L-glutamine with D-glucose stimulates oxidative metabolism and NaCl absorption in piglet jejunum.

L-Glutamine metabolism

with D-glucose stimulates oxidative and NaCl absorption in piglet jejunum

J. MARC RHOADS, EMMANUEL 0. KEKU, J. PAUL WOODARD, SHRIKANT I. BANGDIWALA, JAMES G. LECCE, AND JOHN T. GATZY Department of Pediatrics and Pharmacology, Schools of Medicine and Public Health, University of North Carolina at Chapel Hill; the North Carolina State University Department of Animal Sciences; and the University of North Carolina Center in Gastrointestinal Biology and Disease, Chapel Hill, North Carolina 27599 Rhoads, 3. Marc, Emmanuel 0. Keku, J. Paul Woodard, Shrikant I. Bangdiwala, James G. Lecce, and John T, Gatzy. L-Glutamine with D-glucosestimulatesoxidative metabolismand NaCl absorption in piglet jejunum. Am. J. Physiol. 263 (Gastrointest. Liver Physiol. 26): G960-G966,1992.

-To explore the relationship between intestinal fluid absorption and oxidative metabolism, we measured the effects of amino acids and glucoseon piglet jejunal ion transport and oxygen consumption(Qo~) in vitro. Jejunal Qo, wasstimulated by L-glutamine and D-glucosebut not by the nonmetabolizable organic solutesmethyl /?-D-glucosideor L-phenylalanine. Qo, was maximally enhancedby the combination of D-glucoseand L-glutamine (5 mM). Even though 5 mM L-glutamine was previously found to be insufficient to stimulate NaCl absorption, 5 mM L-glutamine enhancedjejunal NaCl flux when combined with equimolar mucosalD-glucose.Either D-glucoseor methyl B-D-glucosidecausedan increasein short-circuit current (&), an increasein Na+ absorption in excessof I,0 and a decreasein Cl- secretion, when L-glutamine was substituted for D-glucose (10 mM) on the serosalside. This relationship suggeststhat mucOSal sugars,if combinedwith L-glutamine, enhanceneutral NaCl absorption aswell as electrogenicNa+ flow. (Aminooxy)acetate, an inhibitor of alanine aminotransferase,abolishedthe stimulation of Qo, and the NaCl-absorptive response to L-glutamine. We concludethat the oxidative metabolismfueled by L-glutamineis linked to a NaCl-absorptive mechanismin the intestine. We proposethat the CO* produced by glutamine metabolism yields carbonic acid, which dissociatesto H+ and HCO;, which may stimulate parallel antiports in the apical membrane. sodiumions; (aminooxy)acetate; methyl @-D-glucoside; intestinal metabolism GLUTAMINE IS THE PRIMARY metabolic fuelofthe small bowel (26, 30). Enteral or parenteral administration of glutamine enhances intestinal mucosal regeneration after cytotoxic injury (8, 19). Although L-glutamine is known to be cotransported across the intestinal brush border coupled to Na+ (25,28), the effect of L-glutamine on jejunal ion transport appears to be distinct from that of other amino acids. L-Glutamine stimulated both elec(Na+ absorption associated trogenic Na+ absorption with an increase in short-circuit current) and electrically silent absorption of Na+ (coupled to Cl- absorption) in piglet jejunum (21). Furthermore, L-glutamine stimulated neutral NaCl absorption when added to either the mucosal or serosal side of the tissue (21). We wanted to know whether the stimulation of electroneutral Na+ and Cl- absorption by L-glutamine resulted from a metabolic effect. Piglet intestinal salt absorption was studied because of the similarity of porcine and human intestinal processes. The current studies explored the interaction between L-glutamine and D-gluG960

0193-1857/92

case on piglet jejunal oxidative metabolism and electrolyte transport. They also determined the effects of other amino acids and sugars and recognized inhibitors of glutamine metabolism on these processess. A preliminary account of these results has been presented (22). METHODS Animals

