Acid-Base Transport Systems in Gastrointestinal Epithelia

134 Gut 1992;33: 1134-1145

PROGRESS REPORT Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from Acid-base transport systems in gastrointestinal epithelia

D Gleeson

Cell plasma membranes contain a variety of transport systems that carry acid or base into or out ofthe cell. Research over the past decade has led to remarkable advances in our understanding of these acid-base transport systems and has established that they have several major physiological roles. They are the primary means by which cells regulate their internal pH and they also contribute importantly to regulation of cell volume and possibly cell proliferation. Further- more, in the gastrointestinal tract they mediate absorption and secretion, not only of acids and bases but of several other electrolytes and nutrients. The major acid-base transport systems on mammalian cell plasma membranes are illustrated in Figures 1-3; their specific locations on different cell types are discussed later. Hydrogen, potassium (H+,K+) adenosine tri-

phosphatase (ATPase) extrude H+ in exchange http://gut.bmj.com/ for K' using energy derived directly from ATP hydrolysis. They are present on the apical membranes of gastric parietal cells' (Fig 1) and, in some species, colon epithelial cells,2 where they mediate acid secretion. However, they are not widely distributed in gastrointestinal epithelia. Other acid base transport systems derive their H+ HCO3- 3Na+ on September 24, 2021 by guest. Protected copyright. not from ATP but energy, directly hydrolysis, Figure 1: Acid-base transport systems on an activated gastric from coupling the movement of one ion to parietal cell. The apical membrane contains H+-K+ A TPase passive movement of another ion along its and the basolateral membrane contains Na+lH+ exchange, electrochemical gradient. In the case of sodium/ Cl-IHCO3 exchange, and Na+-HCO3 co-transport. The stochiometry ofthe Na+-HCO3 co-transporter is not known hydrogen (Na+/H+) exchange (Figs 1-3), H' (n= 1, 2, or 3). Note also basolateral membrane Na+-K' extrusion from the cell is coupled, in a 1: 1 ratio, A TPase and apical membrane Cl- and K+ channels. For to Na+ entry down its chemical gradient.3-' This details, see text (section 2). As in Figures 2 and 3, the transport systems which derive their energy directlyfrom A TP gradient depends on the sodium pump, Na+,K+ hydrolysis, are shown in closed circles. ATPase, which is present on the basolateral membrane of all gastrointestinal epithelial cells and extrudes 3 Na+ ions in exchange for 2 K+ is a result of the negative intracellular potential, ions, thus maintaining a low intracellular [Na+], which maintains a low intracellular [Cl -]. In the high intracellular [K'], and negative intra- case of Na+-HCO3 co-transport (Figs 1, 3), cellular potential. Na+/H+ exchange is almost HCO3- movement is coupled with that ofNa+ in ubiquitous in mammalian cells and can be the same direction. In most, although not all, inhibited by the diuretic amiloride. cells this sytem operates in the direction of Na+ There are several mechanisms for bicarbonate and HCO3- entry into the cell, the driving force, (HCO3 ) transport across plasma membranes.67 as for Na+/H+ exchange, being the out to in Na+ In most tissues, the enzyme carbonic anhydrase gradient. Typically, Cl-/HCO3- exchange and Gastroenterology Unit, of H' and are not affected Royal Hallamshire mediates rapid equilibration HCO3 Na+-HCO3- co-transport by Hospital, Sheffield with C02, which diffuses freely across all cell amiloride but can be inhibited by the disulphonic D Gleeson membranes. Therefore, HCO3 transport into stilbene DIDS.67 Finally, HCO3- can exit from Correspondence to: the cell is equivalent to H` transport out of the some cells' uncoupled to other ions, via plasma Dr D Gleeson, Gastroenterology Unit, Floor cell and vice versa. membrane HCO3- channels (Fig 3). The driving J, Royal Hallamshire Hospital, In the case of chloride/bicarbonate (Cl-/ force here is the negative intracellular potential. Glossop Road, Sheffield S10 2JF HC03 ) exchange (Figs 1-3), HCO3 extrusion There have been two major approaches to Accepted for publication from the cell is coupled, in a 1:1 ratio, to Cl characterising acid-base transport systems. 4 November 1991 entry along its chemical gradient. This gradirnt Firstly, by studying radiolabelled ion uptake Acid-base transport systems in gastrointestinal epithelia 1135

H+ HCO3-- transmitters, and growth factors, acting via intracellular mediators. On the other hand, 4~ measurement of pHi using fluorescent dyes has usually necessitated cell dispersal, following Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from which epithelial cells often lose their polarised characteristics. Consequently, selective assay of Na+ Ci- apical and basolateral transport systems has usually not been possible. Studies of isolated vesicles and studies of cell pHi, although providing complementary data on acid base transport systems, have defined them in purely functional terms. However, application of molecular biological techniques has recently yielded information regarding the molecular structure of these transport systems. For example, Sardet et al have obtained a human cDNA sequence from human fibroblasts, expression of which restores Na+/H+ exchange activity in a mutant fibroblast which lacks intrinsic Na+/H+ exchange.2627 The putative Na+ 2K+ Na+/H+ exchanger encoded by this cDNA sequence is a 815 amino acid glycoprotein with 10-12 hydrophobic (possibly membrane spanning) domains at its amino terminal. The Cl-/ HCO3- exchanger in red blood cells has also K+ Cl- been cloned and sequenced.28 Very recently, H+ 3Na+ similar approaches have been used to isolate Figure 2: Acid-base transport systems on a NaCI absorbing cDNA sequences encoding Na+/H+ and Cl-/ cell. Examples include ileal villus, colon, and gall bladder HCO3 exchangers from rabbit ileum (see epithelial cells. The apical membrane contains coupled Na+l below). H' and Cl-/HCO3 exchangers. The basolateral membrane contains another Na+lH+ exchanger with different kinetic characteristics and, in addition, Na+-K' ATPase and K' and Cl- channels. In some cells'5 basolateral K' and Cl- exit is via a KCL co-transport system. For details see text (section 3). http://gut.bmj.com/

into isolated plasma membrane vesicles. For example, in many tissues, 22Na can be shown to be concentrated into plasma membrane vesicles in the presence of an in to out transmembrane H+ gradient. Furthermore, a major component

ofthis Na+ uptake is a saturable function ofNa+ on September 24, 2021 by guest. Protected copyright. concentration, is temperature sensitive, is electroneutral (that is, unaffected by transmembrane potential), and is inhibited by amiloride. These properties are characteristic of a Na+/H+ exchange mechanism. Isolated vesicles were used in the first direct demonstration of Na+/H+ exchange, by Murer in rabbit jejunum.9 Similar studies have shown that not only Na+/H+ exchange,"-'7 but also, Cl-HCO3- exchange'8 23 and Na+-HCO3- co-transport2425 are widely distributed in gastrointestinal epithelia from several species including humans. Furthermore, selective isolation of vesicles from basolateral and apical membranes has often revealed a polarised distribution of these transport systems between the two membranes (Figs 1-3). This polarisation, as discussed below, is fundamental to the role of acid-base transport systems in transepithelial transport. H+ 3Na+ Figure 3: Acid-base transport systems on a HCO3- secreting Acid-base transport systems can also be cell. Examples include ileal crypt cells, hepatocytes, and, characterised in intact cells by their effects on probably, duodenal epithelial cells and pancreatic and bile intracellular pH (pHi), an approach facilitated in ductular cells. The apical membrane contains Cl-IHCO3 exchange, Cl- channels, andpossibly (?), separate HCO3 recent years by the advent of pH sensitive channels andlor Na+-HCO3 co-transport. In order to fluorescent dyes. This approach offers the operate in the direction ofHCO3 extrusion, the apical important advantage that the normal environ- Na+-HCO3 co-transporter would have to transport a ment the and has negatively charged species, that is n=2 or 3. The basolateral of transporter is preserved membrane contdins Na+IH+ exchange and Na+/HCO3 co- provided evidence for regulation of acid-base transport (the stochiometry ofwhich is not known), together transport systems by many hormones, neuro- with Na+-K' ATPase. For details see text (section 4). 136 Gleeson

Functional roles of acid base transport coupled to the Na+ gradient, which drives it systems 'uphill,' that is Cl- out of and HCO3- into the cell. Na+ dependent Cl-/HCO3- exchange is an (1) REGULATION OF INTRACELLULAR pH (pHi) important pHi regulatory mechanism in inverte- Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from In most cells, pHi is actively maintained between brate cells and in mammalian mesenchymal 7.00 and 7.40, almost one unit above the value cells67 but has not yet been described in gastro- expected if intracellular and extracellular H+ intestinal epithelia. were in electrochemical equilibrium. Further- more, pHi can recover spontaneously to baseline within minutes after exposure of cells to acute (2) GASTRIC ACID SECRETION (FIG 1) acid and alkaline loads.67 Regulation of pHi is of Gastric acid secretion is mediated by a H+,K+ major importance to cell homeostasis because ATPase on the apical membrane of the parietal many physiological phenomena are pH depen- cell which extrudes H' in exchange for K+.4 dent29; diverse examples include the rate of The enzyme has recently been cloned and glycolysis,30 the level of intracellular calcium,3' sequenced.3' Its molecular weight is about 95 000 and plasma membrane K' permeability which Daltons and it consists of two subunits: a larger may regulate intracellular electrical potential.`23 alpha subunit with several membrane spanning In most cells, including several gastro- domains and a smaller beta subunit. The struc- intestinal cells,3"' the major system regulating tural, enzymatic, and ion transporting properties pHi, in the absence of HCO3-, is Na+/H+ of gastric H+,K+ ATPase have recently been exchange. Under HCO3- free conditions, block- reviewed.' ing the Na+/H+ exchanger with amiloride or In the resting state, H+,K+ ATPase is local- removal of extracellular Na+ typically causes a ised, not on the apical membrane of the parietal fall in pHi and inhibits recovery from an intra- cell, but in tubulo-vesicular structures in the cellular acid load (usually achieved by pulse cytoplasm which are impermeable to K+ and exposure of cells to the weak base ammonia).67 Cl-. Thus, its ability to transport H+ into these Rapid recovery of pHi after intracellular acidifi- vesicles in exchange for K+ is normally limited cation is made possible by a distinctive property by the availability of intravesicular K+. H+ of the Na+/H+ exchanger, first demonstrated in transport into vesicles from resting cells can be renal epithelia by Aronson.42 The exchanger observed however, ifthese vesicles are preloaded contains a modifier site on its intracellular face with K+.`' H+ secretion can also be induced by which is activated by H' ions. Thus, intra- exposure of resting parietal cells to the K+ cellular acidosis causes a greater increase in Na+/ ionophore valinomycin.52 H' exchange activity than that expected from Stimulation of the parietal cell by secreta- the more favourable in to out H' gradient.3 5738 gogues such as histamine, gastrin, and carbachol http://gut.bmj.com/ Conversely, the Na+/H+ exchanger is down results in insertion of these H,K+, ATPase regulated by intracellular alkalosis and is usually containing tubulo-vesicles into the apical inactive above a pHi of 7.20, despite ion membrane, the area of which increases several gradients which favour continued Na+/H+ fold.53 55 The permeability of the apical mem- exchange. Hormones and growth factors that brane to K+ and Cl- also shows a marked regulate the Na+/H+ exchanger often do so increase. This is partly due to the appearance of by shifting this inverse relationship between Cl- conductive pathways in the apical mem- on September 24, 2021 by guest. Protected copyright. activity and pHi 'up' or 'down.' This shift in pHi brane.5158 These pathways are probably Cl- responsiveness may be a result of phosphoryla- selective ion channels, as demonstrated recently tion of the exchanger itself, or of intracellular by patch-clamp techniques.59 Whether the apical proteins which regulate its activity.327 membrane K+ permeability increases via open- When cells are maintained under more physio- ing ofseparate K+ channels,586061 as illustrated in logical conditions, where HCO3 and CO2 are Figure 1, or via activation of a KCI co-transport present, other pHi regulatory systems often system51 7 iS controversial. It is also unclear predominate over Na+/H+ exchange. For whether the increased apical membrane K+ and example, in hepatocytes,' the intestinal cell line Cl- permeabilities result from activation of IEC-6,36 partietal cells,43 and pancreatic acinar latent transport systems or, alternatively, from cells,4' the major mechanism of pHi recovery insertion of a separate class of K+ and Cl- from an acid load is not affected by amiloride but permeable cytoplasmic vesicles into the apical is inhibited by DIDS; furthermore, it is depen- membrane.62 Whatever the mechanism, the dent on the presence of Na+ and HCO3 but not result is that: (a) the K+ accumulated by H+,K+, Cl -. These properties are characteristic of a ATPase can be recycled into the lumen, so that Na+-HCO3 co-transport system. In some H+/K+ exchange is no longer limited by avail- cells,363744 pHi recovery from an intracellular ability of K+; and (b) secretion of Cl- ions can alkaline load is mediated by HCO3- extrusion accompany secretion of H+, thereby preserving which is dependent on the presence of Cl- and is electroneutrality. inhibited by DIDS, properties characteristic of Omeprazole,63-5 one of the group of substi- the Cl -/HCO3- exchanger. This exchanger is in tuted benzimidazoles, is a weak base with a pH of some respects a 'mirror image' of Na+/H+ 4.0. In its uncharged (unprotonated) form, it exchange in that it possesses an intracellular diffuses into acid compartments such as the modifier site which is sensitive to hydroxyl parietal cell lumen, where it becomes protonated (OH-) ions2245 1 and thus, although relatively and positively charged. It is thus trapped in the inactive at baseline pHi, is activated by intra- acid compartment, where it accumulates. The cellular alkalosis. protonated omeprazole is then converted to an In some cells, Cl-/HCO3- exchange is active metabolite, the sulphenamide, which Acid-base transport systems in gastrointestinal epithelia 1137

