interrelationships of chloride,...

Interrelationships of Chloride, Bicarbonate,

Sodium, and Hydrogen Transport in the Human Ileum

LEsLE A. TURNBERG,FREDERICKA. BIEBERDORF, STEPHENG. MORAWSKI,and JOHNS. FORDTRAN

From the Department of Internal Medicine, The University of Texas South-western Medical School at Dallas, Texas 75235

A B S T R A C T Using a triple-lumen constant perfusionsystem, the following observations were made in normalsubjects. First, chloride, bicarbonate, and sodium werefound to exhibit net movement across ileal mucosaagainst electrochemical gradients. Second, during per-fusion with a balanced electrolyte solution simulatingplasma, the ileum generally absorbed, but sometimes se-creted fluid. A reciprocal net movement of chloride andbicarbonate was noted when sodium movement waszero. Increasing rates of sodium absorption were as-sociated with decreasing bicarbonate secretion ratesand finally bicarbonate absorption. Even when bicarbo-nate was absorbed ileal contents were alkalinized (bycontraction of luminal volume). Third, net chloridemovement was found to be sensitive to bicarbonate con-centration in ileal fluid. For instance, chloride was ab-sorbed from solutions containing 14 or 44 mEq/liter ofbicarbonate, but was secreted when ileal fluid contained87 mEq/liter of bicarbonate. Fourth, when chloride-free (sulfate) solutions were infused, the ileum absorbedsodium bicarbonate and the ileal contents were acidified.Fifth, when plasma-like solutions were infused, the po-tential difference (PD) between skin and ileal lumenwas near zero and did not change when chloride wasreplaced by sulfate in the perfusion solution.

These results suggest that ileal electrolyte transportoccurs via a simultaneous double exchange, Cl/HCOsand Na/H. In this model neither the anion nor the ca-tion exchange causes net ion movement; net movementresults from the chemical reaction between hydrogen andbicarbonate. No other unitary model explains all of thefollowing observations: (a) human ileal transport invivo is essentially nonelectrogenic even though Na, Cl,and HCO3 are transported against electrochemical

Dr. Turnberg's present address is Manchester Royal In-firmary, Oxford Road, Manchester 13, England.

Received for publication 3 February 1969 and in revisedform 2 October 1969.

gradients, (b) the ileum can secrete as well as absorb,(c) ileal contents are alkalinized during absorption of orduring secretion into a plasma-like solution, and (d) theileum acidifies its contents when sulfate replaces chlo-ride. Data obtained with a carbonic anhydrase inhibitorsupport the proposed model.

INTRODUCTION

Current concepts of the mechanisms of ileal electrolyteabsorption (1, 2) are derived from both in vitro andin vivo experiments, although in recent years relativelyfew in vivo studies have been carried out. The mostgenerally held view is that electrolyte absorption invitro is due to active sodium transport and that theelectrical potential so generated is the main force foranion absorption (1). In most species that have beentested, electrolyte absorption in vivo is not associatedwith a potential difference across ileal mucosa, and thishas been attributed to simultaneous active transport ofsodium and chloride, both pumps electrogenic but of thesame magnitude so that no net potential difference isgenerated (1). In vivo, concentrations of chloride andbicarbonate vary reciprocally in ileal fluid with theirsum equal to approximately 135 mEq/liter; the bicarbo-nate concentration is generally higher and the chlorideconcentration lower than their respective plasma levels(1, 2). Bucher, Flynn, and Robinson (3) were ap-parently the first to suggest that the appearance of bi-carbonate in ileal fluid is linked to the absorption ofchloride (anion exchange), but it is important to notethat reciprocal net movements of chloride and bicarbo-nate do not occur during absorption of balanced elec-trolyte test solutions but that chloride absorption is al-ways much higher than bicarbonate secretion. Thus,as judged by the rate of bicarbonate secretion, verylittle chloride could be absorbed by anion exchange.

The Journal of Clinical Investigation Volume 49 1970 557

We have recently conducted a series of experimentsin the human ileum in vivo which are not compatiblewith these concepts and which have led us to proposea different mechanism for ileal ion transport.

METHODSThe subjects of this study were normal male and femalevolunteers, aged 21-35 yr. The Ingelfinger triple-lumen per-fusion technique was used exactly as described in detailin earlier publications (4). The intestinal test segment was30 cm and the nonabsorbable marker was polyethylene gly-col. Studies were done when the infusion tip of the triple-lumen tube was 200-250 cm from the teeth. Unless other-wise stated, the infusion rate was 10 ml/min.

The mean concentration of solute in the test segment wascalculated by averaging the concentration in fluid with-drawn from each end of the test segment. Results are ex-pressed in milliliters of water or milliequivalents of elec-trolyte absorbed or secreted per hour per 30 cm of ileum.Usually three different test solutions were studied in thesame individual on one morning.

