±: ±: ±: ±: ±: 0.7 After amiloride 6.1 ±: ±: ±: ±: ±: ±: ±: ±: 0.9 Probability value b <0.001 NS <0.001 NS NS NS <0.001 <0.005 G, conductance; I sc ' short-circuit current; J, ion fluxes; NS, not significant. a The control animals were fed a standard Purina Chow diet (Ralston-Purina, St. Louis, Mo.), whereas the sodium-depleted animals were maintained on a low-sodium diet to ihduce secondary hyperaldosteronism. The long-term aldosterone animals received continuous aldosterone infusions via Alzet osmotic minipumps implanted subcutaneously for 7 or 1 days. The results of these two groups were combined, as the rate of sodium and chloride transport in each group was identical. There were 13, 5, and 10 tissue pairs in the control, sodium-depleted, and long-term aldosterone groups, respectively. The period before amiloride represents the 15-min flux period before the addition of 0.1 mm amiloride to the mucosal bathing solution. Twelve minutes after the addition of amiloride, a second 15-min flux was performed. Results are expressed as mean ±: SEM. b Difference between before-amiloride and after-amiloride periods. given prolonged administration of aldosterone. The studies, however, also reveal that the time-course for the induction of these changes in colonic epithelial cell function by aldosterone is not identical and that both the duration and the magnitude of elevated aldosterone levels are determinants of the observed alterations of sodium, chloride, and potassium transport. Materials and Methods Nonfasting Sprague-Dawley rats weighing g were used in all experiments. After the rats were killed, mucosa stripped of the serosa and part of the muscle layers was mounted in a Lucite chamber as previously described (9). Potential difference, short-circuit current (Isel, and conductance (G) were measured, also as previously described (,9). Unidirectional Na and 36CI fluxes were performed by methods that have also been previously described in detail (). All experiments were performed in Ringer solution, ph 7.4, which contained (in millimoles per liter): Na, 140; K, 5.; Ca, 1.; Mg, 1.; CI, 119.; HC03, 5; HP0 4, 5.; glucose, 10. The solutions were oxygenated with 95% O-5% CO, A control and five experimental groups with elevated aldosterone levels were studied. Two different methods were used to raise plasma aldosterone levels: feeding of a low-sodium diet and continuous infusion of aldosterone administered via a subcutaneously implanted Alzet minipump. The low sodium paste diet contained 0.10 p,eq Na/l00 g food, was prepared as previously described (), and was provided for 7 days to produce secondary aldosteronism. Rats receiving this diet are referred to as the sodium-depleted group. Aldosterone was infused at a rate of 70 p,g/100 g body wt. day. There were four different aldosterone infusion groups: groups in which aldosterone was infused for 4, 4, and 7 h are referred to as the 4-h, 4-h, and 7-h aldosterone groups. The fourth group received an aldosterone infusion for either 7 or 1 days; as their results were identical, these two groups have been combined and are referred to as the long-term aldosterone group. Plasma aldosterone levels were determined by a commercial radioimmunoassay kit (Diagnostic Products Corporation, Los Angeles, Calif.), using 15I-Iabeled aldosterone and antibody-coated tube technology for the final separation of free from bound aldosterone (10). Levels were measured in control, dietary sodium-depleted, and 4-h, 7-h, and long-term aldosterone groups at the time of death (10). Plasma aldosterone level was 1. ±.7 ng/dl and ± 63.7 ng/dl in control and sodium-depleted animals respectively. Plasma aldosterone levels were comparable in the 4-h, 7-h, and long-term aldosterone groups (79.6 ± 77., 99.3 ± 100.6, and ± 5.4 ng/ml, respectively). The paired and unpaired Student's t-tests were performed as indicated (11). A probability value <0.05 was considered statistically significant. Results Table 1 presents the results of unidirectional and net sodium and chloride transport in control and sodium-depleted animals, as well as in animals that received aldosterone for 7-1 days (long-term aldosterone group). In the control group, net sodium