reacts covalently with cysteine residues on the absorption in the ileum,"-0 colon,9' 96 and gall luminal face of the H+,K+ ATPase alpha unit, bladder059790 are largely interdependent pro- resulting in irreversible inhibition of H+,K+ cesses. Furthermore, they are usually inhibited ATPase and a parallel inhibition of acid by amiloride93-%98 " and the carbonic anhydrase Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from secretion.' inhibitor acetazolamide,0093 94 9 "I' and also By itself, apical HCl secretion would pro- (albeit less consistently) inhibited by HCO3-- gressively alkalinise the parietal cell and deplete removal959099101 and by DIDS.94 102 These results it of Cl-. However, parietal cell homeostasis lend support to the model originally proposed by seems to be maintained by modulation of three Turnberg on the basis of studies in humans,0 other acid base transport systems, all localised on whereby Na+ and Cl- are transported across the the basolateral membrane: Cl-IHCO3 apical membrane via coupled Na+/H+ and Cl-/ exchange,346768 which lowers pHi by HCO3 HCO3 exchange. More recent studies have extrusion and Na+/H+ exchange,346768 and Na+- directly confirmed the presence of both Na+/H+ HCO3 co-transport,4369 both of which act to and Cl-/IHCO3- exchangers on the apical mem- alkalinise the cell. After stimulation of HCI branes of gall bladder05 and colonic`5 1621 secretion by histamine, ClW/HCO3- exchange epithelial cells and ileal villus cells'4 '9 (Fig 2). across the basolateral membrane increases The two exchangers seem to be coupled several fold70'7; the HCO3- efflux represents the indirectly via changes in pHi'9; a rise in pHi blood 'alkaline tide.' This increase in Cl-/ resulting from stimulation of apical Na+/H+ HCO3- exchange can be partly ascribed to more exchange activates apical Cl-IHCO3 exchange favourable ion gradients, such as the fall in via its OH- sensitive modifier site, as recently intracellular [Cl-] resulting from apical HCI shown in apical membrane vesicles from ileum-6 exit, but it also reflects a true activation of the and colon.22 The Na+ accumulated in the cell is exchanger. Any rise in pHi resulting from HCI then extruded in exchange for K' by the baso- secretion should activate Cl-/HCO3- exchange lateral Na+,K+ ATPase,'03 and the accumulated via its OH- sensitive modifier site72; the Cl- leaves the cell via basolateral anion channels exchanger may also be directly activated by or in some cells05 via a KCI co-transport system. intracellular mediators, such as cyclic adenosine Intestinal NaCl absorption is regulated by monophosphate (AMP) and calcium, which several hormones and neurotransmitters.83 84 It is activate acid secretion.7' Simultaneously, there is inhibited by agents that increase intracellular downregulation of the basolateral transport cyclic AMP (for example, cholera toxin, prosta- systems which act to alkalinise the cell: Na+- glandin E2, vasoactive intestinal polypeptide HCO3 co-transport73 and, possibly, Na+/H+ (VIP)), intracellular calcium (serotonin, acetyl- exchange7' (although this has been disputed70). choline, substance P), or intracellular cyclic

The net effect ofthese various modifications is guanosine monophosphate (GMP) (Escherichia http://gut.bmj.com/ to oppose the alkalinising effect of apical HCI coli toxin).'03-105 This regulation of NaCl absorp- secretion, with the result that patietal cell pHi tion may be achieved largely by modulation of increases by less than 010 pH units after acid-base transport systems. Cyclic AMP, cyclic stimulation ofHCI secretion.70`7 The parietal cell GMP, and calcium all inhibit Na+/H+ exchange thus provides a striking example of how, by in isolated enterocytes or in ileal apical mem- selective modulation of apical and basolateral brane vesicles.10609 Calcium and cylic AMP also

acid-base transport systems, it is possible to inhibit both NaCl absorption and apical mem- on September 24, 2021 by guest. Protected copyright. reconcile the conflicting demands of varying brane Na+/H1+ exchange in the gall bladder0 "" transepithelial transport and maintaining cell as do aldosterone"' 112 and activators of protein homeostasis. kinase C"13'"4 in the colon. In contrast, gluco- The mechanisms by which secretagogues corticoids"5 and alpha adrenergic agonists"6"7 exert these multiple effects on the parietal cell are increase intestinal NaCl absorption; although incompletely understood. Carbachol, gastrin, the effects of these agents on intestinal Na+/H+ and histamine all bind to specific receptors on exchange are not known, both stimulate renal parietal cell plasma membranes.50 The effects of Na+/H+ exchange."8"9 Intestinal NaCl carbachol and gastrin on acid secretion seem to absorption is also stimulated by respiratory be mediated via an increase in intracellular acidosis99 01 120 and by exposure to volatile fatty calcium,7475 achieved partly, as in many cell acids,'2' interventions which lower pHi and types, by inducing inositol phosphate turn- might thus activate apical membrane Na+/H+ over.75"- Histamine also increases intracellular exchange via an intracellular H+ sensitive calcium,7" by a mechanism independent of modifier site. Indeed, volatile fatty acids have inositol phosphate turnover,75` however, its recently been shown to activate apical membrane effect on acid secretion appears to be mediated Na+/H+ exchange in a human colon cancer cell mainly via a rise in intracellular cyclic AMP.79"0 line.'22 Taken together, these studies provide Calcium, cyclic AMP, and perhaps other media- strong evidence that apical membrane Na+/H+ tors induce phosphorylation of several intracel- exchange is a major regulated step in NaCl lular proteins,"08' which may result in activation absorption. Much less information is available of plasma membrane transport systems. regarding hormonal regulation of apical Cl-/ HCO3- exchange, however the calcium releas- ing agent serotonin inhibits the exchanger in ileal (3) NaCl ABSORPTION (FIG 2) villus cells'23 as does cyclic AMP in the gall Electroneutral NaCl absorption is a major bladder. "0 driving force for intestinal fluid absorption.02 07 Intestinal epithelial cells must also maintain Studies in several species, including humans, homeostasis in the face of varying rates of have shown that electroneutral Na+ and Cl- transepithelial NaCl transport. An increase in 1138 Gleeson

NaCI absorption initiated by stimulation of mediated by intestinal secretion of Cl- and apical Na+/H+ exchange and consequent activa- HCO3- (see below), however, malabsorption of tion of apical Cl-HCO3- exchange via its OH- NaCl, via inhibition ofapical membrane Na+/H+ sensitive modifier site would result not only in exchange and possibly Cl-/HCO3- exchange, Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from some increase in pHi, but also rises in intra- also plays a role. cellular Na+ and Cl- concentrations, cell volume, and cell membrane potential. Such disturbances in cell homeostasis may, as in the (4) HCO3 SECRETION (FIG 3) parietal cell, be minimised by simultaneous HCO3- secretion by the stomach, duodenum, regulation of apical and basolateral membrane and pancreatic and bile ducts is an important transport systems. In gall bladder'24 and means ofneutralising gastric acid and of limiting colonic,'25 epithelia, intracellular [Na+] is main- its damaging effects on the mucosa. Biliary tained virtually constant despite varying rates of HCO3 secretion may be an important driving NaCl absorption, suggesting that there is parellel force for bile acid independent canalicular bile regulation of basolateral Na+/K+ ATPase. The flow,'4' althoughwhether this secretion originates basolateral channels which mediate exit of Cl- from hepatocytes or from small bile ductules is and exit of the K' accumulated by Na+/K+ unclear. HCO3 secretion by the ileum and ATPase are also regulated in parallel with apical colon is an important cause of watery diar- membrane transport systems in several absorp- rhoea.86 103 142 tive epithelia.126 In the stomach and duodenum,143 Intestinal acid-base transport systems may be ileum, c9144-146 colon,9192'47-149 and pancreas,'50 subject to similar coordinated regulation. Recent HCO3- secretion is dependent on the presence observations in isolated vesicles from jejunal," 127 of serosal Na+ and is partly dependent on the ileal,'28-'32 and colonic'5 133 epithelial cells suggest presence ofCl- in the lumen. These findings are that the basolateral membrane also possesses consistent with a model involving Na+ depen- a Na+/H+ exchanger. Furthermore, the dent HCO3 accumulation by the cell across the apical and basolateral exchangers have different basolateral membrane and HCO3 exit across kinetic characteristics; for example, in the apical membrane via Cl -/HCO3 exchange, jejunum'27 and ileum'3 129 131 the apical Na+/H+ the accumulated Cl- recycling via apical mem- exchangerisrelativelyresistant to amiloride. This brane Cl- channels. difference between basolateral and apical Na+/ Such an asymmetrical distribution of trans- H' exchangers was originally shown on a renal port systems has been shown (using isolated epithelial cell line'34 and may be ageneral property vesicles) in cells from ileal crypts, which seem to of epithelial cells. 134A It is thus possible that the be the site of HCO3- secretion.'5' The apical

