Effect of Bicarbonate on Efficacy of Oral Rehydration Therapy: Studies in an Experimental Model of Secretory Diarrhoea

Gut: first published as 10.1136/gut.29.8.1052 on 1 August 1988. Downloaded from

Gut, 1988, 29, 1052-1057

Effect of bicarbonate on efficacy of oral rehydration therapy: studies in an experimental model of secretory diarrhoea

E J ELLIOTT, A J M WATSON, J A WALKER-SMITH, AND M J G FARTHING From the Departments ofGastroenterology and Child Health, St Bartholomew's Hospital, London

SUMMARY In situ perfusion of rat intestine was used to evaluate the effect of bicarbonate on the efficacy of a low sodium (35 mmol/l) glucose-electrolyte oral rehydration solution in normal and cholera toxin-treated rat small intestine. In normal intestine, absorption of water was greater (108 (8-1) .l/min/g; p<001) and sodium secretion less (-4-3 (0.3) [tmol/min/g; p<001) from the oral rehydration solution containing bicarbonate than from the solution in which bicarbonate was replaced by chloride ions (59 5 (7.2) ,tl/min/g and -7-8 (0.8) ,umol/min/g, respectively). Glucose absorption in normal intestine was similar with both solutions. In the secreting intestine, both oral rehydration solutions reversed net water secretion to absorption, but inclusion of bicarbonate resulted in significantly less net absorption of both water (2 18 (6.9) [tl/min/g; p<0-05) and glucose (18.7 (2.1) [tmol/min/g; p<0-001) compared with bicarbonate free oral rehydration solution (19.4 (3.9) ,tl/min/g and 35-8 (3.7) [mol/min/g, respectively). Net sodium secretion occurred in normal and secreting intestine but was significantly less with the bicarbonate containing oral rehydration solution. These findings suggest that the demonstrable advantage ofbicarbonate in promoting water absorption from this oral rehydration solution in normal rat intestine does not apply to cholera toxin http://gut.bmj.com/ treated secreting intestine. on September 25, 2021 by guest. Protected copyright. Glucose-electrolyte oral rehydration solutions bicarbonate or the base precursors have similar (ORSs) are effective in the treatment of dehydration effects on water and sodium absorption in the secret- caused by acute diarrhoeal illnesses including ing intestine. cholera,' and their widespread use has contributed Recent clinical studies suggest that bicarbonate significantly to the decrease in morbidity and free ORSs are effective in correcting the dehydration mortality associated with these infections.2 Contro- and acidosis associated with acute gastroenteritis."- versy still exists regarding the value of an individual Rehydration alone allows the kidney to correct base or base precursor in ORSs. Historically, bicar- acidosis in all but the most severe cases, when bonate or a base precursor (lactate, acetate, citrate) exogenous base may be required. 12 In addition, there has been included in ORSs on the assumption that it are practical difficulties associated with the use of is required for correction of the metabolic acidosis bicarbonate. It is often unobtainable in developing associated with acute gastroenteritis.3 Their inclusion countries, adds bulk and expense to prepackaged has been further justified by the observation that oral rehydration powders and may discolour solu- bicarbonate, acetate and citrate promote water and tions containing glucose by the formation of furfural sodium absorption in the normal human`7 and rat8 compounds.'3 Thus there would be economic and small intestine. There is no evidence, however, that production advantages if bicarbonate could be omitted. For these reasons the World Health Address for correspondence: Dr M J G Farthing, Department of Gastro- enterology, St Bartholomew's Hospital, West Smithfield, London ECIA 7BE, Organisation (WHO) has recently substituted citrate UK. for bicarbonate in their ORS'4 athough its advantage Received for publication 23 February 1988. over a similar solution without citrate has not been 1052 Gut: first published as 10.1136/gut.29.8.1052 on 1 August 1988. Downloaded from

