Short-chain Fatty Acid Transport Research Articles

Mechanism of n-butyrate uptake in the human proximal colonic basolateral membranes.

Current studies were undertaken to characterize the mechanism of short-chain fatty acid (SCFA) transport in isolated human proximal colonic basolateral membrane vesicles (BLMV) utilizing a rapid-filtration n-[(14)C]butyrate uptake technique. Human colonic tissues were obtained from mucosal scrapings from organ donor proximal colons. Our results, consistent with the existence of a HCO(3)(-)/SCFA exchanger in these membranes, are summarized as follows: 1) n-[(14)C]butyrate influx was significantly stimulated into the vesicles in the presence of an outwardly directed HCO(3)(-) and an inwardly directed pH gradient; 2) n-[(14)C]butyrate uptake was markedly inhibited (approximately 40%) by anion exchange inhibitor niflumic acid (1 mM), but SITS and DIDS (5 mM) had no effect; 3) structural analogs e.g., acetate and propionate, significantly inhibited uptake of HCO(3)(-) and pH-gradient-driven n-[(14)C]butyrate; 4) n-[(14)C]butyrate uptake was saturable with a K(m) for butyrate of 17.5 +/- 4.5 mM and a V(max) of 20.9 +/- 1.2 nmol x mg protein(-1) x 5 s(-1); 5) n-[(14)C]butyrate influx into the vesicles demonstrated a transstimulation phenomenon; and 6) intravesicular or extravesicular Cl(-) did not alter the anion-stimulated n-[(14)C]butyrate uptake. Our results indicate the presence of a carrier-mediated HCO(3)(-)/SCFA exchanger on the human colonic basolateral membrane, which appears to be distinct from the previously described anion exchangers in the membranes of colonic epithelia.

N-Butyrate rectal transport in cirrhotic patients.

Data about colonic mucosa transport of short-chain fatty acids in cirrhotic patients are still lacking. The aim of the present study was to compare the rectal mucosa transport of n-butyrate and its effect on transport of other electrolytes and endoluminal pH in normal subjects and in cirrhotic patients by using a rectal dialysis technique. Thirteen subjects with normal hepatic function tests and 17 cirrhotic patients were enrolled. Dialysis bags containing 80 mmol/liter of butyrate in a neutral pH (6.8) electrolyte solution were placed in the rectum of enrolled subjects for 60 min. Net transport rate was calculated for butyrate, sodium, chloride, potassium, and bicarbonate. The differences in pH between initial and final dialysis solutions was also evaluated in the two groups in the study. Net butyrate absorption was significantly lower in cirrhotic patients than in controls (65.2 +/- 38.6 vs 101.2 +/- 45.3 nmol/min/cm2, respectively; P = 0.02). Furthermore, cirrhotic patients showed a lower HCO3 secretion than controls (-26.9 +/- 19.9 vs -45.1 +/- 20.0, respectively; P = 0.01). No differences were found in transport of the other electrolytes. The pH in the final dialysis solution in cirrhotic patients was not significantly lower than in the controls (7.15 vs 7.35; P = 0.1). In conclusion, the impairment of butyrate absorption and the concurrent reduction of bicarbonate secretion observed in cirrhotic patients may suggest a selective hypoactivity of apical HCO3-/SCFA- antiport located at the colonocyte apical membrane.

Short-chain fatty acid (SCFA) uptake into Caco-2 cells by a pH-dependent and carrier mediated transport mechanism.

The short-chain fatty acids, acetate, propionate, and butyrate, are the most abundant organic anions in the human colon. SCFA play a pivotal role in maintaining homeostasis in the colon. Particularly butyrate induces cell differentiation and regulates growth and proliferation of colonic mucosal epithelial cells, whereas it reduces the growth rate of colorectal cancer cell. Previous studies by several groups, including our own, using isolated membrane vesicles have demonstrated that the uptake of butyrate is at least in part mediated by a non-electrogenic SCFA-/HCO3- antiporter. The purpose of the present study was to determine (1) whether Caco-2 cells could serve as an experimental model to assess the mechanisms of SCFA transport, and (2) whether monocarboxlate transporters could play a role in SCFA transport in these cells. Caco-2 cells were found to transport 14C-butyrate in a concentration and time dependent manner. The uptake was sodium independent, but was stimulated by lowering extracellular pH. The uptake of 500 microM butyrate was reduced by 49.6% +/- 3.3% in the presence of propionate and by 57.2% +/- 4.8% in the presence of 10 mM L-lactate. The addition of 1 mM alpha-cyano-4-hydroxycinnamate and phloretin, both known to be potent inhibitors of MCT1, decreased the uptake of 500 microM 14C-butyrate by 59.4% +/- 4.1% and 48.9% +/- 3.3%, respectively, whereas similar concentrations of DIDS did not have any effect. These data suggest that the uptake of butyrate in Caco-2 cells occurs via a carrier mediated transport system specific for monocarboxylic acids, which is in accordance with characteristics of the MCT 1.