two exchangers have distinct functional roles and membrane contains Cl-/HCO3 exchange but http://gut.bmj.com/ are selectively regulated. The apical Na+/H+ not Na+/H+ exchange. This is in contrast to the exchanger might be concerned primarily with apical membrane of the absorptive ileal villus transepithelial transport and respond selectively cells, which contains both exchangers'52 (com- to agents which regulate intestinal NaCl absorp- pare Figs 2 and 3). The ileal crypt cell basolateral tion. On the other hand, the basolateral Na+/H+1 membrane contains two Na+ dependent mecha- exchanger might be concerned mainly with cell nisms for HCO3 accumulation: Na+-HCO3 homeostasis, contributing to regulation of pHi co-transport25 and Na+/H+ exchange,'52 the on September 24, 2021 by guest. Protected copyright. and, possibly also, cell volume and cell prolifera- latter accumulating HCO3- indirectly via its tion (see below). The apical and basolateral alkalinising effect on the cell. Amiloride only HCO3 transport systems might also have modestly inhibits ileal HCO3 secretion,'53 distinct characteristics and functional roles. suggesting that Na+-HCO3 co-transport may Further evaluation of these possibilities awaits be the more important of the two mechanisms. selective study ofapical and basolateral acid-base Hepatocytes have a similar distribution ofacid transport systems and their regulation in intact base transport systems: basolateral Na+/H+ NaCl absorbing epithelia. exchange and Na+-HCO3 co-transport, Malabsorption of NaCl as a result of dysfunc- canalicular membrane Cl-/HCO3- exchange tion of acid-base transport systems may be and Cl- channels.'723'54 ClV/HCO3- exchange important in the pathogenesis of diarrhoeal and Cl- channels are also present on the apical disease. The syndrome of diarrhoea with chlori- membranes of duodenal epithelial cells.'55 dorrhoea and alkalosis, seems to result from an Finally, both Na+/H+ and Cl/HCO33 absence of intestinal Cl /HCO3- exchange.'35 136 exchange are present in intact pancreatic'56 and Other cases of congenital diarrhoea have been bile'57 ductular cells, although in these cells their ascribed to absence of intestinal apical mem- localisation remains to be established. brane Na+/H+ exchange.'37 In several diseases, In the duodenum'43 and ileum,8645 a including cholera, E coli and amoebic infections, major component of HCO3- secretion is bile acid induced diarrhoea, the carcinoid and independent of luminal Cl- and therefore may Werner Morrison syndromes, and inflammatory not be mediated by apical Cl-/HCO3- exchange. bowel disease diarrhoea results from disordered Possible alternative mechanisms for HCO3 exit intestinal electrolyte transport caused by a across the apical membrane include HCO3 variety of hormones, toxins, and neurotrans- permeable channels8 and Na+-HCO3- mitters.8384 103 138~140 These agents may act via co-transport. Na+-HCO3- transport has been increases in enterocyte cyclic AMP (for example, discussed above as a mechanism for cellular cholera toxin, prostaglandin E2, VIP), cyclic accumulation of HCO3-, energised by the out to GMP (E coli toxin), or calcium (serotonin, in Na+ gradient. However, in some epithelia, substance P, bile acids). The diarrhoea is partly such as the proximal renal tubule,'59 the Na+- Acid-base transport systems in gastrointestinal epithelia 1139

HCO3- co-transporter has a stochiometry of constant pHi through varying rates of trans- three HCO3- ions for each Na+ ion. It thus epithelial HCO3- transport. carries a net negative charge and normally Finally, HCO3- secretion may result not from operates in the direction of HCO3- and Na+ direct activation of acid-base transport systems Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from extrusion, the driving force being the negative but from movement of weak acids in their intracellular potential. protonated form in the opposite direction, that is HCO3 secretion, like NaCl absorption, is from lumen to cell. The net result would be under elaborate neurohormonal control. It is consumption ofluminal H' and therefore (in the usually stimulated by agents which increase presence of carbonic anhydrase), generation of intracellular cyclic AMP, for example, secretin HCO3 . Ursodeoxycholic acid'74 and certain in the bile and pancreatic ducts and prosta- other bile acids'75 176 induce a 'hyper' choleresis in glandin E2 in the duodenum.'43 Cholera toxin several species, including man,'77 which results induces severe diarrhoea by stimulating HCO3- mainly from active secretion of HCO3 . This and Cl- secretion throughout the intestine, via HCO3- rich choleresis may partly explain the increases in intracellular cyclic AMP.'03 142 160 161 apparently beneficial effects of ursodeoxycholic HCO3- secretion predominates in vivo and Cl- acid in cholestatic liver diseases,'76 and also its secretion in vitro; the reason for this discrepancy tendency to induce surface gall stone calcifica- is not known. Leukotriones and prostaglandins, tion, one of the factors limiting its efficacy as a often produced in association with intestinal cholesterol gall stone dissolving agent.'79 inflammation, also stimulate HCO3 secre- Recent studies involving measurement ofboth tion. 162 In contrast, HCO3- secretion in the intracellular and canalicular pH in isolated rat duodenum and ileum is inhibited by alpha hepatocyte couplets suggest that ursodeoxycho- adrenergic agonists.146 158 163 lic acid does not directly activate acid base Mediators such as cyclic AMP might induce transport systems in hepatocytes'80 (although HCO3 secretion via several mechanisms. These an effect on bile ductular cells has yet to be include activation of apical membrane Cl- excluded). An alternative hypothesis has been recycling by opening of Cl- channels. Cyclic proposed, based on the observation that these AMP causes opening of apical Cl- channels in 'hyper' choleretic bile acids are excreted by the several secretory epithelia.1' 165 This effect is liver partly in an unconjugated form.5 176 181 absent in patients with cystic fibrosis,'66-'68 who According to this 'chole-hepatic shunt' hypothe- have impaired pancreatic HCO3- secretion'69 sis, the unconjugated bile acid is transported in and may possibly have a generalised defect in its ionised form across the hepatocyte canalicular gastrointestinal HCO3 secretion. Cyclic AMP membrane. Unconjugated bile acids have high and/or other mediators might also directly pKs, and will thus tend to combine with H'

activate apical HCO3 exit: (a) via Cl-IHCO3 in the bile canaliculus, thereby generating a http://gut.bmj.com/ exchange, as demonstrated in duodenal apical HCO3 ion. The protonated uncharged (and membrane vesicles"10; (b) via HCO3- channels, therefore lipid soluble) bile acid then recycles as in salivary acinar cells8; or (c) via Na'-HCO3- across -the apical membrane of the bile ductular co-transport. cell, is returned to the liver, redissociates, and Alternatively, HCO3 secretion may be the bile acid anion is re-excreted. In theory, one induced via activation of basolateral Na+/H+ bile acid molecule could recycle several times,

exchange or Na+-HCO3- co-transport, resulting thereby generating several HCO3- ions in the on September 24, 2021 by guest. Protected copyright. in cellular HCO3- accumulation and thereby canaliculus, and this would explain the 'hyper' increasing the driving force for HCO3- exit choleresis. However, each cycle will result in across the apical membrane. The accompanying hepatocyte accumulation of one H' ion which, cell alkalinisation should also activate apical Cl -/ to maintain the generation of biliary HCO3-, HCO3- exchange via its OH- sensitive modifier must be extruded across the basolateral mem- site.2246 As discussed in the section on NaCl brane. The HCO3- rich 'hyper' choleresis absorption, cyclic AMP and calcium usually induced by ursodeoxycholic acid is partly inhibit Na+/H' exchange in absorptive epithelia. inhibited by amiloride and by Na+ removal,'82 However, recent studies suggest that serotonin,'23 suggesting that the hepatocyte basolateral which increases intracellular calcium, and Na+/H+ exchanger'7 contributes to this H' forskolin,'7' which increases intracellular cyclic extrusion. AMP by direct stimulation of adenylate cyclase, both stimulate the basolateral Na+/H' exchanger in ileal crypt cells. The HCO3 secretagogue (5) ABSORPTION OF OTHER ELECTROLYTES AND carbachol also activates Na+/H+ exchange in NUTRIENTS; THE ACID MICROENVIRONMENT salivary acinar cells.172 Yet another mechanism The luminal pH near the intestinal apical mem- has been suggested for secretin induction of brane is maintained at a more acid level than bulk HCO3 uptake into bile ductular cells: insertion luminal pH. The presence of this acid micro- of cytoplasmic vesicles containing H+ ATPase environment was first suggested by studies in the into the basolateral membrane.'73 1950s on intestinal absorption of weak acids,183 Evaluation of these possible mechanisms and has been subsequently confirmed by awaits more detailed characterisation of acid- microelectrodes in several species, including base transport systems in intact HCO3- secret- man. 184-188 ing epithelia. However, by analogy with the Na+/H+ exchange is present on the apical parietal cell (see section 2), it seems likely that membranes of jejunal epithelial cells.9'-" Here, it HCO3- secretion involves parallel regulation of may function independently of Cl-/HCO3 both basolateral and apical HCO3- transport exchange, the presence of which has not been systems, thereby allowing maintenance of a near convincingly demonstrated in the jejunum.'` A 1140 Gleeson