Effect ofbicarbonate on efficacy oforal rehydration therapy 1053 substantiated. Despite this, ORSs containing bicar- different perfusion solutions were always made bonate are still widely used in the United Kingdom during the same time period after exposure of the and elsewhere. intestine to cholera toxin.`1 The purpose of this study was to evaluate the effect Seventy two rats were studied. Three groups of 12 of bicarbonate on the ability of the most commonly rats received either 150 mmol/l sodium chloride, or used ORS in the United Kingdom to promote water the ORS with bicarbonate, or the ORS without and solute absorption. To investigate this we have bicarbonate after two hour exposure to cholera toxin undertaken in situ perfusion of the entire rat small saline. Another three groups of 12 rats were not intestine excluding the duodenum using methods exposed to cholera toxin, but were perfused with the described previously."5 Studies were done in normal same three solutions. Only one solution was studied intestine and in intestine after induction of a secre- in each animal to ensure the small intestine remained tory state with cholera toxin. functional throughout the perfusion, which had a total duration of 1-5 h. Our previous study using this Methods method confirmed satisfactory absorptive function after three hour perfusion.'5 IN SITU STEADY-STATE PERFUSION OF RAT SMALL INTESTINE TEST SOLUTIONS Male Wistar rats of 180-240 g in weight were fasted The ORS containing bicarbonate (HCO3 18, Na 35, for 18 hours with free access to water, then anaesthe- Cl 37, K 20, glucose 202 mmol/l; 312 mOsm/kg; pH tised with intraperitoneal sodium pentobarbitone (60 8-15) was compared with an ORS in which bicar- mg/kg) and maintained with 30 mg/kg im as required. bonate was replaced by chloride ions (Na 35, Cl 55, At laparotomy cannulae were inserted into the K 20, glucose 202 mmolI1; 312 mOsm/kg; pH 5.89). proximal jejunum and terminal ileum. The proximal Polyethylene glycol (PEG; MW 4000) was added as a cannula was used to deliver the test solution and the non-absorbable marker (5 iCi/l ['4C]-PEG with 2-5 distal cannula to collect the effluent. Body tempera- g/1 unlabelled PEG).'6 The pH of the ORSs was not ture was maintained at 37±0 5°C by a heating pad adjusted to the physiological range for the small and overhead lamp. Before perfusion residual small intestine. intestinal contents were removed by gentle lavage This particular ORS was chosen as it has been with isotonic saline at 37°C. recommended in the British National Formulary http://gut.bmj.com/ In all experiments the entire small intestine apart since the mid-1970s,'7 is commercially available, and from the duodenum, to exclude pancreatic secre- is currently the ORS most widely used in the United tions, was perfused using a Braun ED2 pump at a rate Kingdom. It has a lower osmolality, contains more of 0-5 ml/min." After an equilibration period of 60 glucose and less sodium, chloride and bicarbonate minutes with the test solution, three successive 10 than the solution recommended by the WHO.'3 The minute collections were made by siphonage from 150 mmol/l sodium chloride solution was rendered

each rat via the distal cannula. The variation in isotonic to rat serum osmolality (308 mOsm/kg) by on September 25, 2021 by guest. Protected copyright. polyethylene glycol (PEG) concentration was <10% addition of 25 mmol/l mannitol. All chemicals used about the mean in the three successive collections, were AnalaR grade from British Drug House showing that a steady state had been achieved. At the Chemicals Ltd, Poole, England. end of the experiment rats were killed and the perfused segment of small intestine removed, dried ANALYTICAL METHODS AND CALCULATIONS for 12 hours in a hot air oven at 100°C and weighed. Bicarbonate concentration of the effluents was A secretory state was induced in the intestine of measured immediately as total CO2 using a Corning half the rats before perfusion by instillation of 75 [ig 965 carbon dioxide analyser. Glucose content was pure cholera toxin (Sigma Chemical Co, Poole, estimated by the glucose oxidase method using a Dorset, product no. C-9025, lot 14F-4054) in 5 ml 150 Technicon Autoanalyzer. Sodium and potassium mmol/l sodium chloride into the intestine via the were determined by flame emission spectroscopy distal cannula, which was then ilamped.'5 Cholera (Intrumentation Laboratory 943), osmolality by toxin saline was distributed throughout the small freezing point depression and chloride by coulomb- intestine and the abdomen closed for two hours. The metric titration (CCMI chloride meter). ['4C]-PEG clamp was then removed and the intestine drained. was measured by liquid scintillation spectroscopy Induction of a secretory state was reliably and using an LKB 1210 Ultrobeta counter. pH was reproducibly achieved and confirmed in this model measured immediately using a PTI-6 Universal by determination of water and sodium secretion digital pH meter. during perfusion with isotonic saline before and after Three consecutive 10 minute collections were exposure to cholera toxin. Comparisons between the made from each rat and were analysed. The mean of Gut: first published as 10.1136/gut.29.8.1052 on 1 August 1988. Downloaded from