3 November 196 ALDOSTERONE AND COLONIC ION TRANSPORT 19 absorption Unet Na ) (5.1 ± 0.9 f.teq/h. cm ) was similar to net chloride absorption (Jnet C1 ) (5.5 ± 1.0 f.teq/h. cmz) and was significantly greater than the short-circuit current (Ise). These results are similar to previous studies of sodium and chloride absorption in the distal colon of normal rats that have been interpreted to indicate that the predominant mechanism of sodium and chloride absorption in this epithelium is electroneutral sodium chloride absorption (,9). The addition of 0.1 mm amiloride to the mucosal bathing solution did not alter Jnet Na, Jnet C1, Ise, or G. As in previous studies (), sodium depletion with secondary aldosteronism produced significant changes in ion transport in the distal colon. Although there was a small, but not significant, increase in Jnet Na (7. ± 0.4 vs. 5.1 ± 0.9 f.teqlh. cmz) in the experimental group, sodium depletion resulted in a 10-fold increase in Ise (4.9 ± 0.5 vs. 0.5 ± 0.1 f.teq/h. cm ; p < 0.001) and a significant decrease in Jnet CI (1.5 ± 0.7 vs. 5.5 ± 1.0 f.teq/h. cm ; p < 0.05). The addition of 0.1 mm amiloride to the mucosal bathing solution of the sodium-depleted animals resulted in almost complete inhibition of Jnet Na, primarily as a result of a decrease in Jrns Na. There was no change in Jsrn Na or in unidirectional or net chloride transport after amiloride. Amiloride also induced a reversal of Ise from 4.9 ± 0.5 to -0.7 ± 0. f.teq/h. cm z. These results are consistent with our previous conclusions () that sodium depletion induced amiloride-sensitive electrogenetic sodium absorption, inhibits electro neutral sodium chloride absorption, and stimulates active potassium secretion. Table 1 also presents the results of ion transport studies in the animals that received long-term aldosterone. Excess aldosterone resulted in a modest increase in Jnet Na. In these animals Ise was increased to 7.1 ± 0. from 0.5 ± 0.1 f.teq/h. cm, p < 0.001, and Jnet CI was reduced to 1.7 ± 0.7 from 5.5 ± 1.0 f.teq/h. cm, p < Similar to observations in sodium-depleted animals, the addition of 0.1 mm amiloride resulted in a marked reduction of Jnet Na to 1.0 ± 0.7 f.teq/h. cm z and reversed Ise to -1.6 ± 0.7 f.teq/h. cm z. Sodium and chloride transport was virtually identical in the two groups with elevated aldosterone levels, i.e., the sodium-depleted and long-term aldosterone groups. For example, Jnet Na was 7.0 ± 0.7 and 7. ± 0.4 f.teqlh. cm before amiloride and 1.0 ± 0.7 and 1. ± 0.6 f.teq/h. cm after amiloride in the sodium-depleted and longterm aldosterone groups respectively. Net chloride absorption and Ise were also similar in these two experimental groups. Conductance was significantly increased in the two groups with elevated aldosterone levels compared with the control group. Furthermore, although amiloride did not alter G in the control group, 0.1 mm amiloride added to the mucosal bathing solution resulted in a significant decrease in G in the two experimental groups. These results strongly suggest, therefore, that aldosterone is primarily responsible for the changes in colonic ion transport observed in sodium-depleted animals. Additional experiments were performed in animals that received infusions of aldosterone for 4, 4, and 7 h to determine the time required for the induction of the changes in ion transport produced by aldosterone. In the 4-h aldosterone group Jnet Na, Jnet C1, and Ise were all increased compared with the values observed in the control group (Figure la). The increases in both Jnet Na and Jnet CI (5. ± 1. 7 and N E u.i:... <T ::I.. * p<o.ooi compo red to control 0 t p<o.0 compared to 4h 6 4 Control 4h 4h 7 h Long -term 10 * p<o.005 N E u 6 t p<o.05 compared to 4h *, t..c... <T ::I.. 4 Control 4h 4h 7h Long-term Figure 1. Parameters of electrogenic sodium transport in control, 4-h, 4-h, 7-h, and long-term aldosterone groups. A. Short-circuit current increased as a time-dependent function of aldosterone infusion in that I,c was significantly greater both in the 4-h aldosterone group than in the control group and in the 4-h group than in the 4-h group. B. Amiloride-sensitive component of Jnet Na represents the decrease in Jnet Na after the administration of 0.1 mm amiloride. Amiloride did not produce a significant change in Jnet Na in the control group, but induced a significant decrease in Jnet Na in all of the aldosterone groups. The amiloride-sensitive component of Jnet Na in the long-term aldosterone group was greater than that observed in the 4-h group.