role for apical Na+/H+ exchange in maintaining (6) REGULATION OF CELL VOLUME the acid microenvironment is suggested by When exposed to sudden changes in medium observations that the microenvironment pH is osmolality, cells behave initially as osmometers, increased by luminal perfusion with amiloride or swelling in hypo-osmotic media and shrinking in Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from with Na+ free media,'86 189 although other studies hyperosmotic media. However, cells then tend have failed to confirm these findings.'87 Micro- to return their volume towards normal, despite environment pH is also increased by cyclic continued exposure to anisotonic media.209 AMP, cyclic GMP, and activation of protein Regulatory volume decrease (RVD) and kinase C,'90"'' interventions which, as discussed regulatory volume increase (RVI) are typically in section (3), inhibit intestinal Na+/H+ achieved by activation of plasma membrane ion exchange. In the ileum and colon, microenviron- transport systems, which result ultimately in loss ment pH is higher than in the jejunum,'84 or gain ofcell electrolytes and water respectively. possibly because the apical Na+/H' exchanger is There are two major mechanisms for RVI in largely neutralised by parallel Cl-IHCO3- mammalian cells. The first mechanism is activa- exchange (see section 3 and Fig 2). tion of Na+/H+ exchange with coincidental or The acid microenvironment contributes to secondary (via a rise in pHi) activation of Cl-/ intestinal absorption of weak acids. It favours HCO3- exchange, resulting in cell accumulation conversion of these weak acids to their proto- of NaCl. The accumulated Na+ is extruded by nated uncharged form which can then diffuse Na+/K+ ATPase in exchange for K' and thus across the apical membrane into the cell.'84 An the net effect is cellular accumulation of KCI. example of this process is HCO3- absorption by This mechanism seems to be the major means the jejunum and gall bladder. In the human of RIV in gall bladder210 and some renal2' epithe- jejunum, Na+ and HCO3- absorption are inter- lial cells, hepatocytes,22 and lymphocytes.23 The dependent, are associated with an increase in evidence includes observations that RVI in these intraluminal pCO2, and are inhibited by aceta- cells is dependent on the presence of Na+, Cl, zolamide.'92 '3 These observations suggest that and HCO3- and furthermore, can be inhibited HCO3- is not absorbed directly but rather as a by amiloride and by DIDS. In addition, activa- result of H' secretion via apical Na+/H+ tion of Na+/H+ exchange can be demonstrated exchange. The pCO2 of human gall bladder by measurement of the effects of hyperosmotic bile exceeds that of hepatic bile and blood, stress on pHi.3209213214 The mechanism(s) of suggesting that the gall bladder may also absorb activation remain obscure but may involve phos- HCO3 via H' secretion.'9"'96 Gall bladder H' phorylation of the Na+/H+ exchanger.3 secretion is impaired in patients with calcified The other major mechanism of RVI uses the (calcium carbonate rich) gall stones; the out to in Na+ gradient to drive K' and Cl- into result is an abnormally alkaline gall bladder the cell.219 This electroneutral Na+-K-2Cl- http://gut.bmj.com/ bile that is supersaturated with calcium co-transport system is present on the basolateral carbonate. 11 membranes ofintestinal epithelial cells`4 and can Volatile fatty acids (VFAs), are produced by be inhibited by the diuretic bumetanide. It is this bacterial degradation of non-absorbed dietary system, rather than Na+/H+ and Cl-/HCO3- carbohydrate and are rapidly absorbed by the exchange, which seems to mediate RVI in jejunal colon, where they constitute a significant energy villus enterocytes.2' The mechanisms of RVI in source. 84 197 198 In several species, including other intestinal epithelial cells are not known. on September 24, 2021 by guest. Protected copyright. humans,'99-202 colonic VFA absorption in vivo is Regulatory volume decrease (RVD) after cell associated with HCO3- secretion and a lower swelling in hypo-osmotic media is mediated by luminal pCO2 than in the absence of VFAs. KCI loss, for which there are two main mecha- These findings suggest that VFAs combine with nisms. The first involves opening of K' and Cl- luminal H' (thereby depleting CO2 and generat- channels and mediates RVD in hepatocytes2" ing HCO3 ) and are then absorbed passively in and jejunal villus enterocytes.26 Other cells may their protonated (uncharged) form. Other lose KCI via activation of a bumetanide inhibit- observations suggest that the acid microenviron- ible KCI co-transport system.29 ment, maintained by apical Na+/H+ exchange, The above cell volume regulatory mechanisms also contributes H' to this process and thereby have usually been demonstrated by suddenly assists in VFA absorption. For example, VFA exposing cells to large changes in medium absorption is, like the microenvironment pH,'84 osmolality. It is not clear to what extent these relatively independent of bulk luminal pH.'97 mechanisms are operative in vivo, where extra- Furthermore, VFA absorption is associated with cellular fluid osmolality is usually regulated enhanced colonic Na+ absorption121 99 200 202 and within a narrow range. However, for epithelial can be partly inhibited by amiloride202 and by cells to maintain their volume through varying luminal Na+ removal'99 2"3 A third possible rates of transepithelial transport, net ion fluxes mechanism for VFA absorption is via a VFA- across the apical membrane must balance those HCO3 exchange mechanism, recently demon- across the basolateral membrane. Therefore, cell strated on apical membrane vesicles from human volume regulatory mechanisms might also play a ileum2" and rat colon.205 major role in balancing ion fluxes across the two The acid microenvironment also plays a role in membranes. Indeed, in some epithelia, the intestinal absorption of folic acid20` and oligopep- opening of basolateral membrane K' and Cl- tides.207 Patients with coeliac disease or Crohn's channels after an increase in Na+ absorption disease have an abnormally alkaline jejunal across the apical membrane is mediated partly microenvironment,"'820 which may explain the via the resulting increase in cell volume. 16 frequently observed malabsorption of folic acid Conversely, carbachol induced stimulation of and other nutrients in these patients. HCO3- secretion by salivary acinar cells results Acid-base transport systems ingastrointestinal epithelia 1 141

in activation of basolateral Na+/H+ exchange, branes. They are subject to regulation by many which is mediated partly via a decrease in cell neural, hormonal, and paracrine factors, acting volume,'72 presumably a consequence of apical via intracellular mediators. There is some HCO3 and water exit. evidence that different transport systems, and Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from even similar transport systems on the apical and basolateral membranes, are selectively (7) REGULATION OF CELL PROLIFERATION regulated. These distinctive properties may One of the most intriguing and controversial partly explain how acid-base transport systems aspects of acid-base transport systems is their can play several major physiological roles, possible role in the regulation of cellular pro- including regulation of pHi and cell volume, liferation. In white blood cells, fibroblasts and transepithelial transport, and, perhaps, cell pro- in various invertebrate cells, several agents liferation - roles which might at first seem which induce cell proliferation activate Na/H+ mutually incompatible. exchange5217; the exchanger is also spontane- Our understanding of these systems is, how- ously activated in some tumour cell lines. ever, still incomplete. For example, the role of Furthermore, exposure of cells to amiloride or coordinated regulation of apical and basolateral removal of Na+ often inhibits their prolifera- acid base transport systems in mediating tion.5 Based on these observations, it was sug- intestinal NaCl absorption and HCO3 secretion gested that a rise in pHi resulting from activation needs to be systematically evaluated. This of Na+/H+ exchange was a trigger for initiation evaluation will require selective assay of acid ofcell division. base transport systems on the apical and baso- However, most studies on the effects of mito- lateral membranes of polarised gastrointestinal gens on pHi have been performed in HCO3 free epithelial cells. Such studies have proved media. Under more physiological conditions, difficult because most epithelia are heterogenous when HCO3 is present, mitogens activate not and therefore, measurement of pHi using fluor- only Na+/H+ exchange but also other transport escent dyes in one cell type has usually required systems such as Cl-/HCO3 exchange, which in selective cell isolation, following which the cell's some cells,2829 minimises the changes in pHi. polarised characteristics are often lost. Nevertheless, it is likely that activation of Na+/ Recent advances in methodology offer several H' exchange and also the HCO3- transport possible solutions to this problem. Firstly, systems play a permissive role in cell prolifera- culture of some epithelial cell lines to form tion by maintaining pHi at a level which will monolayers on semipermeable membranes has permit DNA and protein synthesis; these pro- resulted in retention of cell polarity and has cesses often have pH optima above basal pHi and enabled selective study of the apical and baso- are inhibited by intracellular acidosis. Indeed, in lateral membranes of renal'34230 and gastro- http://gut.bmj.com/ some cells, simply raising pHi induces prolifera- intestinal381 22 cell monolayers. Secondly, assay of tion, as was demonstrated recently in fibroblasts acid base transport systems in one cell type in an by inserting and inducing expression of the gene intact heterogeneous epithelium can sometimes for H' ATPase.22' It is also possible that acid be achieved by selective loading of pH sensitive base transport systems regulate cell proliferation dye into that cell type,23' or alternatively by use of not via changes in pHi but via secondary effects single cell fluorescence microscopy. on intracellular Na+ or Cl- concentrations or cell Yet another advance has been the application on September 24, 2021 by guest. Protected copyright. volume. ofmolecular biological techniques to the study of Gastrointestinal epithelial cell proliferation is gastrointestinal acid-base transport systems. Tse influenced by many hormonal, paracrine, and et al'3''32 have recently obtained two related but intraluminal agents.22' It is still unclear whether distinct cDNA sequences from rabbit ileum, acid-base transport systems are involved in which induce expression of Na+/H+ activity in regulating proliferation. However, Na+/H+ a fibroblast cell line that lacks intrinsic Na+/H+ exchange activity is enhanced in several models exchange. The first of these putative Na+/H+ of increased gastrointestinal cell proliferation, exchangers has 95% homology with the pre- including the neonatal liver,222 the pancreas after viously cloned human fibroblast Na+/H+ exposure to trophic hormones,223 and the exchanger,262" with which it also shares several intestine after partial resection224 or exposure to functional characteristics, including extreme the carcinogen 1,2 dimethylhydrazine.225 In con- sensitivity to amiloride. Furthermore, immuno- trast, differentiation and cessation of prolifera- cytochemical studies showed that polyclonal tion in some human colon cancer cell lines is antibodies to a fusion protein incorporating this associated with down-regulation of Na+/H+ sequence localise to basolateral but not apical exchange.226 227 Finally, blocking Na+/H+ membranes from rabbit ileum. The Na+/H+ exchange with amiloride or its analogues inhibits encoded by the second cDNA sequence has been proliferation of a pancreatic cell line and of the less extensively characterised but has some liver and jejunum after partial intestinal resec- properties common to those of the ileal apical tion.228229 Further studies in this area will be of membrane Na+/H+ exchanger, including a great interest. relative resistance to amiloride. 31 32 Detailed studies on the neurohormonal regulation ofthese two putative Na+/H+ exchangers should now be Conclusion possible and are awaited with interest. Progress Gastrointestinal epithelial cells possess a variety has also been made in cloning the ileal Cl -/ of acid-base transport systems. The location of HCO3 exchanger.232 these transport systems is often highly polarised Approaches such as these should lead to between the apical and basolateral plasma mem- further major advances in our understanding of 1142 Gleeson