'1054 Elliott, Watson, Walker-Smith, and Farthing these results was used to calculate net movement of Table l Effect ofbicarbonate in ORS on solute movement solute. Water movement was determined from the in normal andsecreting rat small intestine ratio of PEG concentrations in the perfusate and effluents.'` Net absorption (+) indicates movement Netsmall Normalsmall intestine Secreting small intestine inttestinal of water or solute out of the lumen; net secretion (-) transport ORS without ORS with ORS without ORS with indicates movement into the lumen. Polyethylene bicarbonate bicarbonate bicarbonate bicarbonate glycol recovery was 100.3 (3) % (mean (SE); n= 100). Differences in water and solute fluxes were examined Sodium -7-8(0(8) -4-3 (0.9)* -15.8 (1.2) -9.7 (1 .4)t Chloride +0(3 (1) -46 (1 )t -68 (0.9) -6-0 (1.3)t statistically using the non-parametric Mann-Whitney Potassium +4.9 (0.3) +7.0 (0.3)* +3.9 (0.3) +4-2 (0.6)t U test for unpaired data. Bicarbonate -2-0(0-1) +5-8 (0.4)* -3-8 (0.8) +0-5 (0(5)*

Results Net movement of water is expressed as dI/min/g dry weight of small intestine and of solutes as ftmol/min/g. +denotes absorption, WATER AND SOLUTE MOVEMENT DURING -denotes secretion. Results are the mean (SE) (n= 12). The level of significance of the differences between movement during perfusion PERFUSION WITH ISOTONIC SODIUM CHLORIDE with ORS with and without bicarbonate are shown by: *p<0-0l; Minimal net water absorption (+1 38 (5.0) [tl/min/g tp<(-(X)l; tNS. dry wt) and net sodium absorption (+5.24 (0.8) Rmol/min/g) occurred during perfusion of normal small intestine (n= 12) with isotonic 150 mmol/l free glucose saline ORS resulted in net water absorp- sodium chloride (Fig. 1). Perfusion with the same tion (Fig. 1). This almost doubled with the addition of solution after exposure to cholera toxin (n= 12) 18 mmol/l of bicarbonate to the solution (p<0-01). In resulted in net secretion of water (-47.8 (11.8) the secreting intestine, net water secretion was [tl/min/g; p<0001) and sodium (-3.7 (2.1) fmol/ reversed to net absorption by bicarbonate free min/g; p<0.001) and decreased absorption of glucose saline ORS (Fig. 1), but net absorption was chloride (+6.6 (.11) v +14 (1.7) [tmol/min/g; less than with the same solution in the normal p<001) compared with normal intestine (Fig. 1), intestine (p<0.001). In contrast with the normal confirming induction of a secretory state. intestine, the addition of bicarbonate to the glucose saline ORS did not enhance water absorption, but EFFECT OF BICARBONATE ON WATER resulted in a 10-fold reduction in water absorption http://gut.bmj.com/ MOVEMENT (p<005, Fig. 1). In the normal intestine, perfusion with bicarbonate EFFECT OF BICARBONATE ON SOLUTE -pc

secretion in normal and secreting intestine despite on September 25, 2021 by guest. Protected copyright. net water absorption. The addition of bicarbonate to ' Isotonic saline the ORS significantly reduced the degree of sodium secretion in the normal intestine (p<0.01) and secret- '0 g ORS without bicarbonate a ing intestine (p<0-001), although net sodium absorp- [_] ORS with bicarbonate tion never occurred (Table). r-p<005-1 Chloride movement E In the normal intestine net chloride absorption -0QI occurred from the bicarbonate free ORS and secre- t tion from the ORS containing bicarbonate 0 (p<0001). Secretion of chloride by the secreting intestine was similar with the bicarbonate free and bicarbonate containing solutions (Table). Potassium movement Although net potassium absorption was less from the Normal Secreting bicarbonate free than from the bicarbonate contain- intestine intestine ing ORS in the normal intestine (p<0-01), absorp- Fig. 1 Effect ofbicarbonate on water movement in normal tion was similar from both solutions in the secreting and secreting rat small intestine. intestine (Table). Gut: first published as 10.1136/gut.29.8.1052 on 1 August 1988. Downloaded from