Potential difference (PD) between intestinal lumen andabraded skin was measured during perfusion with varioustest solutions by means of an intraluminal electrode de-scribed in an earlier publication from this laboratory (4).PD was measured during perfusion with the identical solu-tions used for measurement of ion movements and in thesame subjects, although the tests were performed on dif-ferent days.

To validate the use of abraded skin as a reference site PDbetween skin and peritoneum was measured four times inpatients undergoing peritoneal dialysis. With the skin asa reference, peritoneal PD + 2.0, + 0.2, 0.0, and - 0.2 mv inthe four studies. Therefore, our results of PD measurementsbetween skin and gut lumen should be approximately thesame as between lumen and peritoneum.

Electrolyte and polyethylene glycol (PEG) concentrationswere estimated with methods described previously (4). Totalcarbon dioxide was measured by the microgasometricmethod using a Natelson microgasometer (Model 600, Sci-entific Industries, Inc., Springfield, Mass.). pH and PcO2were measured on an Ultramicro pH/Pco2 gas analyzer(Instrumentation Laboratories, Inc., Lexington, Mass.).

The results are given as mean values ± 1 SE of the meanin the tables and figures.

RESULTS

Absorption and secretion ratesChloride. Ileal chloride absorption at varying chlo-

ride concentrations was assessed in nine subjects. Testsolutions contained 0, 50, or 100 mEq/liter of chlorideas the sodium salt. Sodium and potassium bicarbonateand mannitol were added in amounts required to giveeach solution a bicarbonate concentration of 30 mEq/liter, a potassium concentration of 5 mEq/liter, and anosmolality of 285 mOsm/kg. As shown in Fig. 1, chlo-ride absorption increased linearly with increasing meanluminal chloride concentration over the range of 10-94mEq/liter. Absorption occurred against a considerablelumen-to-plasma concentration gradient.

A ChloridemEq/hr/30 cm

- +22.3mV"F (t5.2)

+11.6mV(t 3.1)

+ 3.7 mV P D(t 2.5)

-6hc0

o -4en

-o

-2 -

c0

0u-4)0

(I)

Ok

20 40 60 80 100 120 140CHLORIDE CONCENTRATION

(mEq /liter)

FIGURE 1 Effect of luminal chloride concentration on netchloride movement. Sodium concentration was approximately25 mEq/liter higher than chloride; bicarbonate concentrationwas 30 mEq/liter in all studies. The potential difference(PD) between ileal lumen and skin at different sodiumchloride concentrations is given at the top of the figure. Thesign preceding a PD value indicates the orientation of thepotential, + meaning lumen positive compared to skin and- meaning lumen negative compared to skin.

The PD between skin and ileal lumen was measuredin seven subjects during perfusion of identical solutionsas in the above experiments and the mean values forthese PDs are also shown in Fig. 1. As luminal chloride(and sodium) concentration was reduced, the lumenbecame increasingly electropositive with respect to theskin. Thus chloride absorption occurs against both steepconcentration and steep electrical gradients.

Sodium. Ileal sodium absorption, measured duringperfusion of the test solutions just described, alsooccurred against steep concentration gradients, as pre-viously described in detail (4). However, unlike chlo-ride, sodium absorption occurred in the direction of,and not against, an electrical gradient. Utilizing theNernst equation, it is possible to relate chemical andelectrical gradients and to infer whether sodium move-ment is passive or active, at least as defined by thisequation. For instance, in the present studies sodiumabsorption continued until sodium concentration wasreduced to approximately 27 mEq/liter; at lower con-centrations, sodium was secreted into the ileal lumen.By the Nernst equation, electrical PD would have tobe 43 mv to account for this unequal distribution ofsodium ions between ileal lumen and plasma. The ob-served PD was only approximately 22 mv, indicatingthat sodium was not distributed passively across the ilealmucosa and that ileal sodium absorption occurs againstelectrochemical gradients.

558 L. A. Turnberg, F. A. Bieberdorf, S. G. Morawski, and J. S. Fordtran

.1

TABLE ISlow Perfusion Studies. Changes in Bicarbonate Concentration Compared with Changes in

PEGConcentration and Net Bicarbonate Movement

Increase inBicarbonate bicarbonate Increase in PEG

concentration concentration concentrationat distal as- from proximal from proximal to

Study piration site to distal site distal site A HC03*

mEqlliler % % mEq/hr per 30 cmInfused bicarbonate

concentration30 mEq/liter

1 51.7 48.1 17.9 +0.512 58.3 52.6 29.4 +0.373 31.5 32.3 18.3 +0.294 52.2 56.8 35.3 +0.345 61.9 43.3 35.3 +0.17

35 mEq/liter6 41.6 8.6 36.5 -1.297 42.3 23.0 14.9 +0.248 41.2 38.8 19.6 +0.2791 47.3 18.3 - 21.3T +1.65

40 mEq/liter10 48.3 17.2 41.8 -0.4811 50.1 34.7 43.3 -0.2212 39.1 54.5 7.6 +0.9013 47.6 37.2 102.4 -0.81

* + sign indicates net secretion, - sign indicates net absorption.I Subject in whom net water movement was into the lumen.