Feed-induced Changes in the Transport of Butyrate, Sodium and Chloride Ions Across the Isolated Bovine Rumen Epithelium

A short-chain fatty acid (SCFA) burst feeding strategy to dry cows (periodically high SCFA concentration in the ruminal fluid) has been hypothesized to increase the ruminal surface area and absorptive capacity. The present paper deals with the effects of a SCFA burst feeding strategy on epithelial transport function in dry cows. The epithelial transport capacity for [14C]butyrate and Na+ across the rumen epithelium in vitro was significantly increased by the SCFA burst feeding strategy, compared with the controls. The transport of Cl- was significantly increased by the SCFA burst feeding strategy at the high feeding level, but non-significantly decreased at the low feeding level. The increase in transport capacity could not be attributed to changes in epithelial resistance, to the concentration of Na+/K+-ATP-ase or to the epithelial surface area and structure (data on epithelial surface area and structure were published by Andersen et al., Report No. 23 from the National Institute of Animal Science, Denmark, 1994). The observed changes in transport must reflect a change in either composition of the cell membranes or processes that facilitate transport through the cell, including metabolic processes. The presented results support the existence of a saturable SCFA transport system in the epithelial cells and the results show that the capacity of this transport system can be the subject for feed-induced regulation.

Substrate and Inhibitor Specificity of Butyrate Uptake in Apical Membrane Vesicles of the Rat Distal Colon

Background/Aims: There is substantial evidence that transcellular flux of short-chain fatty acid (SCFA) absorption, at least in part, is mediated by an anion exchange process with bicarbonate. Methods: This anion exchange system was further characterized in apical membrane vesicles of the rat distal colon by studying substrate and inhibitor specificities of a variety of substituted monocarboxylic acids as well as of known inhibitors of the recently described monocarboxylate transporter MCT1 and MCT2. Results: SCFA transport was significantly reduced in the presence of branched and unbranched SCFAs and several bromo, chloro and mercapto analogues as well as nicotinic acid and L-lactate. In contrast, known inhibitors of monocarboxylate transporter proteins like stilbene derivatives, phloretin and 2-cyano-4-hydroxycinnamate did not inhibit bicarbonate-gradient stimulated butyrate transport. Kinetic analysis of increasing substrate concentrations of 3-mercaptopropionate, L-lactate and nicotinic acid showed saturation kinetics with apparent K_i of 6.1, 18.3 and 14.7 mmol/l, respectively. Conclusions: The data not only confirm earlier results that absorption of SCFAs in apical membranes of the rat distal colon is mediated by a relatively low affinity/high capacity SCFA^–/HCO^–₃ exchange mechanism, but also indicate that although this anion transporter shares some functional similarities, is not identical with the recently cloned MCT isoforms.

Ruminal transport and metabolism of short-chain fatty acids (SCFA) in vitro: effect of SCFA chain length and pH