Effect ofbicarbonate on efficacy oforal rehydration therapy 1055

(2) and 290-5 (2) mOsm/kg in those perfused with ORS with and without bicarbonate, respectively. In | 2ORS withoubicorbonoktl secreting intestine, values were 292 (2.7) and 305.8 with bicarbonate (2.4) mOsm/kg in those perfused with ORS with and to r-p<0-00l-n without bicarbonate, respectively. i4 c to Discussion c Oc0 .o .c-~> Perfusion of normal rat intestine has been used previously to evaluate the potential efficacy of Z5o CO.~ established and experimental ORS."1`21 We have o cE developed this rat model of intestinal secretion as we '1 cohsidered it might reflect more closely the patho- 12 physiology of acute diarrhoeal disease, particularly 2 that due to enterotoxin.'` The validity of using an a animal model should always be questioned but our Norma l Secreting intestine intestine findings on the pattern of water absorption in normal Fig. 2 Effect ofbicarbonate on glucose absorption in intestine in our model confirm previous observations normal and secreting rat small intestine. in man that: (i) little or no net jejunal sodium or water absorption occurs from an isotonic sodium chloride solution,'8 (ii) the addition of glucose to an Bicarbonate movement isotonic saline solution enhances water absorption,'8 In the normal and secreting intestines, net bicarbon- and (iii) the addition of bicarbonate to a saline ate secretion occurred during perfusion of the solution enhances small intestinal water and sodium bicarbonate free ORS, and absorption of bicarbon- absorption.'r We have also recently shown that there ate occurred from the ORS containing bicarbonate. is close parallelism between the way in which normal Bicarbonate absorption from ORS containing bi- rat and human small intestine handle a variety of carbonate was severely impaired in the secreting ORSs (unpublished observations), although com- intestine, being 10-fold less than in normal intestine parisons in the secreting intestine have as yet not http://gut.bmj.com/ (p

ate free or bicarbonate containing solution (Fig. 2). mmol/l) of this ORS. It has been shown in the human on September 25, 2021 by guest. Protected copyright. In the secreting intestine, however, the addition of jejunum that sodium movement is critically depen- bicarbonate to the ORS almost halved glucose dent on the luminal sodium concentration and that absorption (p<0-001). net absorption does not occur until a luminal concentration of 90 mmol/l is reached.2' Our own work in the pH and osmolality rat whole small intestine confirms this although a The mean pH of the ORS with bicarbonate was 8 15 luminal concentration of >120 mmol/l was necessary (0.02) and of the ORS without bicarbonate was 5.89 for net sodium absorption in this secreting model. 522 (0.21) and these differed significantly (p<0.01). The Net water absorption occurred despite net sodium mean intraluminal pH, however, differed less, sug- secretion emphasising the importance of osmotic gesting that pH equilibrates rapidly in the small forces in determining water absorption. In this situa- intestine. In the normal intestine the intraluminal pH tion water absorption probably occurs as a result of was similar in those who received the ORS with and the osmotic drive generated by the hyperosmolar without bicarbonate respectively (7-08 (0.03) v 7-01 region at the tip of the villus.23 (0.03); p>0-05). In the secreting intestine, the In the secreting intestine, however, bicarbonate effluent pH was higher (7.13 (0.04)) in those perfused failed to enhance water absorption. Indeed water with ORS with bicarbonate than in those perfused absorption was actually reduced. Bicarbonate did, with ORS without bicarbonate (6.84 (0.05); p<005). however, reduce net sodium secretion (Table, Fig. The mean ORS osmolality was 316 (0.7) mOsm/ 1). The reason for this disjunction of sodium and kg. The mean intraluminal osmolality was similar in water absorption is not clear and requires further all groups. In normal intestine, osmolality was 297-3 investigation. It is of interest though that bicarbonate Gut: first published as 10.1136/gut.29.8.1052 on 1 August 1988. Downloaded from