Bicarbonate. Preliminary studies revealed that netbicarbonate movement' from balanced electrolyte solu-tions was small and difficult to measure accurately withthe standard perfusion technique. Therefore, experi-ments were performed at a slow rate of perfusion, 1.7ml/min, over a long (4 hr) period of time. This slowperfusion accentuates ionic and PEG concentrationchanges and markedly enhances the accuracy of anygiven experimental result. The long perfusion period,which was preceded by 1 hr equilibration, precludedmore than a single experiment on a given day.

Perfused test solutions contained 30, 35, or 40 mEq/liter of bicarbonate, 105 or 110 mEq of chloride, 135 or140 mEq of sodium, and 5 mEq/liter of potassium. Re-sults of 13 slow perfusion studies are shown in Table I.

1 Difficulties arise in deciding which of several mechanismsis responsible for bicarbonate movement. Removal of bi-carbonate from the lumen could be achieved by absorptionof bicarbonate ions or by secretion of hydrogen ions. Theidentical effect to hydrogen secretion could be produced byhydroxyl ion absorption from water which would leavean excess of hydrogen ion in the lumen. Bicarbonate accumu-lation could occur by reversal of any of these processes.Unless otherwise specified the terms bicarbonate "absorp-tion" and "secretion" are used in the loose sense to indicateremoval from or accumulation within the lumen achieved byone of the above processes.

In every study bicarbonate concentration increased asfluid passed from proximal to distal aspiration site.Since water absorption occurred as shown by the risein PEG concentration, some of the increase in bicar-bonate concentration was due to a contraction of luminalvolume. However, as revealed in the table, per cent risein bicarbonate concentration between proximal and dis-tal aspiration sites was higher than the per cent rise inPEGconcentration in 10 of 13 studies. In these experi-ments bicarbonate secretion was at least partly respon-sible for the high concentrations achieved in the ileum.In the three experiments in which bicarbonate absorp-tion was noted, the rise in bicarbonate concentration wasdue entirely to a shrinkage of ileal volume due to rapidwater absorption.

PD was measured four times during slow perfusionof the same test solution. Mean PD was +0.5 ±3.3 mv.

Relation between water and ion absorption ratesduring perfusion with a balanced electrolytesolutionBecause of the high degree of accuracy of the slow

perfusion studies, these experiments were chosen toexamine the relation between sodium and anion move-ments in individual perfusion periods. These results are

Real Electrolyte Transport 559

-8 r

-6

E0

0*

W, on

crwE

-4

-2

+2

NaoIII /" crII ~U

U HCO;

S t AbssorptionI Secretion

+30 +20 +10 0 -10 -20 -30 -40 -50 -60 -70SECRETION ABSORPTION

A WATERml /h r/30 cm

FIGuRE 2 Relation of ion and water movements in slow (1.7ml/min) perfusion studies. Regression lines were drawn by methodof least squares. P values for difference between chloride andsodium and sodium and bicarbonate movements at -zero water flowwere < 0.001 and < 0.001, respectively.

shown in Fig. 2, where sodium, chloride, and bicar-bonate movements are related to water movement. Meanluminal sodium concentration was 137 (range 125-143),mean potassium concentration was 4.9 (4.5-6.0), andmean chloride concentration was 97 (87-108) mEq/liter. Mean bicarbonate concentration varied between32.2 and 52.6 mEq/liter, depending on the rate of waterabsorption and bicarbonate secretion. Several conclu-

sions can be drawn from the data and regression linesshown in Fig. 2. First, it is clear that zero sodiumabsorption is associated with zero water movementwhich is compatible with the thesis that water move-ment is a passive consequence of net solute movement(1). Second, when water and sodium absorption arezero, bicarbonate is secreted and chloride is absorbed atapproximately equal rates. This is suggestive of an

A ChloridemEq/hr/30cm

-4F

c0

.0a4mo

-3-

-21

-I F

0

c0

14)Cf)

+1

+21

+30 10 20 30 40 50 60 70 80 90BICARBONATE CONCENTRATION

(mEq /liter)

FIGURE 3 Effect of luminal bicarbonate concentration on netchloride movement. Chloride and sodium concentrations wereconstant at 40 and 135 mEq/liter, respectively; sulfate wasadded for ionic balance, and all test solutions were isotonic.