Sixty-seven piglets were obtained from a herd of cross-bred pigs maintained by the North Carolina School of Agricultural and Life Sciences.Enteric infections common to neonatal pigs wereminimized. Five days before the projected farrowing dates, sowswere brought to an intensive care facility and scrubbed daily with a solution of povidone-iodine. The piglets were caught on towels, taken to an isolated facility, placed in individual cages,and fed hourly (300 ml. kg-l *day+) an artificial cow’s milk-based formula with an automatic feeding machine (Autosow; 14). Pigs were monitored daily for the presenceof diarrhea. Stools were periodically tested for the presenceof rotavirus antigen by a latex agglutination test (Virogen Rotatest, Wampole Laboratories, Cranbury, NJ). None of the animalsdevelopeddiarrhea or shed rotavirus. Pigs were studied at l-2 wk of age to allow maturation of intestinal transport (21). Toward the end of the study, there was a shortage of colostrum-deprived pigs, so we continued the study with 11 normally suckled pigs, l-2 wk of age. These piglets appearedhealthy, and none had diarrhea. Jejunal enzyme activities [lactase, Na+-K+-adenosinetriphosphatase (ATPase), seeRef. 201were comparableto those of pigs raised on the Autosow. These studies were approved by the University of North Carolina Animal Care Committee. Chemicals

All unlabeledchemicalswere obtained from Sigma (St. Louis, MO). 22NaC1and Na36Cl were obtained from Amersham (Arlington Heights, IL) and ICN Radiochemicals(Irvine, CA), respectively. Oxygen consumption (&02). Segments of stripped jejunum were incubated in physiological salt solution (see Electrical Measurements) with or without amino acids or sugars(5 mM concn unlessotherwise noted) in water-jacketed Lucite chambers with a polarographic Clarke oxygen electrode. The tissue and buffer solutionswerecontinuously stirred at 37°C and bubbled with 95% air-5% C02. Preliminary studies showedthat tissueQo, wasstable for 15-20 min and then decreasedslowly. Consequently,tissueswere kept in ice-cold physiologicalsaline solution (see EZectricaZ Measurements) and were then transferred to the chamberand stirred to equilibrate temperatureand gases(2-3 min). The chamber was sealed,and Qo, was measured for 3-5 min. Tissueswere in the chambersfor no more than 20 min. The contents of each chamber were dried overnight in an oven at 95°C. The weight of an equalvolumeof dried physiological saline (*organic solute) was subtracted from the

