SUMMARY1. 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 Jrm 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 J. 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.

Northern ungulates acclimatize to winter conditions with restricted food supply and unfavorable weather conditions by reducing energy expenditure and voluntary food intake. We investigated in a study on red deer whether rates of peptide and glucose transport in the small intestines are also reduced during winter as part of the thrifty phenotype of winter-acclimatized animals, or whether transport rates are increased during winter in order to exploit poor forage more efficiently. Our results support the latter hypothesis. We found in a feeding experiment that total energy intake was considerably lower during winter despite ad libitum feeding. Together with reduced food intake, mass of visceral organs was significantly lower and body fat reserves were used as metabolic fuel in addition to food. However, efficacy of nutrient absorption seemed to be increased simultaneously. Extraction of crude protein from forage was higher in winter animals, at any level of crude protein intake, as indicated by the lower concentration of crude protein in feces. In line with these in vivo results, Ussing chamber experiments revealed greater electrogenic responses to both peptides and glucose in the small intestines of winter-acclimatized animals, and peptide uptake into jejunal brush-border membrane vesicles was increased. We conclude that reduced appetite of red deer during winter avoids energy expenditure for unproductive search of scarcely available food and further renders the energetically costly maintenance of a large gut and visceral organs unnecessary. Nevertheless, extraction of nutrients from forage is more efficient in the winter to attenuate an inevitably negative energy balance.

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 (J") was higher than mucosal-to-serosal flux(Jlr ), resulting in a net secretory flux of propionate.2. HCO37-CO2-free solution reduced JPr in the caecum and proximal colon markedly; JPr was not (caecum) or little (proximal colon) affected. The subsequent addition of acetazolamide caused a further decrease in Jlr in the proximal colon, but not in the caecum.3. In HC03--containing solutions acetazolamide or ethoxzolamide inhibited Jlr ; Jlr 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 Jlr in the caecum and proximal colon; jPr was not affected. 5.Results support the view that a considerable proportion of short-chain fatty acids (SCFAs) is absorbed via a SCFA--HC03-exchange.

The probiotics Saccharomyces boulardii and Bacillus cereus var. toyoi are nonpathogenic microbes which have been shown to affect certain functions of the mucosal barrier in pig jejunum such as electrogenic ion transport capacity and paracellular permeability. The present studies were performed to investigate potential effects of the probiotics on jejunal nutrient transport such as sodium‐dependent glucose transport or proton‐dependent dipeptide transport. For this purpose the in vitro Ussing‐chamber technique was applied in order to examine net electrogenic ion flux rates (short circuit currents, Isc) across isolated intact jejunal epithelia in the absence and presence of either 10 mmol/l glucose (mucosal side) or two‐fold application of 5 mmol/l glycyl‐ l‐sarcosine or glycyl‐ l‐glutamine to the mucosal bathing solution. Brush border membrane vesicles (BBMV) were prepared in order to characterize kinetic parameters (Vmax, Km) of Na‐dependent glucose transport. Intestinal tissues were obtained from growing pigs in a weight range between 23 and 33 kg. All animals were fed twice daily and received 0.8–0.9 kg/day of a standard diet. After a 9‐ to 10‐day adaptation period the diets for treated animals were either supplemented for 8 days with 1.7×107 colony‐forming units (CFU)/g feed of S. boulardii or for 3 weeks with 106 CFU/g feed B. cereus var. toyoi. Under basal conditions Isc values were not affected by different treatment protocols (controls: 0.74 ± 0.04 µeq/cm2 per h, n=9; S. boulardii: 0.74 ± 0.12 µeq/cm2 per h, n=7; B. cereus 0.68 ± 0.09 µeq/cm2 per h, n=5). Irrespective of dietary treatment, the addition of glucose resulted in significant increases of Isc indicating substantial onset of electrogenic net Na/glucose cotransport. Maximal Isc values occurred within 30 min and reached 2.79 ± 0.41 µeq/cm2 per h in control epithelia. This was significantly lower than found in S. boulardii (4.47 ± 0.43 µeq/cm2 per h, p < 0.05) and B. cereus var. toyoi tissues (4.45 ± 0.31 µeq/cm2 per h, p < 0.05). Gt values were 22.4 ± 1.3 mS/cm2 in control animals and were significantly lower as shown in S. boulardii (p < 0.01) and B. cereus var. toyoi (p < 0.01)‐treated animals (28.4 ± 1.3 and 29.9 ± 0.8 mS/cm2, respectively). Vmax values of Na‐dependent glucose uptake into BBMV differed significantly between controls (0.64 ± 0.08 nmol/mg protein per 10 s; n=5), S. boulardii (0.89 ± 0.06 nmol/mg protein per 10 s; n=5, p < 0.05) and B. cereus var. toyoi preparations (1.08 ± 0.05 nmol/mg protein per 10 s; n=3, p < 0.01). Km values were not significantly affected (control: 0.31 ± 0.04 mmol/l, S. boulardii: 0.29 ± 0.05 mmol/l, B. cereus var. toyoi: 0.21 ± 0.01 mmol/l). Irrespective of dietary treatment, application of the dipeptide model substances glycyl‐ l‐sarcosine or glycyl‐ l‐glutamine resulted in significant increases of Isc indicating marked stimulation of electrogenic net H+/dipeptide cotransport. Highest Isc responses occurred in B. cereus var. toyoi preparations and lowest were found in control tissues. H...

Infectious gastrointestinal diseases are frequently caused by toxins secreted by pathogens which may impair physiological functions of the intestines, for instance by cholera toxin or by heat-labile enterotoxin. To obtain a functional model of the human intestinal epithelium for studying toxin-induced disease mechanisms, differentiated enterocyte-like Caco-2 cells were co-cultured with goblet cell-like HT29-MTX cells. These co-cultures formed a functional epithelial barrier, as characterized by a high electrical resistance and the presence of physiological intestinal properties such as glucose transport and chloride secretion which could be demonstrated electrophysiologically and by measuring protein expression. When the tissues were exposed to cholera toxin or heat-labile enterotoxin in the Ussing chamber, cholera toxin incubation resulted in an increase in short-circuit currents, indicating an increase in apical chloride secretion. This is in line with typical cholera toxin-induced secretory diarrhea in humans, while heat-labile enterotoxin only showed an increase in short-circuit-current in Caco-2 cells. This study characterizes for the first time the simultaneous measurement of physiological properties on a functional and structural level combined with the epithelial responses to bacterial toxins. In conclusion, using this model, physiological responses of the intestine to bacterial toxins can be investigated and characterized. Therefore, this model can serve as an alternative to the use of laboratory animals for characterizing pathophysiological mechanisms of enterotoxins at the intestinal level.