- - - - - - -

..-I

-6 1HC03-4-

-2 - No

0 -

+2

.410 100

-6 -

-4 -

O _

.4

NoAHCO3

CI

10010

"sNo/10 HC03

OC3

'-c Cl N

A BicarbonatemEq /hr/30cm

c

0Cb.0

in

.04

c

0

0U)

+3.7mV PD(±2.5)

20 40 60 80CHLORIDE CONCENTRATION

(mEq /liter)

100

FIGURE 5 Effect of luminal chloride concentration on bi-carbonate movement. Luminal sodium paralleled the chlorideconcentration but was approximately 25 mEq/liter higher ateach of the three points. Luminal bicarbonate concentrationwas constant (24.6-27.7 mEq/liter), regardless of sodiumchloride concentration.

sulfate. Isotonicity was maintained with mannitol. 10subjects were studied with each of the three testsolutions.

As shown in Fig. 3, luminal bicarbonate concentrationinfluenced chloride movement despite the maintenanceof a constant chloride concentration. When the meanluminal bicarbonate concentration was 14 mEq/liter,chloride was absorbed against a concentration gradient.However, chloride absorption was reduced at a bicar-bonate concentration of 44 mEq/liter; and when meanbicarbonate concentration was 87 mEq/liter, chloridewas secreted.

Five additional studies were performed to examinemore closely the net movements of chloride and bi-carbonate as bicarbonate concentration was varied. Inorder to increase accuracy the perfusion rate was re-duced from the standard rate of 10 ml/min to 5 ml/min,and each study period lasted 2 hr instead of 1 hr. Testsolutions had a chloride concentration of 40 mEq/literand bicarbonate concentration was either 10 or 100

mEq/liter. Sodium concentration was 135 mEq/liter inboth solutions with the missing anion being providedby sulfate. As before, isotonicity was maintained withmannitol.

As shown in Fig. 4, raising the bicarbonate concen-tration of the infused solution from 10 to 100 mEq/liter caused chloride absorption to be reduced or chloridesecretion to be induced, and the change in net bicar-bonate movement was approximately the same as thechange in chloride movement, except in those subjectsin whom sodium absorption was different in the twostudies. These experiments confirm a reciprocal netmovement of chloride and bicarbonate as luminal bicar-bonate concentration is varied.

Measurements of PD were made during perfusion ofsolutions with varying anion concentrations while so-dium concentration was maintained at 135 mEq/liter.Table II demonstrates that PD does not change signifi-cantly when chloride and bicarbonate concentrations arevaried reciprocally or when both are largely replacedby sulfate. Therefore, the changes' in chloride movementinduced by altering luminal bicarbonate concentrationoccur in the absence of change in PD.

Effect of chloride concentration on bicarbonatemovementThe effect of varying (sodium) chloride concentration

on ileal bicarbonate movement was assessed in 10 sub-jects. Luminal contents had a constant bicarbonate con-centration (24.6-27.7 mEq/liter) and osmolality wasmade constant at 285 mOsm/kg by addition of appro-priate amounts of mannitol to the test solutions. Asshown in Fig. 5, bicarbonate was secreted at a slowrate when luminal chloride concentration was 94 mEq/liter. However, bicarbonate was absorbed when chlorideconcentration was lowered to 50 and 10 mEq/liter.

PD was +22.3 mv (lumen positive) when lumenchloride concentration was 10 mEq/liter, and +3.7 mvwhen chloride concentration was 94 mEq/liter. Thus ab-

TABLE II IIon Movement in the Presence and Absence of Chloride (Mean Values ± 1 SE).

Concentrations Refer to Mean Concentration in Test Segment

PCo2

AH20 [Na] ANa [K] AK [C1] ACI [HCOa- AHCO3 Infusion Proximal Distal

Chloride- -12.6 142.2 -1.30 5.13 -0.02 11.3 -0.01 15.5 -1.35 53.9 61.8 57.7freesolution +3.1 +0.6 40.50 +0.12 4+0.03 42.1 +0.28 +1.7 40.28 +t1.8 +1.1 +2.3n = 6

Chloride- -24.9 141.0 -3.50 5.30 +0.00 98.5 -4.13 34.3 +0.63 52.1 57.4 56.7containingsolution +9.8 +-0.4 +1.45 +0.09 40.12 +2.3 +-1.34 +2.7 +0.29 +2.1 +2.6 +2.8n = 6


sorption of bicarbonate occurred against a PD of 22.3mv at a chloride concentration of 10 mEq/liter, whileat the highest chloride concentration (94 mEq/liter),bicarbonate movement (secretion) was in the directionfavored by the small PD gradient (3.7 mv).

These experiments show that net bicarbonate move-ment is influenced by luminal sodium chloride concen-tration and that bicarbonate can be absorbed againstelectrochemical gradients (see footnote 1).