these fascinating and versatile 36 Wenzl E, Sjaastad MD, Weintraub WH, Machen TE. Intracellular pH transport systems regulation in IEC-6 cells, a crvptlike intestinal cell line. Am ] Phs'siol over the next decade. 1989; 257: G732-40. 37 Sundaram U, Knickelbein RL, Dobbins JW. Intracellular pH regula- I am grateful to the Wellcome Trust for financial support and to tion in the ileum: Na+/H+ and Cl-/HCO3- exchange in isolated Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from Dr Martin Steward, Department of Physiological Sciences, crypt and villus cells. Am] Phvsiol 1991; 260: G440-9. University of Manchester for critical comment on an earlier draft 38 Watson AJM, Levine S, Donowitz M, Montrose MH. Kinetics and of the manuscript. regulation of a polarized Na+-H+ exchanger from Caco-2 cells, a human intestinal cell line. Amj Phvsiol 1991; 261: G229-38. 1 Rabon EC, Reuben MA. rhe mechanism and structure of the gastric 39 Renner EL, Lake JR, Persico M, Scharschmidt BF. Na+/H+ exchangc H,-K ATPase. Ann Rev Phvsiol 1990; 52: 321-44. activity in rat hepatocytes; role in regulation of intracellular pH. 2 Kaunitz JD, Sachs G. Identification of a vanadate sensitive K+ Am] Phvsiol 1989; 256: G44-52. dependent proton pump from rabbit colon. 7 Biol Chem 1986; 261: 40 Gleeson D, Smith ND, Boyer JL. Bicarbonate dependent and indepen- 14005-10. dent intracellular pH regulatory mechanisms in rat hepatocytes. 3 Grinstein S, Rothstein A. Mechanisms of regulation of the Na/H Evidence for Na-HCO3 cotransport. 7 Clin Invest 1989; 84: 312- exchanger. J7 Membr Biol 1986; 90: 1-12. 21. 4 Aronson P'S. Kinetic properties of the plasma membrane Na+/H+ 41 Muallem S, Lalsbeg P. Intracellular pH regulatory mechanisms in exchanger. Ann Rev Plhysiol 1985; 47: 545-60. pancreatic acinar cells. J. Characteristics of H+ and HCO3- 5 Grinstein S, Rotin D, Mason MJ. Na+/H+ exchange and growth factor transporters. ] Biol Chem 1990; 265: 12806-12. induced cytosolic pH changes. Role in cellular proliferation. Biochim 42 Aronson PS, Nee J, Suhm MA. Modifier role of internal H + in Biophvs Acta 1989; 988: 73-97. activating the Na+-H+ exchanger in renal microvillus membrane 6 Roos A, Boron WF. Intracellular pH. Physiol Rev 1981; 61: 296-432. vesicles. Nature 1982; 229: 161-3. 7 Madshus IH. Regulation of intracellular pH in eukaryotic cells. 43 Townslev MC, Machen TE. Na-HCO3 cotransport in rabbit parietal Biochem,J 1988; 250: 1-8. cells. Amj Phvsiol 1989; 257: G350-6. 8 Lau KR, Elliott AC, Brown PD. Acetylcholine-induced intracellular 44 Benedetti A, Strazzabosco M, Boyer JL, Haddad P, Graf J, Boyen J. acidosis in rabbit salivary gland acinar cells. Am j Phvsiol 1989; 256: Cl /HCO3 exchange in rat hepatocytes: role in regulation of intra- C288-95. cellular pH. AmjPhysiol 1991; 261: G512-21. 9 Murer H, Hopfer U, Kinne R. Sodium/proton antiport in brush border 45 Mason MJ, Smith JD, Garcia-Soto JdeJ, Grinstein S. Internal pH membrane vesicles isolated from rabbit small intestine and kidney. sensitive site couples Cl-/HCO3- exchange to Na+/H+ antiport in Biochemj 1976; 154: 597-604. lymphocytes. Amj Physiol 1989; 256: C428-33. 10 Cassano G, Steiger B, Murer H. Na/H and Cl/HCO3 exchange in rat 46 Mugharbil A, Knickelbein R, Aronson PS, Dobbins JW. Rabbit ileal jejunal and rat proximal tubular brush border membrane vesicles. brush border membrane Cl-/HCO3- exchange is modulated by an Studies with acridine orange. Pflugers Arch 1984; 400: 309-17. internal pH sensitive modifier site. Am J Phvsiol 1990; 259: 11 Kleinman JG, Harig JM, Barry JA, Ramaswamy K. Na+ and H+ G666-70. transport in human jejunal brush border membrane vesicles. 47 Sachs G, Chang HH, Rabon E, Schachman R, Lewin M, Saccarani G. A AmJ7 Phvsiol 1988; 255: G206-1 1. non electrogenic H+ pump in parietal membranes of hog stomach. 12 Kichuchi K, Alumrad NN, Ghishan FK. Na+/H+ exchange by brush J]Biol Chem 1976; 255: G313-8. border membrane sesicles of human ileum. Gastroenterology 1988; 95: 48 Forte JG, Machen TE, Obrink KJ. Mechanisms of gastric H + and Cl - 388-93. transport. Ann Rev Phvsiol 1980; 42: 111-26. 13 Ramaswamy K, Harig JM, Kleinman 1G, Harris MS, Barrv JA. 49 Sachs G. The parietal cell as a therapeutic target. Scand]7 Gastroenlerol Sodium-proton exchange in human ileal brush-border membrane 1986; 21 (suppl 1 18): 1-10. vesicles. Biochim Biophys Acta 1989; 981: 193-9. 50 Wolfe MM, Soll AH. The physiology of gastric acid secretion. N Engi 14 Knickelbein R, Aronson PS, Atherton W, Dobbins JW. Sodium and JMed 1988; 319: 1707-15. chloride transport across rabbit ileal brush border. 1. Evidence for 51 Shull GE, Lingrel JB. Molecular cloning of the rat stomach (H +,K+) Na+/H + exchange. Am3' Physiol 1983; 245: G504-10. ATPase..JBiol Chem 1986; 261: 16788-91. 15 Foster ES, Dudeja PK, Brasitus TA. Na+/H+ exchange in rat colonic 52 Hershey SJ, Steiner L. Stimulation of acid formation in isolated gastric brush border apical membrane vesicles. Am J Physiol 1986; 250: glands by valinomycin. Am] Physiol 1988; 255: G313-8. G781-7. 53 Ito S. Functional gastric morphology. In: Johnson LR, ed. Phvsiologv of 16 Rajendran VM, Binder HJ. Characterisation of Na-H exchange in apical the gastrointestinal tract. New York: Raven Press, 1987: 817-51. membrane vesicles of rat colon. J7 Biol Chem 1990; 265: 8408-14. 54 Forte JG, Forte TM, Black JA, Okamoto C, Wolosin JM. Ultra- 17 Moseley RH, Meier PJ, Aronson Boyer Na-H exchange in rat structural related to PS, JL. changes functional activity in gastric osxyntic cells. http://gut.bmj.com/ liver basolateral but not canalicular plasma membrane vesicles. Am] Physiol 1981; 241: G349-58. AmJ Physiol 1986; 250: G35-43. 55 Smolka A, Helander HF, Sachs G. Monoclonal antibodies against gastric 18 Liedtke CM, Hopfer U. Mechanism of Cl- translocation across small H+,K+-ATPase. AmjPhysiol 1983; 245: G589-96. intestine brush-border membrane. Demonstration of Cl-OH- 56 Perez A, Blissard D, Sachs G, Hershey SJ. Evidence for a chloride exchange and Cl- conductance. AmJI Physiol 1982; 242: G272-80. conductance in the secretory membrane of the parietal cell. 19 Knickelbein R, Aronson PS, Sehron CM, Seifter J, Dobbins JW. Am] Physiol 1989; 256: G299-305. Sodium chloride transport across rabbit ileal brush border. II. 57 Cuppoletti J, Sachs G. Regulation of gastric acid secretion via modula- Evidence for Cl-HCO3 exchange and mechanism of coupling. tion of a chloride conductance. ] Biol Chem 1984; 259: 14952-9. Amj Phyvsiol 1985; 249: G236-45. 58 Reenstra WW, Forte JG. Characterisation of K+ and Cl- conductances 20 Ramaswamy K, Chung M, Barry IA. Chloride/Bicarbonate exchange in in apical membrane vesicles from stimulated rabbit oxyntic cells. human ileal brush border membrane vesicles. Gastroenterology 1988; Am] Ptvsiol 1990; 259: G850-8. on September 24, 2021 by guest. Protected copyright. 94: A366. 59 Saccomani G, Sarnas CGP, Smith PR, Kirk KL, Shoemaker RL. 21 Foster ES, Dudeja PK, Brasitus TA. Cl-OH exchange in rat distal Histamine induced Cl- channels in apical membranes of isolated colon. Amj Physiol 1990; 258: G261-7. rabbit cells. Am]Physiol 1991; 26: C1OOO-l 1. 22 Rajendran VM, Osterlin M, Binder HJ. pH-regulated Cl uptake in 60 Wolosin JM, Forte JG. Stimulation of oxyntic cell triggers K + and Cl - apical membrane vesicles of rat distal colon. Gastroenterology 1991; conductances in the apical H + ,K + ATPase membrane. Am]J Phvsiol 100: A700. 1984; 246: C537-45. 23 Meier PJ, Knickelbein RG, Moseley RH, Dobbins JW, Boyer JL. 61 Gunther RD, Bassilian S, Rabon EC. Cation transport in vesicles from Evidence for carrier mediated chloride/bicarbonate exchange in secreting rabbit stomach. J Biol Chem 1987; 262: 13966-72. canalicular rat liver plasma membrane vesicles.J7 Clin Invest 1985; 75: 62 Im WB, Blackman DP, Davis JP. Studies on K+ permeability of rat 1256-63. gastric microsomes. JBiol Chem 1985; 260: 9452-60. 24 Rajendran VM, Oesterlin M, Binder HJ. Sodium uptake across baso- 63 Sachs G, Carlsson E, Lindberg P, Wallmark B. Gastric H K ATPase as lateral membrane of rat distal colon. Evidence for Na-H exchange and a therapeutic target. Ann Rev Pharmacol Toxicol 1988; 28: 269-84. Na anion cotransport.J Clin Invest 1991; 88: 1379-85. 64 Maton PN. Omeprazole. N Engli Med 1991; 324: 965-75. 25 Zamir Z, Barry IA, Ramaswamy K. Sodium transport in human 65 Wallmark B. Mechanism of action of Omeprazole. Scand] Gastro 1986; intestinal basolateral membrane vesicles. Gastroenterology 1991; 100: 21 (suppl 118): 11-7. A710. 66 Wallmark B, Larsson H, Humble L. The relationship between gastric 26 Sardet C, Franchi A, Pouyssegur J. Molecular cloning, primary acid secretion and gastric H+ ,K + ATPase activity.J Biol Chem 1985; structure and expression of the human growth factor activatible 260: 13681-4. Na+/H+ antiporter. Cell 1989; 56: 271-80. 67 Muallem S, Burnham C, Blissard D, Berglindh T, Sachs G. Electrolyte 27 Sardet C, Counillon L, Franchi A, Pouyssegur J. Growth factors transport across the basolateral membrane of parietal cells. induce phosphorylation of the Na+/H+ antiporter, a glycoprotein J Biol Chem 1985; 260: 6641-53. of 110 kilodaltons. Science 1990; 247: 723-6. 68 Machen TE, Paradiso AM. Regulation of intracellular pH in the 28 Kopito RR, Lodish HF. Primary structure and transmembrane orienta- stomach. Ann Rev Physiol 1987; 49: 19-33. tion of the murine anion exchange protein. Nature 1985; 316: 234-8. 69 Curci S, Debellis L, Fromter E. Evidence for rheogenic sodium 29 Busa WB. Mechanisms and consequences of pH mediated cell regula- bicarbonate cotransport in the basolateral membrane of oxyntic cells tion. Ann Rev Physiol 1986; 48: 389-402. of frog gastric fundus. Pflugers Arch 1987; 408: 497-504. 30 Fidelman ML, Seeholzer SH, Walsh KB, Moore RD. Intracellular pH 70 Muallem S, Blissard D, Cragoe EE, SachsG. Activation of Na+/H+ and mediates action of insulin on glycolysis in frog skeletal muscle. Cl-/HCO3- exchange by stimulation of acid secretion in the parietal AmJ Physiol 1982; 242: C87-93. cell. ] Biol Chem 1988; 263: 14703-11. 31 Siskind MS, McCoy CE, Chobanian A, Schwartz JH. Regulation of 71 Paradiso AM, Townsley MC, Wenzl E, Machen TE. Regulation intracellular calcium by cell pH in vascular smooth muscle cells. of intracellular pH in resting and in stimulated parietal cells. AmJ Physiol 1989; 256: C234-40. Am] Physiol 1988; 257: C554-61. 32 Duffy ME, Dewar DC. IntracellularpH and membrane K+ conductance 72 Wenzl E, Machen TE. Intracellular pH dependence of buffer capacity in rabbit distal colon. AmJ Physiol 1990; 258: C336-43. and anion exchange in the parietal cell. Am] IPhvsiol 1989; 257: 33 Henderson RM, Krumpholtz BF, Boyer JL, Graf I- Effect of intra- G741-7. cellular pH on potassium conductance in liver. Pflugers Arch 1988; 73 Townsley MC, Machen TE. Na/HCO3 cotransport in oxyrntic cells is 412: 334-5. inhibited by histamine. FASE:B]E 1988; 2: A752. 34 Paradiso AM, Tsien RY, Demarest JR, Machen TE. Na/H and 74 Negulescu P, Reenstra WW, Machen TE. Intracellular calcium require- C1/HCO3 exchange in rabbit oxyntic cells using fluorescence ments for stimulus-secretion coupling in parietal cells. Am]7 Phvsiol microscopy. AmJf Physiol 1987; 253: C30-6. 1989; 256: C241-51. 35 Giore I, Hoinard C. Na±+/H±+ exchange in isolated hamster enterocvytes. 75 Chew CS, Brown MR. Release of Ca2+ and eles-ation oSf inossitol Its major role in intracellular pH regulation. Gastroenterology 1989; 97: triphosphate byr secretagognes in parietal and chief cells isolated from 882-7. rabbit gastric mucosa. Biochim Biophvs Acta 1986; 888: 116-25. Acid-base transport systems in gastrointestinal epithelia 1143