The unidirectional transport and metabolism of 14C-labeled acetate, propionate and butyrate across the isolated bovine rumen epithelium was measured in vitro by the Ussing chamber technique. There was a significant, but relatively small, net secretion of acetate and propionate, and a large and significant net absorption of butyrate. The results demonstrate that the mucosal–serosal (MS) pathway for short-chain fatty acids (SCFA) is different from the serosal–mucosal (SM) pathway, and that butyrate is treated differently from acetate and propionate by the epithelium. The results support that the main route for epithelial SCFA transport is transcellular. The correlation between SCFA lipophility and the flux rate was positive but weak at both pH 7.3 and 6.0. Decreasing pH increased all SCFA fluxes significantly, but not proportionally to the increase of protonized SCFA in the bathing solution. There was a significant and apparently non-competitive interaction between the transport of acetate, propionate and butyrate. It seems that mediated transport mechanisms must be involved in epithelial SCFA transport in the bovine rumen, but the data do not exclude that passive diffusion could account for a significant part of the flux. The metabolism of SCFA in the Ussing chamber system was considerable, and there was a clear preference for excretion of CO 2 from this metabolism to the mucosal side, while side preference for non-CO 2 metabolite excretion was not studied. Of the propionate and butyrate transported in the MS direction, 78 and 95% was metabolised, while only 37 and 38% was metabolised in the SM direction (acetate metabolism could not be measured). There was, however, no simple relation between the degree of metabolism and the transport rate or the transport asymmetry of the SCFA.

Metabolism of short-chain fatty acids by rat colonic mucosa in vivo.

To determine the influence of substrate concentration and substrate interactions on short-chain fatty acid metabolism in vivo, a surgical procedure was established. Rats were surgically operated to cannulate a 5-cm segment of proximal colon, isolate the vasculature, and cannulate the right colic vein draining this segment. Thus metabolism was restricted to the defined colonic segment. The appearance of total (14)C and (14)CO(2) in the mesenteric blood stabilized after 30 min of perfusion. Increasing luminal concentrations of butyrate from 2 to 40 mmol/l resulted in linear increases in total (14)C, but (14)CO(2) production from [(14)C]butyrate increased as a function of concentration only up to 10 mmol/l and was stable at higher butyrate concentrations. In addition to CO(2), 3-hydroxybutyrate and lactate were major metabolites of acetate and butyrate in vivo. The presence of a mixture of alternative substrates in the lumen had no influence on the metabolism of butyrate to CO(2) but significantly reduced the metabolism of acetate to CO(2). When compared with young (4 mo old) animals, transport of butyrate was significantly lower for aged (48 mo old) animals, as evidenced by the rate of appearance in blood of total (14)C (P = 0.04) and (14)C in butyrate (P = 0.03), but metabolism was similar, since differences were not significant for (14)C in the major metabolites 3-hydroxybutyrate (P = 0.06) and CO(2) (P = 0.17). These results show that important aspects of short-chain fatty acid transport and metabolism are not predicted from data using isolated colonocytes but require study using an in vivo model.

Cellular SCFA content of colonic epithelial cells (HT29-C1) is dependent on regulation of intracellular and extracellular pH

Colonocyte uptake of short-chain fatty acids (SCFAs) is required for physiologic effects of these weak acids on cell physiology and transepithelial ion transport. We have proposed that changes in pH alter SCFA transport, but testing this model requires measurement of intracellular SCFA content, which has not been possible previously. The aim of this study was to use nuclear magnetic resonance (NMR) to directly measure SCFAs, and question whether changes in pH affected intracellular SCFA content. Methods. HT29-C1 cells grown on polystyrene beads were perfused while spectra were acquired on a GE Omega 400 NMR spectrometer. Interleaved non-diffusion weighted (single pulse acquisition) and diffusion weighted 1H NMR spectroscopy (recording the stimulated echo) was used to measure total SCFA, intracellular SCFA and intracellular water volume. SCFA concentration was estimated by integrating the 13-proton signal of iso-butyrate or the sum of the ct, 13, and /-proton peaks of n-butyrate. Correlative experiments used confocal microscopy of cells loaded with SNARF-1 to measure intracellular pH of HT29-C1 cells on beads. Results. NMR spectra were recorded during exposure of cells to SCFA superfusates containing 130mM Na-SCFA substituted mol:mol for NaC1 in isosmotic medium without added bicarbonate/CO s. Concomitant with detection of SCFA in the perfusion buffer, intracellular iso-butyrate or n-butyrate was detected. After extracellular SCFA levels reached steady state, intracellular SCFA content and cell volume continued to slowly increase. We hypothesized that the slow uptake was due to intracellular pH alkalinization and subsequent SCFA redistribution via nonionic diffusion. Amiloride (1 mM) inhibited intracellular SCFA accumulation by 53%, suggesting that the intracellular alkalinization mediated by Na/H exchange was important for regulating cellular SCFA content. Conversely, acidifying perfusates from pH 7.4 to 7.0 stimulated the accumulation of iso-butyrate (35% increase), consistent with increased SCFA uptake by nonionic diffusion. Conclusions. (1) Proton NMR can be used to report on intracellular SCFA content in substrate-attached epithelial cells. (2) SCFA content (and presumably transepithelial SCFA transport) is regulated by pH. This research was funded by the NIH.