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GLUTAMINE

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JEJUNAL

weight of sampleswith tissue to obtain the tissue dry weight. Results are expressedas microliter of O2 consumedper milligram tissuedry weight per hour. Electrical Measurements Jejuna were prepared as describedpreviously (2‘1). Briefly, proximal jejunum was excised 2-3 cm distal to the suspensory ligament, drawn over a plastic rod, incised longitudinally, and stripped of muscle layers. The stripped tissue was mounted between Lucite half-chambers (aperture = 1.13 cm2) and was bathed at 37°C in 10 ml of gassed(95% 02-5% C02, pH 7.4) Ringer solution with (in mM) 140 Na+, 5.2 K+, 1.2 Ca2+, 1.2 Mg2+ 1198 Cl- 25 HCO;, 0 4 HzPOd, and 2.4 HP04. At the start kf mdst exieriments, &lucose (10 mM) wasaddedto the serosalbath, and equimolar mannitol wasaddedto the mucosal side. The electrical potential difference (PD) acrossthe tissue was measuredby Ringer-agar bridges positioned near the surface of the tissue and connected to calomel electrodesin saturated KCl. Transepithelial PD was maintained at zero by the short-circuit current (I,) generated by automatic voltage clamps (World Precision Instruments, New Haven, CT). Tissueswere mounted in the chamberswithin 30 min of the death of the piglet and continuously short-circuited, except for 5-s intervals every lo-20 min when open-circuit PD was measured. Conductancewascalculatedby Ohm’slaw from the open-circuit PD and IBc. Flux Studies Tissueswere paired so that conductance did not differ by ~25%. Fifteen to 20 min after the tissue was mounted, both **NaCl (2 &i) and Na3V1 (2 &i) were added to either the mucosalor serosalbath. After 20 min, unidirectional Na+ and Cl- fluxes werecalculatedfrom the rate of isotopeappearancein 1.0.ml samplesdrawn from the bath every 20 min. After 40 min, L-glutamine, D-glucose,or both wereaddedto the mucosalbath. Equimolar mannitol wasaddedto the serosalside.Twenty minutes later, we remeasuredunidirectional ion fluxes.for a subsequent 40-min period. Fluxes from mucosato serosa(Jm+S) and from serosato mucosa (Js-m) were determined from paired tissues,and Jnet was taken as the difference between the two fluxes. Occasionally, the conductance acrosstissues began to increaseafter 80-100 min, accompaniedby an increasein unidirectional Na+ and Cl- fluxes. To minimize the chanceof such a changein the presenceof metabolic inhibition, flux and equilibration periods were shortened to 10 and 15 min, respectively, and the average of three or four flux periods during the experimental period was recorded. Inhibitors (Aminooxy)acetate is a competitive inhibitor of alanine aminotransferase,which prevents glutamine from being converted to ar-ketoglutarate,a substrate that enters the Krebs cycle (10, 11, 15). Another inhibitor of glutamine metabolism,the structural analogue6-diazo-5-oxonorleucine,irreversibly inhibits (in rat intestine) the first and rate-limiting enzyme in mitochondrial glutamine metabolism,phosphate-dependentglutaminase (30). The effects of these two inhibitors on electrolyte absorption in the absenceand presenceof L-glutamine were examined at the concentrations that were reported to maximally inhibit glutamine metabolismby rat intestine (10 and 2.5 mM, respectively). Inhibitors were addedto both baths at t = 0 (i.e., 40 min before L-glutamine was added).

METABOLISM

G961

AND ABSORPTION

require that the data have a normal distribution. Data are summarized as meanst SE RESULTS

Jejunal Qo2 Figure 1 shows Qo, of stripped piglet jejunum bathed in Ringer b uffer and exposed sequentially at 7-min intervals to increasing concentrations of L-glutamine. Mannito1 (an unabsorbed nonmetabolizable solute) was added at the same concentrations to the chamber with the control segment. At 0.5 mM concentration, mannitol and glutamine tended to enhance jejunal QoZ slightly but not significantly. L-glutamine stimulated Qo, maximally at 5 mM (41 t 16% increase). Higher concentrations of glutamine did not affect Qo, significantly. Smaller Qo, values in the presence of higher concentrations of glutamine could indicate spontaneous waning of tissue respiration after 20 min in vitro or toxic effects of supraphysiological glutamine. A toxic effect seems unlikely, given our finding that Qo~ of tissue immediately placed in 30 mM L-glutamine was 11.6 -+ 1.8 &mg-l*h-l (n = 5), similar to that of tissue immediately placed in 5 mM L-glutamine [13.1 t 0.8 &rng-l l h-l (n = 12)]. Based on these findings, we subsequently measured &o, once in paired strips of tissue added to buffer alone or buffer containing optimal L-glutamine, D-glucose, or other sugars and amino acids. Qo, was higher in tissues exposed to 5 mM L-glutamine (41% increment) or D-glucose (46% increment; Fig. 2). Qo, was 120% greater in tissues exposed to the combination of L-glutamine and D-glucose. The effect of glutamine plus glucose on Qo, was not significantly greater than the sum of the iesponses to the individual substrates. &a was no greater when tissues were exposed to 30 mM L-glutamine $us 30 mM D-glucose (not shown). When an inhibitor of glutamine metabolism, (aminooxy)acetate (10 mM), was present in the bath (with glutamine), Qo, was equivalent to Qo, of tissues not exposed to glutamine (Fig. 2). The structural analogue of D-glucose, methyl /3-D-ghcoside (5 mM), which is cotransported with Na+ across the apical membrane, did not stimulate Qo, (Fig. 2). When 30 mM methyl /3-D-glucoside was added to maximize sugar-coupled Na+ absorption (24), jejunal Qo2 (9.1 9