1056 Elliott, Watson, Walker-Smith, and Farthing absorption was markedly decreased in the secreting and decreases the shelf life of ORSs. Bicarbonate is intestine compared with the normal intestine. A probably required only in the treatment of acute further unexpected observation was that glucose gastroenteritis when acidosis is severe, although it absorption was decreased in the secreting intestine in seems likely that on these occasions parenteral fluid the presence of bicarbonate whereas in the normal replacement would also be necessary. intestine bicarbonate as expected had no effect on glucose absorption (Fig. 2). Thus in our model of the MJGF is a Wellcome Trust Senior Lecturer and secreting intestine bicarbonate inhibits water and gratefully acknowledges the financial assistance of glucose absorption. the Wellcome Trust. We thank Nutricia Cow and The pH of the ORSs perfused was not adjusted to Gate Ltd for financial support. We thank Ms Nicola the physiological range for the small intestine as no Herrera for typing the manuscript. such adjustment is made when ORSs are given therapeutically. We do not consider that this differ- ence in initial pH can explain our findings because the References stimulatory effect of bicarbonate on sodium and 1 Tripp JH, Candy DCA. Oral rehydration fluids. Arch water absorption is independent of luminal pH.4 Dis Child 1984; 59: 99-101. Measurement of luminal pH in our study suggests 2 Pierce NF, Hirschhorn N. Oral fluid - a simple weapon that ORS pH equilibrates rapidly in the small against dehydration in diarrhoea. WHO Chron 1977; 31: intestine. 87-93. Luminal osmolality did not differ substantially 3 Elliott EJ, Walker-Smith JA, Farthing MJG. The role of from the original ORS osmolality, suggesting that the bicarbonate and base precursors in the treatment of observed water absorption is closely related to over- acute gastroenteritis. Arch Dis Child 1987; 62: 91-5. all absorption of osmotically active particles. Thus, in 4 Sladen GE, Dawson AM. Effect of bicarbonate on the secreting intestine, the fall in water absorption is sodium absorption by the human jejunum. Nature 1968; 218: 267-8. caused primarily by the fall in glucose absorption and 5 Turnberg LA, Fordtran JS. Carter NW, Rector FC. this might explain the disjunction of sodium and Mechanism of bicarbonate absorption and its relation- water absorption. ship to sodium transport in the human jejunum. J C/lin The adverse effect of bicarbonate seen in the Invest 1970; 49: 548-56. secreting intestine may relate to an adverse effect on 6 Turnberg LA, Bieberdorf FA, Morawski SG, Fordtran http://gut.bmj.com/ enterocyte function by an alkaline solution. This JS. Interrelationships of chloride, bicarbonate, sodium seems unlikely, however, as a similar effect was not and hydrogen transport in the human ileum. J Clin observed in the normal intestine. Further, the Invest 1970; 49: 557-66. jejunum is constantly exposed to large quantities of 7 Rolston DDK, Morilrty KJ. Farthing MJG, Kelly MJ, Clark ML. Dawson AM. Acetate aind citrate stimulate bicarbonate rich endogenous secretions from the watter aind sodium absorption in the normal human liver and pancreas. jejunum. Digestion 1986; 34: 101-4. These data show that in this model bicarbonate

8 Mahalanabis D, Patra FC. In search of a super oral on September 25, 2021 by guest. Protected copyright. does not enhance the action of this ORS and indeed rehydration solution: Can optimum use of organic appears to have a detrimental effect with respect to solute-mediated sodium absorption lead to the develop- water movement. Our findings also emphasise the ment of an absorption promoting drug? J Diarrhoeal Dis fact that the normal and secreting intestines behave Res 1983; 1: 76-81. differently and that results obtained in the normal gut 9 Islam MR, Ahmed SM. Oral rehydration solution do not necessarily apply to disease states. One must without bicarbonate. Arch Dis Child 1984; 59: 1072-5. 10 Clements ML, Levine MM, Cleaves FT, et al. Compari- be cautious when extrapolating these findings to son of simple sugar/salt versus glucose/electrolyte oral man. When taken by mouth the solute concentration rehydration solutions in infant diarrhoea. J Trop Med and osmolality of an ORS is almost certainly altered Hyg 1981; 84: 189-94. before it enters the jejunum and so these studies may 11 Elliott EJ, Armitstead JCM, Farthing MJG, Walker- not be an accurate reflection of the clinical situation. Smith JA. Oral rehydration therapy without bicarbon- There is, however, support for our observations from ate: A double blind controlled trial in children with clinical trials. ORSs without bicarbonate have been acute gastroenteritis in the United Kingdom [Abstracti. shown to be effective for correction of both dehydra- Gut 1986; 27: A 1274-5. tion and acidosis.W" Furthermore, in some studies 12 Kappy MS, Morrow G. A diagnostic approach to metabolic acidosis in children. Pediatrics 1980; 65: comparing bicarbonate and citrate, stool volume was 35 1-6. higher when bicarbonate was included in ORS.'42 13 WHO/UNICEF. Oral rehydration salts. Planning, Thus there seems little experimental or clinical establishment and operation of production. Geneva: evidence to justify the addition of bicarbonate to WHO/CDD/SER 1985; 85: 1-136. ORS- particularly as its addition increases the cost, 14 World Heillth Oroanisation Diarrhoeial Diseases Gut: first published as 10.1136/gut.29.8.1052 on 1 August 1988. Downloaded from

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