4 130 HALEVY ET AL. GASTROENTEROLOGY Vol. 91, No p<o.ooi compared with those in the 4-h aldosterone group (Table and Figures 1-3). Amiloride resulted in identical decreases in conductance in all three aldosterone groups '" E u ~... IY :t Control 4h 4h 7 h Long-term Figure. Short-circuit current after amiloride addition in contro\' 4-h, 4-h, 7-h, and long-term aldosterone groups. Short-circuit current was reversed after addition of 0.1 mm amiloride in all aldosterone groups, but not in the control group, and most likely represents active potassium secretion (7). The reversed Ise after amiloride was similar in all four aldosterone groups. * Discussion Dietary sodium deprivation has frequently been used to study the effect of aldosterone on 10 '" E u 6 p<0.05 t p<0.05 compared to 4 h 4.9 ±.0 /LEq/h cmz, respectively) were greater than the elevation of Ise (1.9 ± 0.6 /LEq/h. cmz). The addition of 0.1 mm amiloride resulted in a modest but significant decrease in Joel Na of. ± 0.6 /LEq/h. cmz, did not alter J net Cl, and decreased Ise by 3.9 ± 0.6 /LEq/h. cm z to -1.5 ± 0.1 /LEq/h. cm z (Figure ). The decrease in Joet Na produced by amiloride (i.e., amiloride-sensitive component) in the 4-h aldosterone group was significantly smaller than that seen in the long-term aldosterone group (. ± 0.6 vs. 5.9 ± 1.0 /LEq/h. cmz) (Figure lb). Net sodium absorption after the addition of amiloride (i.e., the amiloride-i:'''3nsitive component) was greater in the 4-h aldosterone group (.1 ± 1.4 /LEq/h. cmz) than in both the control and the longterm aldosterone groups (4.3 ± 0.9 and 1.0 ± 0.7 /LEq/h. cmz, respectively) (Figure 3B). In contrast, Ise after amiloride addition was identical in the two aldosterone groups (-1.5 ± 0.1 vs ± 0. /LEq/h cmz) (Figure ). In the 4-h aldosterone group Ise was increased to 5. ± 1.1 /LEq/h. cm, which was greater than that in both the 4-h aldosterone and the control groups (Table and Figure la). The short-circuit current was also reversed to -1.7 ± 0.1 /LEq/h. cm by mucosal amiloride in this group (Figure ). Net sodium absorption and Joet C1 values were both less than those observed in the 4-h aldosterone group, although J nel Na was inhibited by 0.1 mm amiloride by - 50%. Essentially identical results were observed in all parameters in the 7-h aldosterone group 10 '" E u 6.c... IY :t. 4 Long-term * p< 0.0 t p<0.05 compared to 4h Control 4h 4h 7h Long-term Figure 3. Parameters o f electroneutral sodium chloride absorption in control, 4-h, 4-h, 7-h, and long-term aldosterone groups. A. Net sodium absorption minus the electrogenic component Unc,Na - I,e) represents the electroneutral component of sodium chloride a bsorption. B. Amiloride-insensitive component of ] oc,na is the rate of sodium absorption observed after addition of 0.1 mm amiloride. These two parameters of electroneutral sodium absorption were significantly increased after aldosterone infusion for 4 h compared with the control group. Both parameters were significantly decreased in the 4-h aldosterone group compared with that observed in the 4-h group. In the long-term aldosterone group these parameters were significantly less than those observed in the control and the 4-h aldosterone groups and were not significantly different from zero. t

5 November 196 ALDOSTERONE AND COLONIC ION TRANSPORT 131 Table. Unidirectional and Net Sodium and Chloride Fluxes in Control, 4-h, 4B-h, and 7-h Aldosterone Groupso Sodium fluxes (JLEq/h. em ) Chloride fluxes (JLEq/h. em ) Ise G Jms Jsm Jne' Jms Jsm Jne' (JLEq/h. em ) (ms/em ) Control Before amiloride 9.5 ± ± ± ± ± ± ± ± 0.6 After amiloride.5 ± ± ± ± ± ± ± ± 0.7 Probability value b < 0.01 NS NS NS NS NS NS NS Aldosterone: 4 h Before amiloride 15. ± ± ± ± ± ± 1..4 ± ± 1. After amiloride 13.0 ± 1.3 4,9 ± ± ± ± ± ± ± 1.3 Probability value b < NS <0.005 NS NS NS < < 0.05 Aldosterone: 4 h Before amiloride 1.4 ± ± ± ± ± ±. 