24

1

0 0

-O-

I

I

5

10

Glutamine

1 15

1

20

Substrate concentration (mM) Fig. 1. Oxygen consumption (Qo~, &rng-l l rein-l) of stripped piglet Statistics jejunum in vitro. Response to sequential addition (%min time intervals) Differences between mean fluxes of matched pairs were as- of increasing concentrations (0.5-20 mM) of L-glutamine or mannitol. sessedby the Wilcoxon signedranks test. This test was chosen n = 6 tissues from 5 pigs for glutamine response; n = 4 tissues from 3 becausesamplesizeswere relatively small,and this test doesnot pigs for mannitol response. Values are means2 SE.


G962

GLUTAMINE

ENHANCES

JEJUNAL

METABOLISM

AND ABSORPTION

+

25

Buffer

Ringer

Gin

Gln/AOA

Glc

f3MC

Gln/Glc

(12)

(12)

(8)

(7)

(11)

(7)

Fig. 2. Jejunal Qo, in paired tissue segments from normal piglets l- to 2-wk old placed directly in buffer containing 5 mM L-glutamine (Gln), D-glucose (Glc), or methyl @-D-glucopyranoside (BMG). (Aminooxy) acetate (AOA), where noted, is present at 10 mM. Number of piglets studied is in parentheses. Selection of tissues to be exposed to various substrates was done in a random order. There were no sequential additions. Although sample sizes are different, tissues were paired and each of the Wilcoxon tests was performed only on the relevant matched pairs. *P < 0.05,** P < 0.01 compared with Ringer buffer alone. + P < 0.05 compared with tissue exposed to glutamine or glucose.

&mg-l~h-l, n = 2) was similar to the rate in tissues exposed to the lower concentration. Table 1 shows jejunal Qo, response to sequential addition of 5 mM L-phenylalanine (an amino acid cotransported with Na+; 21) or mannitol, and methyl p-D-&coside. None of these three substrates individually stimulated oxidative metabolism. However, methyl P-Dglucoside, when added after glutamine, significantly augmented Qo2. Glucose, added after glutamine, caused an even greater stimulation of Qo2. Figure 3 shows Qo2 of jejunal strips incubated in a buffer in which Na+ was replaced by equimolar ALmethyl-D-glucamine. Na+ replacement resulted in a 60% reduction in tissue respiration. Na+ replacement also prevented the Qo, responses to glutamine, glucose, or the combination of the two. Table 1. Response of jejunal Original

Solution

Qo2 to sequential

addition

+

Phenylalanine 11.9k2.6

Ringer 14.8k2.8 (4

4

Mannitol(5 15.921.5

Glutamine (5 mM) 20.0*2.2 (5)

--b

Cln/Glc

Glc

Ion Transport Studies Effect of mucosal L-glutamine and o-glucose on ion transport. L-Glutamine, added to the serosal reservoir at

“serum concentration” = 0.5 mM did not enhance Na+ or Cl- absorption (n = 3 piglets). We tested the effect of the nutrient concentrations that maximally stimulated Qo, (5 mM) on Na+ and Cl- fluxes. These concentrations were expected to minimally affect NaCl absorption, because previous studies had shown that 5 mM mucosal L-glutamine did not affect absorption of either Na+ or Cl- (21). When 5 mM L-glutamine plus 5 mM D-glucose were added to the mucosal bath, e; increased more than ISc (2.1 t 0.5 compared with 1.1 t 0.6 peq*cm-2*h-1, respectively, P < 0.055 by signed ranks test). At the same time, JE+S increased and Cl- secretion was reduced (AcAt = 0.8 t 0.3 peg* crnB2. h-l, P c 0.05). These observations are summarized in Fig. 4. Tissue conductance did not significantly change after glucose and glutamine were added (not shown). Effect of serosal L-glutamine (10 mM) on Na+ absorption stimulated by o-glucose (30 mM). To determine the effect

of L-glutamine on D-glucose-stimulated Na+ flow, we studied the response of jejunum to mucosal D-glucose in tissues exposed to serosal glutamine in place of glucose (Table 2). Resting net flux of Na+ did not significantly

of amino acids and methyl P-o-glucoside

First Additions

Ringer 12.Ok2.8 (4)