Nonionic diffusion of short-chain fatty acids across rat colon.

Short-chain fatty acid (SCFA) transport across the colon may occur by nonionic diffusion and/or via apical membrane SCFA-/HCO3- exchange. To examine the relative importance of these processes, stripped segments of rat (Ratus ratus) proximal and distal colon were studied in Ussing chambers, and the unidirectional fluxes of radiolabeled SCFA butyrate, propionate, or weakly metabolized isobutyrate were measured. In N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) or 1 or 5 mM HCO3- Ringer, decreases in mucosal pH stimulated mucosal-to-serosal flux (Jm-->s) of all SCFA, decreases in serosal pH stimulated serosal-to-mucosal flux (Js-->m), and bilateral pH decreases stimulated both fluxes equally. These effects were observed whether the SCFA was present on one or both sides of the tissue, in both proximal and distal colon, in the absence of luminal Na+, and in the presence of either luminal or serosal ouabain. Changes in intracellular pH or intracellular [HCO3-] did not account for the effects of extracellular pH. Luminal Cl- removal, to evaluate the role of apical membrane Cl-/SCFA- exchange, had no effect on Jm-->s but decreased Js-->m 32% at pH 6.5 and 22% at 7.2. Increasing SCFA concentration from 1 to 100 mM, at pH 6.4 or 7.4, caused a linear increase in Jm-->s. We conclude that SCFA are mainly transported across the rat colon by nonionic diffusion.

In vitro Studies on Transport and Metabolism of Short-Chain Fatty Acids in Pig Hindgut

An analytical method based on alkaline freeze drying, ultracentrifugation, and quantitative gas chromatography was established to differentiate between mucosal uptake, tissue accumulation, and serosal release of SCFA in pig hindgut. It was shown that serosal release of SCFA was substantially lower than mucosal uptake and tissue accumulation, indicating substantial degradation and/or metabolism during transepithelial movement.

Transport of propionate across the distal colonic epithelium of guinea pig in the presence and absence of bicarbonate and of chloride.

Short-chain fatty acids (SCFA) are rapidly absorbed in the large intestine. However, the mechanisms involved have not been fully delineated. Findings indicate that absorption of SCFA occurs in the undissociated form as well as by an anion exchange, whereby marked species and segmental differences are present. The guinea pig distal colon has certain peculiarities. Unidirectional fluxes of propionate across guinea pig distal colon were studied under short-circuit current conditions in Ussing chambers. Removal of bicarbonate caused reduction of mucosal-to-serosal fluxes by 30%, serosal-to-mucosal fluxes were little affected. In chloride-free solution unidirectional fluxes were also reduced. However, in the presence of transepithelial chloride-gradients with the Cl(-)-free solution only at the luminal side, no such effects were seen. Findings support the presence of SCFA(-)-HCO3- exchange in guinea pig distal colon. Chloride seems not of major importance for SCFA transport.

Transport of n-butyrate into human colonic luminal membrane vesicles.

Human colonic short-chain fatty acid (SCFA) absorption is associated with increased luminal pH and HCO3- and enhanced Na+ absorption. Therefore, the mechanism of colonic SCFA transport, its dependence on Na+ and HCO3- and interactions with Cl-/HCO3- and Na+/H+ exchangers were characterized. Luminal membrane vesicles (LMV) isolated by divalent cation precipitation from organ donor colons were used for n-butyrate transport. Uptake of n-butyrate into the human colonic LMV was minimal even in the presence of an inward pH gradient, but an outward HCO3- gradient significantly increased uptake rates. HCO3(-)-stimulated butyrate uptake was saturable with an apparent Michaelis constant of 1.5 +/- 0.2 mM and maximal velocity of 105 +/- 3 nmol.mg protein-1.3 s-1. Intravesicular butyrate resulted in trans-stimulation of n-[1-14C]butyrate uptake. Butyrate uptake was inhibited approximately 25-40% by C2-C5 SCFAs and approximately 40% by niflumic acid. Butyrate uptake was not affected by extravesicular Na+, and 22Na uptake was unaltered by extravesicular butyrate. Butyrate uptake was independent of extra- or intravesicular CI-, and butyrate loading produced no changes in 36Cl uptake. We conclude that the predominant mechanism of n-butyrate transport across the human colonic luminal membrane appears to be via a HCO3-/SCFA antiport system independent of Cl-/HCO3- exchange and Na+ transport.