76 Puurtinen J, Schwabe U. Effect of gastric secretagogues on the formation 115 Turmanian SG, Binder HJ. Aldosteronc and glucocorticoid rccptor- ofinositol phosphates in isolated gastric cells of the rat. BrJ7 Pharmtacol specific agonists regulatc ioni transport in rat proximal colon. 1987; 90: 479-90. Am] Phvsiol 1990; 258: G492-8. 77 Chiba r, Fisher SK, Park J, Seguin EB, Agranoff BW, Yamada 'T. 116 Chang EB, Field M, Miller RJ. Alpha2 adrenergic receptor regulation of Carbamylcholine and gastrin induce inositol lipid turnover in canine ion transport in rabbit ileum. Am' Phtsvisol 1982; 242: G237-42. Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from gastric parietal cells. AmJ Phvsiol 1988; 255: G99-105. 117 Sellin JH, DeSoigne R. Regulation of Na-Cl absorption in rabbit 78 Ljungstrom M, Chew CS. Calcium oscillations and morphological proximal colon in vitro. Am]' Phvsiol 1987; 252: G45-5 1. transformations in single cultured gastric parietal cells. AmJI Physiol 118 Kinsella JL, Freiberg JM, Sacktor B. (ilucocorticoid activation oft 1991; 260: C67-78. Na t/H+ exchange in renal bruish border sesicles; kinetic effects. 79 Soll AH, Wollin A. Histamine and cyclic AMP in isolated canine parietal Am]' Phvsiol 1985; 248: F233-9. cells. AmJ Phvsiol 1979; 237: E444-52. 119 Nord EP, Howard MIJ, Hafezi A, Moradehagi P, Zavjstub S, Insel P'A. 80 Brown MR, Chew CS. Carbachol-induced phosphorylation in parietal Alpha2 adrenergic agonists stimulate Na/H antiport activity in the cells: regulation by [Ca2 + J. AmJ7 Phvsiol 1989; 257: G99-1 10. rabbit renal proximal tubule. ] Clin Invest 1987; 80: 1755-62. 81 Urushidani T, Hanzel DK, Forte JG. Protein phosphorylation associ- 120 Vacarezza SC, Charnev AN. Acid base effects on ileal sodium ated with stimulation of gastric glands. Biochim BRiophvs Acta 1987; chloride absorption in vitro. Am]7 Phtsiol 1988; 254: G329-33. 930: 209-19. 121 Binder HJ, Mehta P. Short chain fatty acids stimulate active soditim and 82 Hopfer U, Liedtke CM. Proton and bicarbonate transport mechanisms chloride absorption in vitro in the rat distal colon. Gastroenterologs in the intestine. Ann Rev Phvsiol 1987; 49: 51-67. 1989; 96: 989-96. 83 Binder HJ. Absorption and secretion of water and electrolytes by 122 Montrose MH. Transport of short chain fatty acids in cultured huiman small and large intestine. In: Sleisenger MH, Fordtran JS, eds. colon epithelial cells. Gastroenterologv 1990; 98: A549. Gastrointestinal disease: pathop/vsiology, diagnosis, management. 123 Sundaram U, Knickelbein RG, Dobbins JW. Mechanism of intestinal Philadelphia: W B Saunders, 1988: 1022-44. secretion. Effect of serotonin on rabbit ileal crypt and sillus ccils. 84 Field M, Rao MC, Chang EB. Intestinal electrolyte transport and J Clin Invest 1991; 87: 743-6. diarrhoeal disease. N Engl3 Med 1989; 321: 800-6, 879-83. 124 Moran WM, Hudson RL, Schultz SG. TIranscellular Na absorption and 85 Reuss L. Ion transport across gallbladder epithelium. Phlvsiol Rev 1989; intracellular Na activities in rabbit gallbladder. Am 7 Phtvsiol 1986; 69: 503-45. 251: G155-9. 86 Sullivan SK, Field M. Ion transport across mammalian small intestine. 125 Turnheim K, Hudson RL, Schultz S(i. Cell Na+ actisities and In: Schultz SG, Field M, Frizzell RA, eds. Handbook of p/vsiologv. transcellular Na+ absorption by descending colon from normal and Section 6. The gastrointestinal system. Vol IV. Intestinal absorption Na+ deprived rabbits. Pflugers Arch 1987; 410: 279-83. and secretion. New York: Oxford University Press, 1991: 287-301. 126 Dawson DC, Richards NW. Basolateral K conductance: rolc in regula- 87 Halm DR, Frizzell RA. Ion transport across the large intestine. In: tion of NaCI absorption and secretion. Am ] PhYtsiol 19911; 259: Schultz SG, Field M, Frizzell RA, eds. Handbook of Physiology. C181-95. Section 6. The gastrointestinal system. Vol IV. Intestinal absorption 127 Acra SA, Dykes W, Nylander W, Ghishan FK. Characterisation of a and secretion. New York: Oxford University Press, 1991: 257-73. distinct Na+/H + exchanger in the basolateral membranes of human 88 Turnberg LA, Bieberdorf FA, Morawski SG, Fordtran JS. Inter- jejunum. Gasiroenterologv 1991; 100: A678. relationships ofchloride, bicarbonate, sodium and hydrogen transport 128 Barros F, Dominguez P, Velasco G, Lazo PS. Na'/H' exchange is in the human ileum. J Clin Invest 1970; 49: 557-67. present in the basolateral membranes from rabbit small intestine. 89 Podesta RB, Mettrick DF. HCO3- and H+ secretion in rat ileum in Biochem Biophvs Res Commun 1986; 134: 827-34. vivo. Aml Phyvsiol 1977; 232: E574-9. 129 Knickelbein RG, Aronson PS, Dobbins JW. Characterisatiotl of 90 Nellans HN, Frizzell RA, Schultz SG. Brush border processes and Na+/H t exchangers on villus cells in rabbit ileum. Am 7 P/htsiol transepithelial Na and Cl transport by rabbit ileum. Am J Phvsiol 1990; 259: G802-6. 1974;226: 1131-41. 130 Ma Al, Tse CM, Watson AJM, et al. Expression and extracellular 91 Davis GR, Morawski SG, Santa Anna CA, Fordtran JS. Evaluation of regulation of the rabbit ilea! villus basolateral membrane Na + /H + chloride/bicarbonate exchange in the human colon in vivo. exchanger in a fibroblast. Gastroenterologv 1991; 100: A695. J Clin Invest 1983; 71: 201-7. 131 lIse CM, Watson AJM, Ma Al, Pouyssegur J, Donowitz M. Cloning and 92 Deverode GJ, Phillips SF. Conservation of sodium, chloride and water functional expression of a second novel rabbit ileal villus epithelial cell by the human colon. Gastroenterology 1969; 56: 101-9. Na+/H+ exchanger. Gastroenterologs% 1991;100: A258. 93 Binder HJ, Foster ES, Budinger ME, Hayslett JP. Mechanism of 132 Tse CM, Ma Al, Yang VW, et al. Molecular cloning and expression of electroneutral sodium-chloride absorption in the distal colon of the a cDNA encoding the rabbit ileal villus basolateral membrane Na/H rat. Gastroenterology 1987; 93: 449-56. exchanger. EMBO] 1991; 10: 1957-67. 94 Lubke R, Haag K, Berger E, Knauf H, Gerok W. Ion transport in rat 133 Dude'ja PK, Foster ES, Brasitus TA. Na + -H + antiporter of rat colonic proximal colon in vivo. AmJ Phvsiol 1986; 256: G132-9. basolateral membrane sesicles. Am] Phtvsiol 1989; 257: G624-32. http://gut.bmj.com/ 95 Foster ES, Budinger ME, Hayslett JP, Binder HJ. Ion transport in the 134 Haggerty JG, Agarwal N, Reilly RF, Adelberg A, Slayman C\X'. proximal colon of the rat. Sodium depletion stimulates neutral sodium Pharmacologicallv different Na/H antiporters on the apical and chloride absorption.JI Clin Invest 1986; 77: 228-35. basolateral surfaces of cultured porcine kidney cells (LLC-IPK 96 Sellin JH, DeSoignie R. Rabbit proximal colon: a distinct transport Proc Natl Acad Sci USA. 1988; 85: 6797-801. epithelium. AmJ Phvsiol 1984; 246: G603-10. 134A Clark DJ, Limbird LE. Na-H' exchange subtypes: a predictive 97 Frizzell RA, Dugas MC, Schultz SG. Sodium chloride transport by review. Am] Physiol 1991; 261: C945-53. rabbit gallbladder. Direct evidence for a coupled NaCl influx process. 135 Turnberg LA. Abnormalities in intestinal electrolyte transport in J Gen Physiol 1975; 65: 769-95. congenital chloridorrhoea. Gut 1971; 12: 5-44-50. 98 O'Grady SM, Wolters PJ. Sodium and chloride transport across isolated 136 Bieberdorf FA, Gorden P, Fordtran JS. Pathogenesis of conlgenital