Ion transport during sodium sulfate perfusionSince the studies recorded in Fig. 5 indicated that

bicarbonate can be absorbed against electrochemicalgradients when luminal chloride concentration is low, itwas decided to study the effect of chloride removal ingreater detail, especially in regard to its effect on Naand K movements and on pH and Pco2 of luminal fluid.To make these observations, the ileum was perfusedwith an electrolyte solution containing 140 mEq/literNa, 5 mEq/liter K, 25 mEq/liter HCOs, and no Cl.The missing anion was sulfate, and mannitol was addedto bring the final osmolality to 290 mOsm/kg. A con-trol solution had the same bicarbonate, sodium, andpotassium concentrations but contained 100 mEq/literof chloride rather than sulfate. The infused test solu-tions were bubbled with 5% C02 for 30 min before andduring the intestinal perfusions.

7.6 F

7.51-

7.4 H

pH 7.3 F7.21

7.1p-

7.0

6.9 p Infusionsolution

Proximalsite

Distalsite

FIGURE 6 Effect of chloride and chloride-free (sulfate)solutions on pH of infused test solution and fluid collectedfrom the proximal and distal collection sites of the three-lumen tube. Test solutions were bubbled with 5%o C02 beforeinfusion. Sodium, potassium, and bicarbonate concentrationsof both test solutions were 140, 5, and 25 mEq/liter,respectively.

c

oEca

000

-

.- Cl-free solution -FIO--cI soluton A|

-40 -

-30 _

-20 _

-lo0

o

0 20 40 60 80 100 120

TIME IN MINUTESFIGURE 7 Potential difference (lumen to skin) during per-fusion with chloride and chloride-free (sulfate) test solu-tions. All test solutions contained 140, 5, and 25 mEq/literof sodium, potassium, and bicarbonate, respectively. A posi-tive PD indicates lumen positive with respect to skin; anegative potential means lumen negative with respect to skin.

Table III shows the mean concentration of ions inthe test segment and their movements for the two infu-sion solutions. Substitution of sulfate for chloride wasassociated with a reduction in sodium absorption from3.5 to 1.3 mEq/hr. Potassium movement was the samein the presence or absence of chloride. Bicarbonate wassecreted in the presence of chloride and absorbed in itsabsence.

Fig. 6 shows pH of the infusion solutions and fluidcollected at proximal and distal sites under a layer ofoil. The pH values of both infusion solutions rangedfrom 7.30 to 7.40. Whereas the chloride-containing solu-tions either became more alkaline or did not show apH change, the pH values of the chloride-free solutionsfell. At the proximal collecting site, the pH had fallento values between 7.09 and 7.25. Between the proximaland distal sites, the pH continued to fall to values be-tween 6.94 and 7.16. As shown in Table III, luminalPco2 was approximately the same during perfusion ofthe two test solutions.

As shown in Fig. 7, substitution of sulfate for chlo-ride had no demonstrable effect on PD. Three otherstudies gave similar results.

Carbonic anhydrase inhibitor.The effects of acetazolamide' (200 mg/liter of per-

fused fluid) on sodium, chloride, and bicarbonate move-ments were tested during perfusion of a balanced electro-lyte solution. As shown in Fig. 8, absorption of sodiumand chloride was markedly inhibited by acetazolamide.

2 Diamox, American Cyanamid Co., Lederle LaboratoriesDiv., Pearl River, N. Y.

Heal Electrolyte Transport 563

There was no consistent effect on net bicarbonate move-ment which was near zero with and without aceta-zolamide.

Sodium diffusion potentialAs shown in Table II and in Fig. 7, the PD between

ileal lumen and skin is not affected by reciprocal varia-tion of chloride and bicarbonate or chloride and sulfate.By contrast, progressive lowering of luminal sodiumconcentration is associated with a progressive increasein PD with the lumen positive to skin. These results,illustrated in Fig. 9, are compatible with a sodiumdiffusion potential. Similar results were obtained in therat by Wright (5) and this conclusion is compatiblewith PD measurements in man reported by Gustke,Whalen, Geenen, and Soergel (6). As shown in TableII, the average PD across ileal mucosa is close to zerowhen the ileum is perfused with an electrolyte solutionsimulating plasma; when glucose is added to such solu-tions, the PD becomes substantially negative, as pre-viously reported (4).

DISCUSSIONThese experiments were designed primarily to elucidatethree aspects of ileal transport. First, we measured ionmovement and PD as ion concentrations varied in orderto establish which ions equilibrate passively and whichmust be subjected to some special transport process.Second, we determined the interrelationship between themovement of various ions by carefully measuring Na,K, Cl, and HCO3 movements during perfusion of abalanced electrolyte solution and by assessing the effectof the concentration of one ion on the movement of theothers. Finally, by measuring potential difference under

Nao

c0

0.

a.04

0

-

cl

-4 1 I I ICON- ACET. CON- ACET.TROL TROL

FIGURE 8 Effect of acetazolamide on sodium and chloridemovements during perfusion with a balanced electrolytesolution.