An Na(+)-independent short-chain fatty acid transporter contributes to intracellular pH regulation in murine colonocytes.

Short-chain fatty acids (SCFAs) are the major anions in the colonic lumen. Experiments studied how intracellular pH (pHi) of isolated colonocytes was affected by exposure to SCFAs normally found in the colon. Isolated crypt fragments were loaded with SNARF-1 (a fluorescent dye with pH-sensitive excitation and emission spectra) and studied in a digital imaging microscope. Intracellular pH was measured in individual colonocytes as the ratio of fluorescence intensity in response to alternating excitation wavelengths (575/505 nm). After exposure to 65 mM acetate, propionate, n-butyrate, or iso-butyrate in isosmotic Na(+)-free media (substituted with tetramethylammonia), all colonocytes acidified rapidly and then > 90% demonstrated a pHi alkalinization (Na(+)-independent pHi recovery). Upon subsequent removal of the SCFA, pHi alkalinized beyond the starting pHi (a pHi overshoot). Using propionate as a test SCFA, experiments demonstrate that the acidification and pHi overshoot are explained by transmembrane influx and efflux of nonionized SCFA, respectively. The basis for the pHi overshoot is shown to be accumulation of propionate during pHi alkalinization. The Na(+)-independent pHi recovery (a) demonstrates saturable propionate activation kinetics; (b) demonstrates substrate specificity for unmodified aliphatic carbon chains; (c) occurs after exposure to SCFAs of widely different metabolic activity, (d) is electroneutral; and (e) is not inhibited by changes in the K+ gradient, Cl- gradient or addition of the anion transport inhibitors DIDS (1 mM), SITS (1 mM), alpha-cyano-4-hydroxycinnamate (4 mM), or probenicid (1 mM). Results suggest that most mouse colonocytes have a previously unreported SCFA transporter which mediates Na(+)-independent pHi recovery.

In vivo colonic electrolyte flux in dogs fed soyprotein, casein or meat as their primary protein source

Fifteen adult, mongrel dogs were used in a study to assess the effects of dietary protein on canine colonic electrolyte transport. The dietary protein sources used in the study were soyprotein, casein and meat (i.e., freeze-dried beef muscle). The experiment was a completely random allocation of treatmens, with dogs as the experimental unit. Each animal was adapted to their respective diet for a minimum of 14 days prior to beginning the laboratory phase of the study. The colon of each animal was then cleansed and perfused with a known test solution to assess the apparent absorption/secretion of sodium, chloride and short chain fatty acids (SCFA) in the intact animal. The net flux of total and individual SCFA was affected by dietary treatment. Soyprotein fed dogs had greater total SCFA and individual SCFA (acetate, propionate & butyrate) absorption than did the meat fed animals. Transport of total and individual SCFA for the casein fed dogs was generally between that of the soyprotein and meat fed animals. Casein fed dogs absorbed significantly more acetate and propionate than the meat fed, and significantly less proionate and butyrate than the soyprotein fed animals. Sodium and chloride transport was not affected by protein source.

Functional role of bicarbonate in propionate transport across guinea-pig isolated caecum and proximal colon.

1. Unidirectional fluxes of propionate across isolated epithelia from the guinea-pig caecum and proximal colon were measured under short-circuit current conditions. In the caecum and proximal colon the serosal-to-mucosal propionate flux (JPrsm) was higher than mucosal-to-serosal flux (JPrms), resulting in a net secretory flux of propionate. 2. HCO3(-)-CO2-free solution reduced JPrms in the caecum and proximal colon markedly; JPrsm was not (caecum) or little (proximal colon) affected. The subsequent addition of acetazolamide caused a further decrease in JPrms in the proximal colon, but not in the caecum. 3. In HCO3(-)-containing solutions acetazolamide or ethoxzolamide inhibited JPrms; JPrsm was not affected. A macromolecular carbonic anhydrase inhibitor, prontosil-dextran, had no effect on propionate fluxes, indicating that the intracellular carbonic anhydrase is of importance for short-chain fatty acid transport. 4. Subsequent to carbonic anhydrase inhibition, mucosal addition of amiloride caused a slight further decrease of JPrms in the caecum and proximal colon; JPrsm was not affected. 5. Results support the view that a considerable proportion of short-chain fatty acids (SCFAs) is absorbed via a SCFA(-)-HCO3- exchange.