porcine gallbladder. Am3' Phvsiol 1989; 257: C45-5 1. alkalosis with diarrhoea. Implications for the physiology of normal on September 24, 2021 by guest. Protected copyright. 99 Goldfarb DS, Egnor RW, Charney AN. Effect of acid base variables on ileal electrolyte absorption and secretion. ] Clin Invest 1972; 51: ion transport in rat colon. J Clin Invest 1988; 81: 1903-10. 1958-68. 100 Nellans HN, Frizzell RA, Schultz SG. Effect of acetazolamide on 137 Booth IW, Strange G, Murer H, Fenton FR, Milla P. Defective jeptinal sodium and chloride transport by in vitro rabbit ileum. AmJI Phvsiol brush border Na+/H+ exchange: a cause of congenital secretory 1975; 228: 1808-14. diarrhoea. Lanceg 1985; i: 1066-9. 101 DeSoigne R, Sellin JH. Acid base regulation of ion transport in rabbit 138 Fondacaro J. Intestinal ion transport and diarrhoeal disease. ileum in vitro. Gastroenterology 1990; 99: 132-41. Am ] Physiol 1986; 250: G1-8. 102 Hatch M, Freel RW, Goldner AM, Earnest DL. Oxalate and chloride 139 McGowan K, Piver G, StoffJS, Donowitz M. Role of prostaglandins and absorption by the rabbit colon; sensitivity to metabolic and anion calcium in the effects of Entamoeba Histolvtica on colonic electrolytc transport inhibitors. Gut 1984; 25: 232-7. transport. Gastroenterologv 1990; 98: 873-80. 103 Field M. Secretion of water and electrolytes by mammalian small 140 Dharmsathaphorn K, Huott PA, Vongkovit P, Beuerlein G, Pandol SI, intestine. In: Johnson LR, ed. Physiology of the gastrointestinal tract. Ammon HV. Cl - secretion by bile salts. A study of the mechanism ott New York: Raven Press, 1981: 963-82. action based on a cultured colonic epithelial cell line. 7 C/int Invest 104 Donowitz M, Welsh MJ. Ca+ + and cyclic AMP in regulation of 1989; 84: 945-53. intestinal Na, K and Cl transport. Ann Rev Phvsiol 1986; 48: 135-50. 141 Hardison WGM, Wood CA. Importance of bicarbonate in the bile acid 105 Guandalini S, Rao MC, Smith PL, Field M. cGMP modulation of independent fraction of bile flow. Am]7 Phvsiol 1978; 285: E158-64. ileal ion transport: in vitro effects of Escherichia coli heat stable 142 Moore WL Jr, Bieberdorf FA, Morawski SG, Finkelstein RA, Fordtran enterotoxin. Am J Physiol 1982; 243: G36-41. JS. Ion transport during cholera induced ileal secretion on the dtog. 106 Semrad CE, Chang EB. Calcium mediated cAMP inhibition of Na +/H + 7 Clin Invest 1971; 50: 312-8. exchange in isolated chick enterocytes. Am J7 Physiol 1987; 252: 143 Flemstrom G, Garner A. Gastroduodenal HCO3 transport: character- C315-22. istics and proposed role in acidity regulation and mucosal protection. 107 Hirose R, Chang EB. Effects of serotonin on Na+/H+ exchange and Am ] Phvsiol 1982; 242: G183-93. intracellular calcium in isolated chick ettterocvtes. Am7 Phvsiol 1988; 144 Hubel KA. Effect of luminal chloride concentration on bicarbonatc 254: G891-7. secretion in rat ileum. Am] Phvsiol 1969; 217: 40-5. 108 Semrad CE, Cragoe EJ Jr, Chang EB. Inhibition of Na/H exchange in 145 Smith PL, Cascairo MA, Sullivan SK. Sodium dependence of luminal avian intestine by atrial natiuretic factor. J Clin Invest 1990; 86: alkalinisation by rabbit ileal mucosa. Am ] Phvsiol 1985; 249: 585-9 1. G358-68. 109 Emmer E, Rood RP, Wesolek JH, ei al. Role of calcium and calmodulin 146 Sellin JH, DeSoignie R. Regulation of bicarbonate transport in rabbit in the regulation of the rabbit ileal brush border membrane Na + /H + ileum: pH stat studies. Am] P'hvsiol 1989; 257: G607-15. antiporter. J7 Membr Biol 1989; 108: 207-15. 147 Sullivan SK, Smith PL. Bicarbonate secretion by rabbit proximal colon. 110 Reuss L, Segal Y, Altenberg G. Regulation of ion transport across Am] Ph/vsiol 1986; 251: G436-45. gallbladder epithelium. Ann Rev Phvsiol 1991; 53: 361-73. 148 Frizzell RA, Koch MI-, Schultz SG. Itn transport by rabbit colon. 1. Ill Foster ES, Zimmerman TW, Havslett JP, Binder HJ. Corticostertsid Actis-e and passis-ecomponents.]7,elmbr ioI 1976; 27: 297-316. alteration of active electrolyte transport in rat distal ctolsn. 149 Feldman GM, Stephenson SL.. H and HCO3- flux across aptcal AmJ7 Physiol 1983; 245: G668-75. surface otf rat diistal colon. Am]7 Phtssiol 1990); 259: C35-40. 112 Rajendran VM, Kashgarian M, Binder HI. Aldosterone indtlction olf 150 Kuijpers G.AJ, 1)e Pont JHIM1. Role otf proton andi btcarbonatc tratsport electrogenic stsdium transport in apical membrane sresicles tsf rat distal in pancreatic cell function. Ann evP/2'tvsiol 1987; 49: 87-103. colon. J Bio/ (,hem 1989; 264: 18638-44. 151 Welsh MI1, Smith 1'1., Fromm M, Frizzell RA. C rspts are thc sttc of 113 Dtnowitz M, Cheng HY, Sharp GW. Effects tof phorbol esters tsn intestinal fluidi andei lectrolyte secretionl. Scienlej 1982; 218: 1219-21. sodium and chloride transport in rat cslon. A^m .7 Pstysiol 1986; 251: 152 Knickelbein R(,, Aronson l'S, 1)obbins 1W'. Mecmbrant dtistritbutton of (i509- 17. sodiium:hvdrogen and chloride:bicarbonate exchangcrs to crspt andt 114 Ahn I, Chang EC, Field M\. I'hosrbol ester inhibition of Na/H exchange villus cell membranes f~rom rabbit leinu. .7 (,i/tn Inveo 1988; 82: in rabbit proximal colon. .Am]7 tsPsiss/ 1985; 249: C527-31). 2158-63. 1144 Gleeson

153 AMinhas B, Sullivan SK, Field M. Ileal HCO3- secretion in vitro: effects of Esterichia coli STa enterotoxin and other secretagogues on mucosal of Na and Cl. Gasiroenterology 1990; 98: A548. surface pH of rat small intestine in viso. Proc R Soc Lond 1988; 234: 154 Renner EL, Lake JR, Scharschmidt BF, Zimmereli B, Meier PJ. Rat 219-37. hepatocytes exhibit basolateral Na- HCO3 cotransport. J Clin Invest 191 Shimada T, Hoshi 'F. Na+ dependent elevation of the acidic cell surface 1989; 83: 125-35. pH (microclimate pH) of rat jejunal villus cells induced by cyclic Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from 155 Brown CDA, Dunk CR, Turnberg LA. Cl- HCO3 exchange and anion nucleotides and phorbol ester: possible mediators of the regulation of conductance in rat duodenal apical membrane vesicles. AmJI Phvsiol the Na +/H + antiporter. Biochim Biophvs Acta 1988; 937: 328-34. 1989; 257: G661-7. 192 Sladen GE, Dawson AM. Effect of bicarbonate on sodium absorption by 156 Stimenhill El, Machen TE, Williams JA. pH regulatory mechanisms in the human jejunum. Nature 1968; 218: 267-8. rat pancreatic duct cells. AmJ Physiol 1988; 254: G925-30. 193 Turnberg LA, Fordtran JS, Carter NW, Rector FC Jr. Mechanism of 157 Strazzabosco M, Mennone A, Boyer JL. Intracellular pH regulation in bicarbonate absorption and its relationship to sodium transport in the isolated rat bile duct epithelial cells. J Clin Invest 1991; 87: 1503-12. human jejunum.J7 Clin Invest 1970; 49: 548-56. 158 Sheerin HE, Field M. Ileal HCO3 secretion: relationship to Na and Cl 194 Shiffman ML, Sugerman HJ, Moore EW. Human gallbladder mucosal transport and effect oftheophylline. Amj Physiol 1975; 228: 1065-74. function. Effect of concentration and acidification of bile on 159 Boron WF, Boulpaep EL. Intracellular pH regulation in the renal cholesterol and calcium solubilitv. Gastroenterologv 1990; 99: 1452-9. proximal tubule of the salamander. J Gen Phvsiol 1983; 81: 53-94. 195 Martheau C, Sastre B, Iconiomidis N, Portugal H, Pauli A-M, Gerolami 160 Banwell JG, Pierce NF, Mitra RK, Brigham KL, et al. Intestinal fluid A. pH regulation in human gallbladder bile: study in patients with and and electrolyte transport in human cholera. J Clin Invest 1970; 49: without gallstones. Hepatology 1990; 11: 997-1002. 183-95. 196 Gleeson D, Hood KA, Murphy GM, Dowling RH. Calcium and 161 D)onowitz M, Binder HJ. Effect of enterotoxins of V. cholera, E. coli, carbonate ion concentrations in gallbladder and hepatic bile. Results and S. dysenteriae type 1 on fluid and electrolyte transport in the in patients with radiolucent and calcified stones. Gastroenterology colon. J Infect Dis 1976; 134: 135-43. 1992; 102: 1707-16. 162 Chang EB, Musch MW, Mayer L. Interleukins 1 and 3 stimulate anion 197 Cummings JH. Short chain fatty acids in the human colon. Gui 1981; 22: secretion in chicken intestine. Gastroenterologv 1990; 98: 1518-24. 763-79. 163 Nvlander 0, Flemstrom G. Effects of alpha adrenergic agonists and 198 Fleming SE, Arce DS. Volatile fatty acids: their production, absorption, antagonists on duodenal surface epithelial HCO3 secretion in the rat in utilization and roles in human health. Clin Gastroenterol 1986; 15: vivo. Acta Physiol Scand 1986; 26: 433-41. 787-814. 164 Gray MA, Greenwell JR, Argent BE. Secretin regulated chloride 199 Rubsamen K, von Engelhardt W. Absorption ot Na, H ions and short channel on the apical plasma membrane of pancreatic duct cells. chain fatty acids from the sheep colon. Pflugers Arch 1981; 391: J MembrBiol 1988; 105: 131-42. 141-6. 165 Welsh MJ. Electrolyte transport by airway epithelia. Phvsiol Rev 1987; 200 Ruppin H, Bar-Meir S, Soergel KH, Wood CM, Schmitt MG Jr. 67: 1143-84. Absorption of short chain fatty acids by the human colon. 166 Welsh MJ, Liedtke CM. Chloride and potassium channels in cystic Gastroenierology 1980; 78: 1500-7. fibrosis airway epithelia. Nature 1986; 322: 467-70. 201 Argenzio RA, Whipp SC. Inter-relationship of sodium, chloride, 167 Shoumacher RA, Shoemaker RL, Halm DR, Tallant EA, Wallace RW, bicarbonate and acetate transport by the colon of the pig. J7 Phvsiol Frizzell RA. Phosphorylation fails to activate chloride channels from 1979; 295: 365-81. cystic fibrosis airway cells. Nature 1987; 330: 752-4. 202 Sellin JH, DeSoigne R. Short chain fatty acid absorption in rabbit colon 168 Taylor CJ, Baxter PS, Hardcastle J, Hardcastle PT. Failure to induce in vitro. Gastroenterology 1990; 99: 676-83. secretion in jejunal biopsies from children with cystic fibrosis. Gut 203 Umesaki Y, Yajima T, Yokokura r, Mutal M. Effect of organic acid 1988; 29: 957-62. absorption on bicarbonate transport in the rat colon. I'flugers Arch 169 Gaskin KJ, Durie PR, Carey M, Wei P, Forster GG. Evidence for 1979; 379: 43-7. a primarv defect of pancreatic HCO3 secretion in cystic fibrosis. 204 Harig JM, Soergel KH, Barry JA, Ramaswamv K. Transport of Pediatr Res 1982; 16: 554-7. propionate by human ileal brush border membrane vesicles. 170 Dunk CR, Brown CDA. Turnberg LA. Regulation of Cl/HCO3 AmJPhvsiot 1991; 260: G776-82. exchange in rat duodenal brush border vesicles by cAMP. Pflugers 205 Mascolo N, Rajendran VM, Binder HJ. Mechanism of short-chain Arch 1989; 414: 701-5. fatty acid uptake by apical membrane vesicles of rat distal colon. 171 Sundaram U, Knickelbein R, Dobbins JW. Mechanisms of cAMP Gastroenterology 1991; 101: 331-8. mediated ileal secretion. Gastroenterology 1991; 100: A705. 206 Sellub J, Rosenberg IH. Folate transport in isolated brush border 172 Manganel M, Turner RJ. Rapid secretagogue induced activation of vesicles from rat intestine.J Biol Chem 1985; 256: 4489-93. Na+/H + exchange in rat parotid acinar cells. J Biol Chem 1991; 266: 207 Ganapathy V, Leibech L. Role of pH gradient and membrane potential