Glutamine (5 mM) 17.2k2.7 (10)

Gln

Fig. 3. Qo, rate of piglet jejunum bathed in a buffer in which NaCl was replaced by equimolar N-methyl-D-glucamine chloride and NaHC03 was replaced by equimolar choline bicarbonate. Note that ordinate is expanded compared with Fig. 1. Tissues from 3 different piglets were studied in parallel under each of the test conditions. See Fig. 1 legend for additional information.

Second Additions

(5 mM)

mM)

-+

--+

Methyl p-D-glucoside (5 mM) 10.6t3.0 Methyl p-D-glucoside (5 mM) 13.lk1.8

Methyl @-D-glucoside (5 mM) 19.8k2.9” Glucose (5 mM) 29.6t4.5’

--+ Glutamine (5 mM) Glutamine (5 mM) 21.3t6.6 18.7k5.8 (4) Values are means ,t SE; (n) = no. of piglet jejunal segments. Shown are rates of oxygen consumption (Qo~; in ~1. mg-l h-l). * P < 0.05 by signed ranks test. l


GLUTAMINE

ENHANCES

JEJUNAL

METABOLISM

-5

-

J”, Fig. 4. Unidirectional and net Na+ and Cl- fluxes (ez and cAt, respectively) across piglet jejunum. Black bars indicate basal fluxes. Striped bars are fluxes in presence of mucosal Glc (5 mM) and Gln (5 mM). Mannitol(l0 mM) was added to the serosal bath. Values are means t, SE; n = 7 tissue pairs from 7 pigs. J!$ Na flux from mucosa to serosa; J’!$ Na flux from serosa to mucosa; PA, Cl flux from mucosa to serosa; fl:, Cl flux from serosa to mucosa; Jnet = Jms - J,,; Jr$ residual flux; 1=, short-circuit current. * P < 0.05, ** P < 0.01 compared with basal flux.

differ from zero, but Cl- was secreted. D-ghCW! enhanced Na+ absorption. Part of the increase was electrogenie (AIBc = 1.7 peqcmB2. h-l). Net Cl- secretion decreased by 1.0 & 0.3 peq cm -2. h-l, a change that resulted from a significant increase in Jc,’ S. Effect of inhibitors of glutami& metabolism on NaCl absorptioe reSpOltSe to L-glutamine. Electrolyte transport by tissues pretreated with 10 mM (aminooxy)acetate and then exposed to 20 mM mucosal L-glutamine were determined (Fig. 5). These concentrations were chosen because 20 mM L-glutamine enhances Jm+ and net absorption of Na+ and Cl- (21) and because increasing the bath osmolarity by 30 mM (with mannitol) does not influence Na+ or Cl- transport rates (21). (Aminooxy)acetate did not significantly affect basal jejunal ion flows compared with flows under control conditions. (Aminooxy)acetate abolished the NaCl-absorptive transport response to mucosal glutamine (Fig. 5). However, tissue preincubated with (aminooxy)acetate responded to glutamine with the expected increase in IBc (A.&,, = 0.9 k 0.3 peqe crnw2 h-l, P < 0.05, Fig. 6). Tissue conductance, remained stable throughout the experiment. With (aminooxy)acetate in the bath, 30 mM mucosal D-glucose (added 60 min after glutamine) caused an increase in I,, = 0.7 t 0.2 ~eqcrn-~. h-l (P < 0.05, n = 8 tissue segments from 4

G963

AND ABSORPTION

J”,

J”,,

J=‘,

J=‘,

J=’

net

JR

Fig. 5. Unidirectional and net ion flows in pig jejunum under basal conditions in the presence of 10 mM (aminooxy)acetate (black bars) and after 20 mM mucosal L-glutamine has been added (striped bars). n = 6 pigs. Flux rates are not significantly different in the presence and absence of glutamine.

l

l

Pigs).