5. ± ± 0.9 After amiloride.7 ± ± ± ± ± ± ± ± 1.4 Probability value b < NS < NS NS NS < < Aldosterone: 7 h Before amiloride 13.4 ± ± ± ± ± ± ± ± 0.9 After amiloride.0 ± ± ± ± ± ± 1.6 -:1 ± ± 1.0 Probability value b < 0.05 <0.05 <0.05 NS NS NS < < NS, not significant. a The aldosterone animals received continuous infusion of aldosterone administered via subcutaneously implanted Alzet minipumps for 4, 4, or 7 h. There were 13,7,6, and 6 tissue pairs in the control and the three experimental groups respectively. See the footnote to Table 1 for additional details. b Difference between periods before and after amiloride addition. epithelial cell function (1,1). In previous studies designed to assess the role of aldosterone in the regulation of colonic electrolyte transport in the rat distal colon, we have determined sodium, chloride, and potassium fluxes across isolated colonic mucosa under voltage clamp conditions, as well as the electrophysiologic properties of this epithelium in normal and sodium-depleted rats (7,,13). Animals with dietary induced sodium depletion were used, as infusion of aldosterone to euvolemic animals can induce potassium depletion and may also elicit the "escape" phenomenon. Both electroneutral sodium chloride absorption and active potassium absorption are present in the distal colon of normal rats (7,). Amiloride-sensitive sodium absorption is either minimal or absent in normal animals, and it is likely that electroneutral sodium chloride absorption is a result of parallel ion exchanges, Le., Na-H and CI-HC0 3. Active potassium absorption is also electroneutral and is independent of both sodium and chloride, observations that are most consistent with the presence of a K-H exchange (7). In contrast to these findings in normal animals, sodium depletion induces several changes in electrolyte transport in the distal rat colon. Sodium depletion induces both amiloride-sensitive electrogenic sodium transport and amiloride-insensitive electrogenic potassium secretion (7,). In addition, electroneutral sodium chloride absorption is not present in sodium-depleted animals. These conclusions are based on the observations that sodium absorption in norma.l animals is chloride dependent and is not inhibited by 0.1 mm amiloride, but is inhibited by 1.0 mm amiloride (Le., high concentrations) and by theophylline and acetazolamide (). As it is generally recognized that cyclic adenosine monophosphate, carbonic anhydrase inhibitors, and high amiloride concentrations inhibit electroneutral sodium chloride absorption, these results indicate that electroneutral sodium chloride absorption is the predominant sodium absorptive process in the distal colon of the rat. In contrast, in sodium-depleted animals, sodium absorption is chloride independent, and is inhibited by 0.1 mm amiloride but not by theophylline. Sodium depletion also stimulates active potassium secretion (). These changes in electrolyte transport are accompanied by decreases in apical membrane voltage, apical membrane resistance, and basolateral membrane voltage (13). Although sodium depletion elevates plasma aldosterone levels, it is possible that these changes in colonic electrolyte transport are in part secondary to factor(s) other than aldosterone. e therefore designed studies to compare electrolyte transport in the distal colon of rats that had received continuous aldosterone infusions via implanted mini pumps for 7-1 days with sodium-depleted animals. Infusion of aldosterone resulted in plasma aldosterone levels that were comparable to those observed in dietary sodium deprivation. Electrogenic sodium absorption is best represented by an increase in Isc (Figure la) and the amiloride-sensitive component of Jnet Na (Fig-

6 13 HALEVY ET AL. GASTROENTEROLOGY Vol. 91, No.5 ure lb). Three parameters can be used to identify electroneutral sodium chloride absorption: (a) amiloride-insensitive component of Jnet Na (Figure 3B), (b) JnetNa-Isc (Figure 3A), and (c) Jnet C1. Previous studies of transmural potassium fluxes have established that in sodium-depleted animals, after the addition of amiloride to the mucosal bathing solution active potassium secretion is equivalent to the reversed Ise (see Figure ) (7). Based on these criteria, in both the sodiumdepleted and the long-term aldosterone groups, there is evidence for the induction of both amiloridesensitive, electrogenic sodium transport and active potassium secretion, and for inhibition of electroneutral sodium chloride absorption. Indeed, as shown in Table 1, electrolyte transport in these two groups is identical. These results establish that aldosterone is responsible for most, if not all, of the changes in electrolyte transport seen in the distal colon of sodium-depleted rats. The method of aldosterone infusion developed for these studies also provided an opportunity to determine whether the changes