Segmental differences in short-chain fatty acid transport in rabbit colon: effect of pH and Na.

Short-chain fatty acids (SCFAs) are the predominant luminal anion in the mammalian colon. Although they are rapidly absorbed in vivo, little is known about the mechanisms of transepithelial transport in vitro. Previous studies have suggested that SCFA transport may be linked to Na absorption or an anion exchange mechanism. We compared the transport of propionate under short-circuit conditions in rabbit proximal and distal colon to determine whether there were segmental differences, how SCFAs may be linked to either Na absorption or anion transport, and whether SCFAs, as weak electrolytes, may be affected by transepithelial pH gradients. In distal colon, propionate transport was not significantly altered by stimulation of electrogenic Na absorption, epinephrine or Cl removal. However, a modest transepithelial pH gradient (luminal 6.8/serosal 7.4) stimulated propionate absorption. In proximal colon, propionate transport was significantly altered by maneuvers that either stimulated (lowered [Na] in the bathing media) or inhibited (theophylline) apical Na-H exchange. Neither Cl removal, nor the anion exchange inhibitor DIDS, nor a transepithelial bicarbonate gradient, altered propionate transport. A transepithelial pH gradient inhibited propionate secretion, but not in a manner entirely consistent with the effect of pH on the distribution of a weak electrolyte. These results suggest that there is significant segmental heterogeneity in colonic SCFA transport; that transepithelial propionate fluxes are altered by changes in pH or electroneutral Na absorption (Na-H exchange), but not by chloride removal, bicarbonate gradients or electrogenic Na absorption. Regulation of SCFA transport may be an important factor in the physiology of colonic fluid balance.

Bicarbonate-stimulated [14C]butyrate uptake in basolateral membrane vesicles of rat distal colon

Background: The mechanism of short-chain fatty acid (SCFA) absorption by the colon is not known. The aim of these experiments was to identify the transport mechanisms present in the basolateral membrane to develop an overall model of colonic SCFA absorption. Methods: These studies determined the uptake of [14C]-butyrate (used as a model SCFA) by basolateral membrane vesicles prepared from rat distal colonic mucosa. Results: Significantly higher [14C]butyrate uptake under an acidic environment (extravesicular pH [pH0] = intravesicular pH [pH1] = 5.5) than that under alkaline environment (pH0 = pH1 = 7.5) indicates the presence of nonionic diffusion. In the absence of a pH gradient (pH0/pH1 = 7.5/7.5), outward gradients of bicarbonate significantly stimulated [14C]butyrate uptake. Additional presence of a pH gradient (pH0/pH1 = 6.0/7.5) further enhanced the bicarbonate gradient-stimulated [14C]butyrate uptake that was not inhibited by voltage clamping but was inhibited substantially by an anion exchange inhibitor 4,4′-diisothiocyanatostilbene-2,2′disulfonic acid (DIDS) (82%). Both increasing extravesicular butyrate concentration and intravesicular bicarbonate concentration saturated bicarbonate/pH gradient-stimulated [14C]butyrate uptake with an apparent Michaelis constant (Km) for butyrate of 6.9 mmol/L and an apparent Km for bicarbonate of 27.4 mmol/L. Conclusions: Butyrate uptake by basolateral membrane vesicles represents both nonionic diffusion and a carrier-mediated SCFA-bicarbonate exchange process that differs from the SCFA-bicarbonate exchange recently identified in apical membrane vesicles. Thus, two distinct carrier-mediated anion exchange processes located in apical and basolateral membranes mediate transcellular SCFA transport in colonocytes.

Effects of amiloride and ouabain on short‐chain fatty acid transport in guinea‐pig large intestine.