The L-glutamine analogue 6-diazo-5-oxonorleucine did not affect glutamine’s stimulatory effect on Qo2 (not shown). 6-Diazo-5-oxonorleucine also did not inhibit the transport response to mucosal L-glutamine. Responses to Table 2. Piglet jejunal eJNa m-4

transport

20 I 20

O0

JNa net

JELI

1 60

4 80

t 100

TIME (min.)

Fig. 6. 1= response of jejunal sheets in Ussing chambers before (t = 0 min) and after (t = -40-160 min) mucosal addition of 5 mM L-glutamine and 5 mM D-&cost? (open triangles); 30 mM methyl B-Dglucoside with serosal glutamine (open circles); 30 n&l methyl /?-Da glucoside with serosal glucose (closed circles); or 20 mM L-glutamine in the presence of 10 mM (aminooxy)acetate (closed t&n&s). 1= increased (P < 0.05) at t = 60 min in each of the 4 instances. In experiments with glucose or methyl /3-D-glucoside, 1= was determined at 200 min intervals for 100 min. In experiments with the inhibitor (aminooxy)acetate, flux periods were 10 min, and the total study period was 80 min. For the purposes of graphic comparison, the time scale was adjusted so that addition of glutamine would coincide with t = 40 min; Iw values at 20-min intervals are shown. Values are means * SE; n = 6 or 7 piglets for each study. When not shown, SE was smaller than size of symbol.

mucosal L-glutamine (30 mM) in tissues bathed by 2.5 mM 6-diazo-5-oxonorleucine were AJ!$ = 2.8 k 1.0; APnet1 = 1.3 t 0.5; A&= = 1.0 k 0.3 peqecm-2.h-1 (P < 0.05, n = 6). Effect of a rwnmetabolizable sugar on Na+ and Cltransport in the presence of glutamine. Figure 7 shows the effect on jejunal electrolyte transport of mucosal methyl @-D-glucoside. We compared the response to methyl B-Dglucoside in tissues exposed to serosal D-glucose (10 mM)

response to mucosal o-glucose (30 mM) with se&d

JNa km

1 40

Jz!ln

JEt

L-gUarnine (IO mM) J!L

LC

G

Basal 5.3*0.7 6.0*0.6 -0.720.7 4.OkO.4 6.6k0.5 -2.6k0.7 -0.2kO.4 1.7*0.3 14Jkto.9 D-Glucose 9.2a.4 7.3*0.9 2.0k0.8 6.1kO.8 7.7kO.9 -1.6k0.7 -0.kto.5 3.4*0.9 l&1*2.2 A 3.9a.1 1.320.6 2.6k0.7 2.1H.O l.lkO.6 l.OkO.3 0. MO.2 1.7kO.6 3.3k2.1 P co.05 NS 5 mM) when added in the presence of mucosal or serosal glucose stimulated all three processes. These findings support those of Frizzell et al. (9) who showed that neither glucose nor proline increased Qo, by rabbit ileum, even though both are transported with Na+ across the apical membrane (27). In contrast, they found that Qo, was stimulated by glutamine, alanine, and glutamate, three metabolizable substrates (9). Oxidation of glutamine appears to be necessary for the observed increases in jejunal Na+ and Cl- absorption. (Aminooxy)acetate inhibits the second step in glutamine catabolism, the conversion of glutamate to a-ketoglutarate, catalyzed by either of the two enzymes alanine aminotransferase or glutamate dehydrogenase. In suck10 mM (aminooxy)acetate reduces ling rat jejunum, glutamine oxidation by 95% (1 l), suggesting that the predominant metabolic pathway is through alanine aminotransferase. We, too, found that in newborn animals (aminooxy)acetate abolished the increase in Qo,, as well as the neutral NaCl-absorptive responses to L-glutamine. Although a nonspecific inhibitory effect of (aminooxy)acetate on electrolyte transport cannot be ruled out, the expected electrical response to L-glutamine in the presence of the inhibitor (Fig. 6) suggests that apical L-glutamine-Na+ cotransport is unaffected. Furthermore, the increase in I,, after mucosal D-ghcose (added after 2 h incubation) ruled against a general toxic effect. Another inhibitor of glutamine metabolism, 6-diazo-&oxonorleutine, did not inhibit glutamine’s stimulation of neutral NaCl absorption. We postulate that this inhibitor either failed to interact with porcine glutaminase at the concentration tested or it was degraded by the tissue. Weber et al. (29) showed that intraluminal glucose enhanced glutamine uptake and catabolism in canine jejunum. The nonmetabolizable hexose 3-0-methylglucose enhanced fluid absorption but not glutamine metabolism. They concluded that glucose promotes glutamine oxida-