seen in the long-term aldosterone group were dependent on time. That is, was the same duration of time, during which plasma aldosterone levels were elevated, required to induce the several changes in sodium, chloride, and potassium transport? Recent studies indicate that inhibition of electroneutral sodium chloride absorption may be a function of the magnitude of the increase in plasma aldosterone levels. In animals fed an excess in dietary potassium, which is associated with a 10-fold increase in plasma aldosterone levels, both amiloride-sensitive electrogenic sodium absorption and active electrogenic potassium secretion are induced, but electroneutral sodium chloride absorption is not altered (14). These observations suggest that the inhibition of electro neutral sodium chloride absorption requires higher levels of plasma aldosterone than does the stimulation of electrogenic sodium and potassium transport. Analysis of sodium and chloride transport in the animals that received aldosterone for 4, 4, and 7 h reveals than the several changes in ion transport induced by aldosterone are not identical, but rather occur after varying periods of aldosterone infusions. Evidence of stimulation of electrogenic sodium absorption is observed after 4 h of aldosterone infusion, in that Ise and amiloride-sensitive Jnet Na in this group (.4 ± 0.6 and. ± 0.7 J-LEq/h. cm z, respectively) is significantly greater than that in the control group (Figure la). Furthermore, 0.1 mm amiloride induced comparable decreases in both Ise and Jnet Na in this group. It is likely that the initial induction of amiloride sensitivity and electrogenic sodium absorption occur at a considerably earlier time period, inasmuch as both amiloride sensitivity and an increase in potential difference have been observed 5 h after the administration of dexamethasone (15). Although dexamethasone has both glucocorticosteroid and mineralocorticosteroid actions bn ion transport (16,17)' recent studies suggest that amiloride sensitivity is a manifestation of the mineralocorticoid not the glucocorticoid effect. It also appears that aldosterone does not exert its maximal effect on electrogenic sodium transport at 4 h, as quantitatively greater changes in both Ise and the amiloride-sensitive component of Jnet a are observed in the 4-h and longterm groups (Figure 1). As the changes in these parameters are almost identical in the 4-h, 7-h, and long-term groups, aldosterone most likely induces changes in electrogenic sodium transport that begin between 5 and 4 h and become maximal after 4 h. Assessment of the effect of prolonged aldosterone on electroneutral sodium chloride absorption produced unexpected findings. Results presented in Table are best interpreted to indicate that electro neutral sodium chloride absorption is initially stimulated and then inhibited. It is clear that in both the sodium-depleted and the long-term aldosterone groups electroneutral sodium chloride absorption is abolished as Jnet C1 is reduced toward zero and Jnet a approximates Ise. In the 4-h aldosterone group, although Jnet Na is increased, which might solely represent an induction of electrogenic sodium absorption, Jnet C1 is also augmented. Furthermore, the increases in Jnet Na (5. J-LEq/h. cmz) and Jnet Na - Ise (.5 J-LEq/h. cmz) (Figure 3A) and the amilorideinsensitive component of Jnet Na (.1 J-LEq/h. cmz) (Figure 3B) are greater than the increase either in Ise (1.9 J-LEq/h. cmz) (Figure la) or in the amiloridesensitive component of Jnet Na (.0 J-LEq/h. cmz) (Figure lb), suggesting that both the electroneutral and the electrogenic components of sodium absorption are stimulated by aldosterone infusions given for 4 h. An adequate explanation for the stimulation of electroneutral sodium chloride absorption is not readily evident. As aldosterone increases sodiumpotassium-stimulated adenosine triphosphatase activity, an increase in sodium extrusion may increase the sodium-proton exchange with subsequent coupling to the Cl-HC0 3 exchange. It must be noted that in the proximal colon of the rat, sodium depletion has recently been shown to stimulate electroneutral sodium chloride absorption but does not include amiloride-sensitive sodium transport (1). In the 4-h and 7-h aldosterone groups, Jnet Na and Jnet C1 are similar to control values but less than those seen in the 4-h group. It is likely that infusion of aldosterone for 4 h inhibits electroneutral sodium chloride