10182-8. in dipeptide transport in intestinal and renal brush border membrane http://gut.bmj.com/ 173 Buanes T, Grotmol T, Landsverh T, Raeder MG. Secretin empties bile vesicles from the rabbit..7 Biol Chem 1986; 258: 14189-92. duct cell cytoplasm of vesicles when it initiated ductular HCO3- 208 Kitis G, Lucas ML, Bishop H, et al. Altered jejunal surface pH in coeliac secretion in the pig. Gastroenterology 1988; 95: 417-24. disease: its effect on propranalol and folic acid absorption. Clin Sci 174 Dumont M, Erlinger S, Uchman S. Hypercholeresis induced by 1982; 63: 373-80. ursodeoxycholic acid and 7-ketolithocholic acid in the rat: possible 209 Hoffmann EK, Simonsen LO. Membrane mechanisnis in volume and role of bicarbonate transport. Gastroenterology 1980; 79: 82-9. .pH regulation in vertebrate cells. Phvsiol Rev 1989; 69: 315-82. 175 Yoon YB, Hagey LR, Hofmann AF, Guranz D, Michelotti EL, 210 Erikson AE, Spring KR. Volume regulation by Nectarus gallbladder: Steinbach JH. Effect of side chain shortening on the physiological apical Na+/H+ and Cl-/HCO3- exchange. Am3r Phvsiol 1982; 243: properties of bile acids: hepatic transport and effect on biliary C146-50. secretion pf 23-nor-ttrsodeoxycholate in rodents. Gastroenterology 211 Herbert SC. Hypertonic cell volume regulation in mouse thick limbs. II. 1986; 90: 837-52. Na+/H+ and Cl-/HCO3- exchange in basolateral membranes. on September 24, 2021 by guest. Protected copyright. 176 Palmer KR, Gurantz D, Hofmann AF, Clayton LM, Hagey LR, AmJ Physiol 1986; 250: C920-3 1. CeLcchetti S. Hypercholeresis induced by nor-chenodeoxycholate in 212 Corasanti 1G, Gleeson D, Boyer JL. Effects of osmotic stresses on the biliary fistula rodent. Amj Physiol 1987; 252: G219-28. isolated rat hepatocytes I. Ionic mechanisms ofcell volume regulation. 177 Knyrim K, Vakil N, Pfab R, Classen M. The effects of intraduodenal AmJ' Phvsiol 1990; 258: G290-8. bile acid administration on biliary secretion of ionized calcium and 213 Grinstein S, Rothstein A, Sarkadi B, Gelfand EW. Responses of carbonate in man. Hepatology 1989; 10: 134-42. lymphocytes to anisotonic media: volume regulating behaviour. 178 de Caestecker JS, Jazrawi RP, Petroni ML, Northfield TC. AmJPhvsiotl 1984; 246: C204-15. Ursodeoxycholic acid in chronic liver disease. Gut 1991; 32: 1061-5. 214 Gleeson D, Corasanti JG, Boyer JL. Effects of osmotic stresses on 179 Gleeson D, Ruppin DC, Murphy GM, Dowling RH. Final outcome of isolated rat hepatocytes II. Modulation of intracellular pH. ursodeoxycholic acid treatment in 126 patients with radiolucent Am3' Phvsiotl 1990; 258: G299-307. gallstones. QJ Med 1990; 229: 711-29. 215 McLeod RI, Hamilton JR. Regulatory solume increase in mammalian 180 StrazzaboscoM, Sakisaka S, HayakawaT, Boyer JL. Effect of UDCA on jejunal villus cells is due to bumetanide sensitive NaKC12 intracellular and biliary pH in isolated rat hepatocyte couplets and cotransport. AmJ7 Phvsiol 1990; 258: G665-74. perfused liver. Amj Physiol 1991; 260: G58-69. 216 McLeod RI, Hamilton JR. Volume regulation initiated by Na+- 181 Guranz D, Schteingartt CD, Hagey LR, Steinbach JH, Grotmol T, nutrient cotransport in isolated mammalian villus enterocytes. Hofmann AF. Hypercholeresis induced by unconjugated bile acid AmJ7 Physiol 1991; 260: G26-33. infusion correlated with recovery in bile of conjugated bile acids. 217 Moolenaar WH. Effect of growth factors on intracellular pH regulation. Hepatology 1991; 13: 540-50. Ann Rev Ptvsiol 1986; 48: 363-75. 182 LakeJR, Van DykeRW, Scharschmidt BF. EffectsofNa+ replacement 218 Bierman AJ, Cragoe EJ Jr, deLaat SW, Moolenaar WH. Bicarbonate and amiloride on ursodeoxycholic acid stimulated choleresis and determines cytoplasmic pH and suppresses mitogen induced biliary bicarbonate secretion. Amj Physiol 1987; 252: G163-9. alkalinisation in fibroblastic cells.J7 Biol Chem 1988; 263: 15253-6. 183 Hogben CAM, Tocco DJ, Brodie BB, Schanker LS. On the mechanism 219 Ganz MB, Boyarski G, Sterzel RB, Boron WF. Argenine vasopressin of intestinal absorption of drugs. J Pharmacol Exp Ther 1959; 125: enhancess pHi regulation in the presence of HCO3- by stimulating 275-82. three acid-base transport systems. Naiure 1989; 337: 648-51. 184 Rechkemmer G. Transport of weak electrolytes. In: Schultz SG, Field 220 Perona R, Serrano R. Increased pH and tumourgenicity of fibroblasts M, Frizzell RA, eds. Handbook of physiology. Section 6. The expressing a yeast proton pump. Naiure 1988; 334: 438-40. gastrointestinal system. Vol IV. Intestinal absorption and secretion. 221 Johnson LR. Regulation ofgastrointestinal mucosal growth. Phvsiol Rev New York: Oxford University Press, 1991: 371-88. 1988; 68: 456-502. 185 Lucas ML, Blair JA. The relationship of the acid microclimate in 222 Goodrich AL, Suchy FJ. Na+/H+ exchange in basolateral plasma proximal jejunum and its relationship to luminal acidification. membrane vesicles from neonatal rat liver. Am J Phvsiol 1990; 259: Proc R Soc Lond 1975; 200: 27-41. G334-9. 186 Lucas ML, Kei FH, Blair1IA. The influence of buffer pH, glucos and 223 Bastie MJ, Delvaux M, Dufresne M, Saunier-Blache IS, Vavsse N, sodium ion concentratiott on the acid microclimate of rat proximal Ribet A. Distinct activation oif Na/H exchange by gastrin and CCK jejunum in vitro. Pflugers Arch 1980; 385: 137-42. peptide in acini from guinea pig. AmJ7 Phvsiot 1988; 254: G25-32. 187 Shiau YF, Fernandez P, Jackson MI, McMonagle S. Mechanisms 224 Sacks Al, Acra 5, Dykes W, Polk B, Barnard J, Ghishan FK. The: ileal maintaining a low-pH microclimate in the intestine. Am J Phyvsiol Na/H exchanger is upregulated by small intestine resection in the 1985; 248: G6()8-17. weanling rat. Gasiroenierotogy 1991; 100: A701. 188 Lucas ML, Cooper BT, Lei FH, Jo)hnson IT, Holmes GKT, Blair IA, 225 Brasitus TA, Dudeja PK, Foster ES. 1,2 dimethylhydrazine induced et al. Acid microclimate in coeliac and Crohns disease: a model for alteratiomns in Na+/H+ exchange in rat colonic brush bordfer mem- folate malabsorption. Gut 1978; 19: 735-42. brane vesicles. Biochim Biophvs Acita 1988; 938: 483-8. 189 Shimada T. Factors affecting the microclimate pH in rat jejunum. 226 Corasanti JG, Deinhart IA. Na+/H + exchange actisity in Hr1-29 colon 7JPhysiol 1987; 392: 113-27. carcinoma cells saries as a function osf differentiation. Gtasiro¢enieroto>gy 190 MclEwan GiTA, Daniel H, Fett C, Burgess MN, Lucas ML. I'he effect 1990; 98: A539. Acid-base transport systems in gastrointestinal epithelia 1145

227 Corasanti JG, Deinhart JA, Grinciro A. Sodium butyrate down regulates tion in AR42J pancreatic cell line. Am]7 PhYsiol 1990; 259: (842-9. Na+/H+ exchange activity in T-84 colon carcinoma cells. 230 Montrose IH, Murer H. Polarity and kinetics of Na t /H t exchangc in Gastroenierology 1991; 100: A683. cultured opossum kidney cells. ,m]7 /hv-vsi'ol 1990; 258: Cl 21-33. 228 Ulrich-Baker MG, Wang P, Fitzpatrick L, Johnson LR. Amiloride 231 Carter KJ, Saario 1, Seidler U, Silen W. Effect of pCO2 on intracellullar inhibits rat mucosal ornithine decarboxvlase activity and DNA pH in in sitro frog gastric mucosa. Amjt Pht'siol 1999; 256: G2()6- 13. Gut: first published as 10.1136/gut.33.8.1134 on 1 August 1992. Downloaded from synthesis. AmJ7 Phvsiol 1988; 254: G408-15. 232 Chow A, Dobbins JWX, Aronson PS, Igarashi 1. Cloning of a rabbit ileal 229 Delvaux M, Bastie MJ, Chentoufi J, Cragoe EJ Jr, Vaysse N, Ribet A. band 3 related protein, a member of the anion exchangcr gene tamily. Amiloride and its analogues inhibit Na/H exchange and cell prolifera- Gastroenterologv 1991; 100: A682. http://gut.bmj.com/ on September 24, 2021 by guest. Protected copyright.