tion directly, independent of its effect on Na+ absorption. In our studies, glucose added after glutamine appeared to have an additive effect on metabolic rate (Fig. 2), but without measuring release of NH3 from glutamine, we cannot conclude that glucose accelerated glutamine metabolism. Like Weber et al. (29), we observed similar effects of glucose and a nonmetabolizable analogue on intestinal water/electrolyte absorption (Fig. 6) when the tissue was provided with glutamine as a fuel. In the absence of glutamine, methyl p-D-glucoside had a minor effect on Na+ absorption, whereas D-glucose was an effective stimulator. Model for L-Glutamine-stimulated and NaCl absorption

Na+

Although we have not identified the mechanism by which glutamine activates NaCl transport, we propose a model on which further experimentation can be based. Parallel Na+-H+ and Cl--HCO; antiports have been described in pig jejunum and rabbit ileum (7,13). A coupled NaCl absorptive process has not been identified in pig jejunal brush border (7). Mitochondrial oxidation of glutamine to CO2 must yield intracellular H+ through the formation of carbonic acid. which dissociates to H+ and HCO,. Therefore, glutamine oxidation produces both ionic species that are transported by the parallel antiport systems. When a sugar is present that enters by apical Na+-coupled transport, one would expect increased activity of the Na+-K+-ATPase to extrude the additional intracellular Na+. The increased “pump” activity could, in theory, enhance glutamine metabolism, H+ and HCO, generation, and transepithelial NaCl absorption. In addition to acid production, glutamine metabolism would be expected to generate organic anions. In rabbit kidney there are apical antiport systems that exchange Cl- for several of the anionic products of glutamine metabolism, including lactate, oxaloacetate, and a-ketogutarate (1). Implications for Oral Rehydration Therapy

A research priority for the World Health Organization is to improve the glucose-based ORS used to treat patients with diarrhea1 dehydration. Oral treatment solutions that contain glucose plus amino acids effectively replace fluid deficits and reduce stool output of patients