+30

+25

' +20

_ +150

.O-

> +10E0X. +5

0

50 75 100 15

SODIUM CONCENTRATION(mEq /liter)

FIGURE 9 Effect of luminal sodium concentration on poten-tial difference (mean ±+1 SE) between ileal lumen and skin.Orientation of PD is the same as in Fig. 1.

a variety of experimental conditions, we attempted todetermine whether or not ion transport in the ileumin vivo is electrogenic.

Ion movement against electrochemical gradientsThe results presented in Fig. 1 provide clear evidence

for absorption of chloride against a steep electrochemicalgradient. Similarly, the data in Table I suggest thatbicarbonate secretion usually occurs against an electro-chemical gradient when luminal chloride concentrationis approximately 100 mEq/liter. However, when lumi-nal chloride concentration is low, bicarbonate movementin the reverse direction, i.e. absorption, occurs againstelectrochemical gradients (Fig. 5, Table III). Thus,under different conditions there is evidence for move-ment of bicarbonate against an electrochemical gradientin either direction across the membrane. In addition tothe apparently active nature of these transport processes,it is clear that the concentration of one anion markedlyinfluences movement of the other.

The present results confirm previous investigations re-ported from this laboratory suggesting that sodium isactively transported in the human ileum (4). In thepresent studies the orientation of the PD developedacross the ileal membrane (lumen positive) favoredsodium absorption from ileal solutions having a lowsodium concentration; however, the magnitude of thisPD could not alone account for the observed concentra-tion gradient against which sodium was absorbed.Furthermore, when 135-140 mm saline solutions plusglucose are perfused, the ileal lumen becomes electro-negative (Table II and reference 4); sodium absorptioncontinues under these conditions demonstrating conclu-


sively that sodium can be absorbed against electro-chemical gradients. We. found no evidence for potassiummovement against an electrochemical gradient in thenormal human ileum.

Interrelation between ion movementsVariation of normal ileal function during balanced

electrolyte perfusion. In previous ileal perfusion ex-periments, wide variation in the rates of sodium andwater absorption from day to day in the same subjectand between different normal subjects has been noted.Secretion rather than absorption is occasionally ob-served (unpublished observations of the authors). Simi-lar findings were seen in the slow perfusion studiesreported in the present paper, and the high accuracy ofthese studies precludes experimental error as an explana-tion for this variation in ileal behavior. Secretion ofsodium, chloride, bicarbonate, and water was observedat one end of the scale and absorption of all these atthe other. This marked diversity of ileal activity oc-curred in response to factors not defined in the presentstudy; unknown hormonal or circulatory phenomenamay have played a part.

The interrelation between different ion movements atvarying rates of absorption or secretion was also ofinterest. Only when sodium movement was zero did anequimolar anion exchange become obvious. Sodiumabsorption was associated with a decrease in bicarbonatesecretion and at high sodium absorption rates, bicar-bonate was absorbed; sodium secretion, on the otherhand, was associated with enhanced bicarbonate secre-tion. Regardless of the rate or direction of fluid move-ment, ileal fluid becomes alkaline. During fluid secretion,alkalinity results from bicarbonate secretion; during ab-sorption, alkalinity results from water absorption (whichcauses luminal bicarbonate concentration to rise) andin many instances also from bicarbonate secretion.

Effect of bicarbonate concentration on chloride move-ment. The data presented in Figs. 3 and 4 clearlydemonstrate that chloride movement is dependent uponluminal bicarbonate concentration. This effect was notmediated by a change in PD, as shown in Table II. Al-though these observations, like the reciprocal anionmovement at zero sodium movement during balancedelectrolyte perfusion, suggest a chloride/bicarbonate ex-change, it is clear that net chloride absorption fromphysiologic solutions is much greater than net bicar-bonate accumulation within the lumen. For instance,when luminal contents were similar in ionic compositionto plasma, chloride was absorbed at a rate of 6.5 mEq/hr per 30 cm, mainly as sodium chloride, while bicar-bonate was secreted at a rate of only 0.1 mEq/hr per30 cm (compare Figs. 1 and 5). Thus, that portion ofchloride absorption normally mediated via an exchange

must be exceedingly small if judged by the net bicar-bonate accumulation rate. However, this ignores thepossibility that some of the exchanged, i.e. secreted,bicarbonate may have been removed by a separateprocess, such as hydrogen secretion.