7 November 196 ALDOSTERONE AND COLONIC ION TRANSPORT 133 absorption and that aldosterone inhibition is greater than its stirilulation of electroneutral transport. Electroneutral sodium chloride absorption is completely inhibited after 7-1 days of aldosterone infusion. The addition of amiloride to all experimental groups not only decreases but also reverses Ise (Figure ). In both sodium-depleted and potassiumloaded animals, active potassium secretion as determined by unidirectional 4K fluxes under voltage clamp conditions is responsible for this reversed Ise after amiloride addition (7,19). Thus, based on these studies we believe that the reversal of Ise by amiloride also represents active potassium secretion. It is therefore evident that aldosterone induces a maximal rate of active potassium secretion by 4 h, because after amiloride addition, the reversed Ise is similar in all five experimental groups with elevated plasma aldosterone levels (Figure ). Thus, the duration of time that aldosterone is in excess affects active sodium and potassium transport differently. These experiments do not provide an adequate explanation of how aldosterone modifies the apical membrane such that electroneutral sodium chloride transport is abolished. Nonetheless, it appears that aldosterone induces several significant changes in apical membrane function. The effects of aldosterone on e l e ~ t r osodium g e n i absorption, c electroneutral sodium chloride absorption, and active potassium secretion are not identical. Also, there is evidence that the effect of aldosterone is influenced by the magnitude of the elevation of plasma aldosterone levels as well as the duration of time in which excess aldosterone levels are present. References 1. Hayslett JP, Binder HJ. Mechanism of potassium adaptation. Am J Physiol 19;43:F Edmonds q. Transport of potassium by the colon of normal and sodium-depleted rats. J PhysioI1967;193: Edmonds q, Marriott J. Sodium transport and short-circuit current in rat colon in vivo and the effect of aldosterone. J Physiol (Lond) 1970;10: ill PC, Lebovitz FL, Hopfer U. Induction of amiloridesensitive sodium transport in the rat colon by mineralocorticoids. Am J PhysioI190;3:F Kashgarian M, Taylor CR, Binder HI. Hayslett JP. Amplification of cell membrane surface in potassium adaptation. Lab Invest 190;4: Hayslett JP, Myketey N, Binder HI. Aronson PS. Mechanism of increased potassium secretion in potassium loading and sodium deprivation. Am J PhysioI190;39:F Foster ES, Hayslett JP, Binder HJ. Mechanism of active potassium absorption and secretion in the rat colon. Am J Physiol 194;46:G Foster ES, Zimmerman T, Hayslett JP, Binder HJ. Corticosteroid alteration of active electrolyte transport in rat distal colon. Am J PhysioI193;45:G Binder HI. Rawlins CL. Electrolyte transport across isolated large intestinal mucosa. Am J PhysioI1973;5: Martin RS, Jones I. Hayslett JP. Animal model to study the effect of adrenal hormones on epitheljal function. Kidney Int 193;4: Snedecor G, Cochran G. Statistical methods. 6th ed. Ames, Iowa: Iowa State University Press, 1967: Edmonds q. Transport of sodium and secretion of potassium and bicarbonate by the colon of normal and sodium-depleted rats. J Physiol (Lond) 1967;193: Sandie GI, Hayslett JP, Binder HJ. Effect of chronic hyperaldosteronism on the electrophysiology of rat distal colon. Pfiugers Arch 194;401 : Budinger ME, Foster ES, Hayslett JP, Binder HJ. Sodium and chloride transport in the large intestine of potassium-loaded rats. Am J Physio!196;51:G Sandie GI, Lewis SA, Hayslett JP, Binder HJ. Dexamethasone induces amiloride-sensitive Na transport in rat distal colon (abstr). Clin Sci 19;6: Marusic ET, Hayslett JP, Binder HJ. Corticosteroid-binding studies in cytosol of colonic mucosa of the rat. Am J Physiol 191;40:G Binder HJ, hite A, hiting D, Hayslett J. Demonstration of specific high affinity receptors for aldosterone in cytosol of rat colan. Endocrirtology 196;11: Faster ES, Budinger ME. Hayslett JP, Binder HJ. Ion transport in proximal colon of the rat: sodium depletion stimulates neutral sodium chloride absorption. J Clin Invest 196; 77: Foster ES, Sandie GI, Hayslett JP, Binder HJ. Dietary potassium modulates active potassium absorption and secretion in rat distal colon. Am J Physiol (in press).