G966

GLUTAMINE

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JEJUNAL

with secretory diarrhea (20), by stimulating absorption across separate Na+-coupled carriers in the apical membrane (19). Glutamine, the most abundant circulating amino acid in humans, has been considered a promising component of “super ORS.” When infused enterally in volunteers, L-glutamine is absorbed efficiently (6). By serving as a fuel, glutamine reduced the rate of lipolysis in these patients (6). L-Glutamine stimulates both neutral NaCl absorption and Na+ absorption in the intestine, even in rotavirus-damaged epithelium (23). However, because of the high concentrations required to enhance absorption, extrapolation of our findings to ORS may be limited. The efficacies of L-glutamine-based and glucosebased ORS in dehydrated patients are presently being compared (0. Fontaine, personal communication; Ref. 5). The authors thank Drs. Don Powell and Arthur Finn for review of the manuscript, R. John MacLeod and Dr. Douglas W. Wilmore for helpful suggestions, Shih-Chia Chang-Liu for assistance in statistical analysis, and William Dorsey and Jan Huggins for expert animal care. This study was supported by grants from the World Health Organization (HQ/88/106496) and the National Institute of Diabetes and Digestive and Kidney Diseases (DK-K08-HD00945). Address for reprint requests: M. Rhoads, Dept. of Pediatrics, Div. of Gastroenterology, Univ. of North Carolina at Chapel Hill, Campus Box 7220, Chapel Hill, NC 27599. Received 21 October 1991; accepted in final form 3 August 1992. 1. Aronson, P. S. The renal proximal tubule: a model for diversity of anion exchangers and stilbene-sensitive anion transporters. Annu. Rev. Physiol. 51: 419-441, 1989. 2. Bern, M. J., C. W. Sturbaum, S. S. Karayalcin, H. M. Berschneider, 3. T. Wachsman, and D. W. Powell. Immune system control of rat and rabbit colonic electrolyte transport: role of prostaglandins and enteric nervous system. J. Clin. Invest. 83: 1810-1820, 1989. 3. Bessman, S. P., and E, C. Layne. The stimulation of the non-enzymatic decarboxylation of oxalacetic acid by amino acids. Arch. Biochem. 26: 25-32, 1950. 4. Dahlqvist, A. Method for assay of intestinal disaccharidases. Anal. Biochem. 7: 18-25, 1964. 5. Dechelotte B., G. A&an, B. Hecketsweiler, and P. Hecketsweiler.*Effect of glutamine and alanine on water and electrolyte fluxes in human jejunum during experimental hypersecretion (Abstract). Gastroenterology 100: A683, 1991. 6. Dechelotte, P., D. Darmaun, M. Rongier, B. Hecketsweiler, 0. Rigal, and J.-F. Desjeux. Absorption and metabolic effects of enterally administered glutamine in humans. Am. J. Physiol. 260 (Gastrointest. Liver Physiol. 23): G677-G682,1991. 7. Forsyth, G. W., and S. E. Gabriel. Chloride ion transport into pig jejunal brush-border membrane vesicles. J. Physiol. Lord. 402: 555-564, 1988. 8. Fox, A. D., S. A. Kripke, J. De Paula, 3. M. Berman, R. G. Settle, and J. L. Rombeau. Effect of a glutamine-supplemented enteral diet on methotrexate-induced enterocolitis. J. Parenter. Enteral Nutr. 12: 325-31, 1988. 9. Frizzell, R. A., L. Markscheid-Kaspi, and S. G. Schultz. Oxidative metabolism of rabbit ileal mucosa. Am. J. Physiol. 226: 1142-1148, 1974. 10. John, R. A., A. Charteris, and L. J. Fowler. The reaction of amino-oxyacetate with pyridoxal phosphate-dependent enzymes. Biochem. J. 171: 771-779,1978.

METABOLISM

AND ABSORPTION

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Acetate absorption in the normal and secreting rat jejunum.

HCO3 transport in rat jejunum: relationship to NaCl and H2O transport in vivo.

Renojejunal reflex controlling jejunal absorption of fluid and NaCl.

Insulin stimulates SGLT2-mediated tubular glucose absorption via oxidative stress generation.

Thiazide-sensitive NaCl absorption in rat cortical collecting duct.

Absorption of lactose from colon of newborn piglet.

Lipid absorption and metabolism.

Absorption and metabolism of piceatannol in rats.

Unstirred layer and kinetics of electrogenic glucose absorption in the human jejunum in situ.

Oxidative stress, antioxidants and intestinal calcium absorption.

Water and solute absorption from hypotonic glucose-electrolyte solutions in human jejunum.

Stimulation of active and passive sodium absorption by sugars in the human jejunum.

Genistein stimulates jejunum chloride secretion via an Akt-mediated pathway in intact female mice.

Oxidative metabolism of dimethylnitrosamine: correlation with toxicity.

Intravenous injection of hypertonic NaCl solution stimulates pulmonary C-fibers in dogs.

Percutaneous absorption and metabolism of Irgasan DP300.

Absorption, metabolism, and excretion of hydrochlorothiazide.

Absorption, metabolism, and functions of β-cryptoxanthin.

Absorption and metabolism of nicotine from cigarettes.

Vitamin D metabolism and calcium absorption.

Intestinal absorption and metabolism of xenobiotics.

Absorption, transport and metabolism of vitamin E.

Absorption from the jejunum is increased by forward and backward pacing.

Further studies on the action of prolactin on fluid and ion absorption by the rat jejunum.