Real ion movements during perfusion of chloride-freesolutions. The studies described in Table III and Fig.6 demonstrate that when sodium plus a nonabsorbableanion (sulfate instead of chloride) is perfused, bicar-bonate is absorbed and hydrogen ions accumulate in theileal lumen. Several mechanisms might explain this ob-servation. Active sodium absorption in the absence ofan absorbable anion might cause the intraluminal PDto become negative with the result that hydrogen andpotassium would be secreted passively. This possibilityis unlikely since PD was the same when the ileum wasperfused with chloride-containing or chloride-free solu-tions and since sodium absorption from chloride-freesolutions did not stimulate potassium secretion. At leasttwo possible explanations remain. First, a neutral so-dium bicarbonate pump might become activated whenchloride is removed from the ileal lumen. Second, asodium-hydrogen exchange carrier could effect sodiumbicarbonate absorption without a change in PD. Ourdata do not, unfortunately, provide a clear indicationthat either of the last two possibilities is correct. How-ever, two facts are clear. The ileum continues to absorbsodium in the absence of luminal chloride, and sodiumabsorption under these conditions is associated withbicarbonate absorption and acidification of ileal contents.

Nonelectrogenic nature of ileal ion transportin man

The experiments reported here suggest that ion trans-port in the normal human ileum in vivo does not gen-erate an electrical potential across ileal mucosa. Thisstatement is based on the following pieces of evidence:

(a) when balanced electrolyte solutions are perfusedthrough the human ileum, the average PD between skinand ileal lumen (which should approximate lumen/peri-toneum PD as discussed in Methods) is near zero; thelumen is slightly positive as often as it is slightly nega-tive. This observation has also been made in ileum ofexperimental animals in vivo, although the interpreta-tion was that chloride and sodium were actively ab-sorbed at the same rates by separate (and negating)electrogenic pumps (1).

(b) When the human ileum is perfused with a bal-anced electrolyte solution in which chloride has beenreplaced by poorly absorbed sulfate, the PD betweenskin and lumen is the same as when a chloride-contain-ing solution is perfused. This suggests that an electro-genic chloride pump is not negating the potential of anelectrogenic sodium pump when chloride-containing so-

lleal Electrolyte Transport 565

cl I , _ Cl

ILEALLUMEN PLASMA

FIGURE 10 A double exchange model that best explains our observa-tions on the relation of different ion movements in the ileum.

lutions are perfused.3 Sodium (bicarbonate) absorptionfrom a chloride-free solution is not accompanied bypotassium secretion which is further evidence against achange in PD since potassium probably equilibratespassively across ileal mucosa (1, 2).

(c) In a low resistance epithelium in which net ionfluxes are small compared to unidirectional fluxes,sodium transport might be electrogenic but produce alow potential that could not be distinguished experi-mentally from zero. (We probably could not pick up aPDchange less than 2-3 mv by our system.) Fortunately,however, we have some idea of the magnitude of PDthat an electrogenic sodium pump would have to gen-erate in order to account for the bicarbonate concentra-tion gradient that is developed across ileal mucosaduring perfusion of chloride-free solutions. As shownin Table III, the mean bicarbonate concentration in thestudy segment fell to 15.5 mEq/liter. The average con-centration of bicarbonate at the end of the study seg-

3 This analysis assumes equal passive permeability of theileum to sulfate and chloride. Only if the ileum were morepermeable to sulfate than to chloride would the observedresults be compatible with electrogenic sodium transport(sulfate diffusion potential cancels sodium transport poten-tial). Had the lumen become negative under these experi-mental conditions, this might indicate either electrogenicsodium transport or a higher ileal permeability to chloridethan sulfate (chloride diffusion potential).

ment was 13 mEq/liter. To account for this chemicalgradient on an electrical basis would require a PD of17 mv, assuming a plasma bicarbonate concentration of25 mEq/liter. A PD change (as chloride is replaced bysulfate) of this magnitude would be easily detectable.Since no PD change at all could be observed, an elec-trically neutral mechanism must be postulated forsodium bicarbonate absorption under these experimentalconditions.

The PD change observed when the sodium chlorideconcentration of ileal contents was progressively low-ered was clearly related to sodium and not to chlorideconcentration, i.e., a sodium diffusion potential. Theseresults suggest a much higher permeability of ileal mu-cosa to sodium than to anions and are consistent withthe finding that the maximum electrochemical gradientsagainst which sodium can be absorbed is much smallerthan that against which chloride can be absorbed. Thismay also help explain why glucose added to electrolytesolutions perfusing the human ileum causes the lumen tobecome electrically negative even though sodium ab-sorption is not increased (4). Glucose enhances ilealwater absorption, and its effect on PD might be due toa streaming potential (7). This possible mechanism forthe glucose effect has not, to our knowledge, been ruledout in vitro.


A model for ion transport in the ileumAny complete model for ileal transport must take into

account the ability of the ileum to secrete as well asabsorb, the interrelations of ionic movements at differ-ent rates of absorption or secretion, the effect of bicar-bonate concentration on chloride movement, the bicar-bonate absorption and acidification of luminal contentswhen the ileum is perfused with sodium plus a non-absorbable anion, and the nonelectrogenic nature of iontransport. The simplest model which fits our data andobservations is one in which a chloride-bicarbonate ex-change is linked with a sodium-hydrogen exchange.