1. Effects of H+ secreting mechanisms on unidirectional passage of the short-chain fatty acids (SCFA) acetate, propionate and n-butyrate across isolated guinea-pig caecum, proximal and distal colon were studied under short-circuit current conditions in Ussing chamber isotope flux experiments. 2. In the caecum and the proximal colon the serosal-to-mucosal fluxes (Jsm) were higher than the mucosal-to-serosal fluxes (Jms). Thus a net secretion of SCFA was present in the caecum and proximal colon. The higher Jsm appears to be coupled to the Na+ gradient established by the basolateral membrane Na(+)-K(+)-ATPase, whereas Jms is related to the operation of an apical membrane Na(+)-H+ exchanger. Inhibition of Na(+)-H+ exchange by amiloride (1 mM) added to the mucosal solution decreased Jms of SCFA in caecum and in proximal colon, but had no major effect in distal colon. 3. In distal colon Jms exceeds Jsm and thus a net absorption of SCFA was observed. Jms is Na+ independent and coupled to the activity of the apical membrane K(+)-H(+)-ATPase. Inhibition of the K(+)-H(+)-ATPase by addition of ouabain (0.1 mM) to the mucosal solution diminished Jms in the distal colon; in the caecum and proximal colon ouabain had no effect on Jms. 4. Neither amiloride nor ouabain caused major changes in Jsm in any of the large intestinal segments. 5. In conclusion, absorption of SCFA, e.g. Jms, in all large intestinal segments is related to the presence and activity of H+ secreting systems located in the apical membrane of colonocytes. In the caecum and proximal colon the predominant system appears to be Na(+)-H+ exchange, and in the distal colon K(+)-H(+)-ATPase. Supply of H+ ions allows protonation of SCFA anions and subsequent permeation by non-ionic diffusion across the apical membrane. These mechanisms account for 35, 40-50 and 60-80% of SCFA transport in the caecum, the proximal and the distal colon of guinea-pig, respectively. The nature of the Na(+)-dependent secretory pathway in the caecum and proximal colon remains to be determined.

Segmental differences of short-chain fatty acid transport across guinea-pig large intestine.

Unidirectional fluxes of short-chain fatty acids (SCFAs) were measured under short-circuit current conditions across guinea-pig caecum, proximal and distal colon. Fluxes increased linearly with concentration. In the caecum with equal concentrations on both sides of the mucosa the serosal-to-mucosal (sm) fluxes were nearly twice the mucosal-to-serosal (ms) fluxes for all SCFAs; in the distal colon ms fluxes were always higher than sm fluxes. Thus, in the caecum the net effect was secretion while in the distal colon net absorption occurred. In caecum, ms fluxes decreased with chain length while sm fluxes were similar for the three fatty acids, acetic, propionic and butyric acid. In the distal colon both unidirectional fluxes increased with chain length. Fluxes across the mucosa of the proximal colon were intermediate to those in caecum and distal colon. A paracellular transport of short-chain fatty acids is not present. The results indicate that other processes are involved in transcellular transport of SCFA besides non-ionic diffusion.

Transport of propionate by human ileal brush-border membrane vesicles

Human ileal brush-border membrane vesicles were employed to study the mechanisms of short-chain fatty acid (propionate) absorption especially to determine the effects of intravesicular HCO3- and the component of nonionic diffusion. Preloading the vesicles with HCO3- resulted in up to 20-fold "overshoots" of transport, and this effect was not seen with other intravesicular anions. This transport process was very fast (peak uptake 6 s) and was not due to intravesicular buffering by HCO3-. Radiolabeled propionate transport demonstrated transstimulation when the vesicles were preloaded with unlabeled propionate. An inward H+ gradient led to stimulation of propionate transport much smaller than in the presence of trans-HCO3-, whereas an inward Na+ gradient had no effect. Propionate transport was attenuated by the anion exchange inhibitors SITS and DIDS. Under HCO3- gradient conditions, propionate transport exhibited saturation kinetics with an apparent Km of 21 +/- 3 mM and a Vmax of 50 +/- 3 nmol.mg protein-1.3 s-1. Propionate transport was inhibited up to 40% by 2-5 carbon short-chain fatty acids (10 mM) but not by other organic anions. Short-chain fatty acid transport in the human ileum is Na+ independent and occurs mostly via a specific anion exchange mechanism with HCO3-. Our results also demonstrate a small component of nonionic diffusion of the protonated fatty acid (or anion exchange for OH-).