According to this double exchange hypothesis, illus-trated in Fig. 10, neither the anion nor the cationexchange brings about net ion movement with the resultthat an electrical potential difference is not generated.Net movement observed by measuring the disappearanceor accumulation of solute is the consequence of thechemical reaction between hydrogen and bicarbonateto form water and C02. If the rate of the anion andcation exchange is exactly equal, the model functions asa NaCl pump and observed bicarbonate movement iszero; on the other hand, if the rate of the two exchangesis not equal, net bicarbonate movement will be observed.For instance, when chloride absorption exceeds sodiumabsorption, more bicarbonate than hydrogen would enterthe lumen resulting in net bicarbonate secretion. Con-versely, when sodium absorption exceeds chloride ab-sorption, more hydrogen than bicarbonate enters thelumen resulting in net bicarbonate absorption. Ilealsecretion can occur by reversing the usual direction ofthe two exchange mechanisms.'

Since the rate of movement of sodium and chloride isoften different and since sodium is absorbed even whenchloride is completely replaced by sulfate, it is clearthat the two exchange processes can operate inde-pendently of each other. However, independent cation oranion exchange would cause rapid development of steepion concentration gradients across ileal mucosa and thispresumably creates a passive functional link between thetwo exchanges so that activity in one exchange processtends to be followed by similar activity in the other.

The inhibition of sodium and chloride absorption bythe carbonic anhydrase inhibitor, acetazolamide, alsolends support to the model proposed in Fig. 10. Inhibi-tion of carbonic anhydrase, which is known to be presentin the ileum of laboratory animals (8, 9), would be ex-

Alternately, secretion might be explained by a separatedouble exchange carrier system, oriented to secrete sodiumand chloride in exchange for absorbed hydrogen and bicar-bonate. This separate system might be located in the sameor in different cells of the intestinal mucosa. Secretion wouldresult when the magnitude of transport in this second systemwas greater than that of the NaCl absorption oriented doubleexchange.

pected to inhibit sodium and chloride absorption if, in theproposed model, hydrogen and/or bicarbonate exchangedfor sodium and chloride are derived from carbon dioxideand water under the influence of carbonic anhydrase.These experiments cannot be used as conclusive prooffor the model, however, since it is possible that aceta-zolamide exerts its effect directly on sodium and chlorideabsorption and not via inhibition of carbonic anhydrase(10).

On the basis of our data, it is not possible or neces-sary to specify the location of the exchange processes;these could reside on one or more of the several mem-branes that separate luminal fluid from plastha.

Although we are well aware that we have not pre-sented definitive proof for this hypothesis, it is worthnoting that we and many others have been unable todesign or suggest an alternate unitary model that satis-factorily explains our experimental observations.

ACKNOWLEDGMENTSDr. Floyd Rector helped us in many ways and this papercould not have been written without his generous support.Weare also grateful to Dr. Norman Carter who supervisedthe PD measurements between skin and peritoneal cavity.

This work was supported under U. S. Public HealthService Training Grant 5 TO1 AM5490 and Research Grant5 RO1AM06506.

REFERENCES1. Schultz, S. G., and P. F. Curran. 1968. Intestinal ab-

sorption of sodium chloride and water. Handbook ofPhysiology. American Physiological Society, Washing-ton, D. C. 3(Section 6): 1245.

2. Fordtran, J. S., and F. J. Ingelfinger. 1968. Absorptionof water, electrolytes, and sugars from the human gut.Handbook of Physiology. American Physiological So-ciety, Washington, D. C. 3 (Section 6): 1457.

3. Bucher, G. R., J. C. Flynn, and C. S. Robinson. 1944.The action of the human small intestine in altering thecomposition of physiological saline. J. Biol. Chemn. 155:305.

4. Fordtran, J. S., F. C. Rector, and N. W. Carter. 1968.The mechanisms of sodium absorption in the humansmall intestine. J. Clin. Invest. 47: 884.

5. Wright, E. M. 1966. Diffusion potentials across the smallintestine. Nature (London). 212: 189.

6. Gustke, R. F., G. E. Whalen, J. E. Geenen, and K. H.Soergel. 1967. Mucosal potential difference in the intacthuman small intestine. Gastroenterology. 52: 1134.(Abstr.).

7. Dietschy, J. M. 1964. Water and solute movement acrossthe wall of the everted rabbit gallbladder. Gastroenter-ology. 47: 395.

8. Maren, T. M. 1967. Carbonic anhydrase: chemistry,physiology and inhibition. Physiol. Rev. 47: 595.

9. Carter, M. J., and D. S. Parsons. 1968. Carbonic anhy-drase activity of mucosa of small intestine and colon.Nature (London). 219: 176.

10. Kitahara, S., K. R. Fox, and C. A. M. Hogben. 1967.Depression of chloride transport by carbonic anhydraseinhibitors in the absence of carbonic anhydrase. Nature(London). 214: 836.

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