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NaCl absorption in the rabbit ileum. Effect of acid-base variables.

In vivo and in vitro studies suggest that acid-base variables regulate ion transport in the rabbit ileum. The relative importance of these variables on active Na+ and Cl- absorption has not been defined. Isolated, stripped ileal segments were studied under short-circuited conditions in the Ussing flux chamber. Unidirectional 22Na and 36Cl fluxes were measured after changes in bathing solution pH, PCO2, and/or [HCO3-]. When pH was decreased from 7.6 to 7.1, net flux of Na+ increased from 0.1 +/- 0.7 to 2.6 +/- 0.7 mu Eq/cm2 per hour and net flux of Cl- increased from -2.0 +/- 0.9 to 1.3 +/- 0.6 mu Eq/cm2 per hour. These changes were rapid in onset, completely reversible, and accounted for by changes in mucosal-to-serosal fluxes of these ions. They were accompanied by small decreases in short-circuit current, but there were no changes in residual flux. These pH effects were not inhibited by the presence of luminal bumetanide (1 mmol/L), furosemide (1 mmol/L), hydroflumethiazide (1 mmol/L), or 4,4'-diisothiocyanostilbene-2,2'-disulfonate (1 mmol/L), or by the carbonic anhydrase inhibitor methazolamide (1 mmol/L). When data from all combinations of acid-base conditions were combined and analyzed by linear regression, pH was the only variable that correlated with mucosal-to-serosal fluxes (r = -0.84) and net flux (r = -0.85) for Na+, mucosal-to-serosal fluxes (r = -0.96) and net flux (r = -0.99) for Cl-, and short-circuit current (r = 0.97). These findings suggest that extracellular pH modulates active Na+ and Cl- absorption in the rabbit ileum.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

Accuracy of cutaneous recordings of gastric electrical activity.

BACKGROUND: This study was undertaken to determine the accuracy of cutaneous electrogastrography (EGG). METHODS: The ability of EGG to assess gastric contractions and electrical frequency, phase lag, and waveform was studied in 4 healthy volunteers with cutaneous electrodes before and after eating, 4 healthy fasting volunteers with intraluminal pressure recording tubes and cutaneous electrodes, 1 patient with surgically implanted gastric serosal electrodes, and 4 anesthetized dogs with serosal force transducers and implanted and cutaneous electrodes. In 2 of the dogs, an intragastric distending balloon was introduced. RESULTS: The use of long-distance serosal electrodes allowed direct comparison of internal and cutaneous signals. Cutaneous electrodes recognized 80%-85% of changes in frequency with spectral analysis. Only 30%-40% of increases in EGG amplitudes were associated with gastric contractions. Gradual distention of the canine stomach after blocking contractions with atropine and glucagon increased the EGG amplitudes. No appreciable phase lag could be discerned with EGG. The descending portion of the EGG waveform was predominant. CONCLUSIONS: Frequency dynamics is currently the only reliable cutaneous EGG parameter.

Adult↗

Effect of endothelin 1 on ion transport in isolated rat colon.

BACKGROUND: Endothelin may play a significant role in the regulation of gastrointestinal function because it has a variety of biological activities and because endothelin-like immunoreactivity as well as its specific binding sites have been found in the gastrointestinal tract. This study investigated the secretory effect and mechanism of action of endothelin 1 in mammalian large intestine. METHODS: Distal colonic segments from Sprague-Dawley rats were stripped of their muscle layers and mounted in Ussing chambers. The effects of endothelin 1 on short-circuit current in rat colonic mucosa were studied in the absence or presence of specific inhibitors. Transmural unidirectional 22Na+ and 36Cl- fluxes and endothelin 1-induced prostacyclin release were also measured. RESULTS: Serosal addition of endothelin 1 evoked a sustained increase in short-circuit current that was significantly reduced by tetrodotoxin or atropine, and virtually abolished by a selective endothelin A receptor antagonist (BQ-123), furosemide, piroxicam, d,I-verapamil, or removal of serosal calcium. Hexamethonium, amiloride, diphenhydramine, or a specific platelet-activating factor antagonist (CV-6209) did not influence the response to endothelin 1. Endothelin 1 significantly decreased net sodium and net chloride absorption and induced a marked increase in prostacyclin release from the serosal surface of stripped colonic mucosa. CONCLUSIONS: Endothelin 1 has a secretory effect in rat colon. Its action seems to be mediated by cyclo-oxygenase products and enteric nerves via the activation of an endothelin A receptor.

Animals↗

HCO3- secretion by rat distal colon: effects of inhibitors and extracellular Na+.

BACKGROUND/AIMS: The large intestine secretes HCO3- via a Cl-/HCO3- exchange mechanism located in the apical membrane of colonocytes. However, an additional transport system(s) must facilitate HCO3- (OH-) entry or H+ exit across the basolateral cell surface. The aim of this study was to determine that mechanism(s). METHODS: A modified Ussing apparatus was used to measure net HCO3- secretion in segments of rat distal colon. RESULTS: When added to the serosal solution, 10 mmol/L 4-acetamido-4'-isothiocyano-2,2'-disulfonic acid stilbene (SITS), 1 mmol/L SITS and 0.1 mmol/L diisothiocyanostilbene-2,2'-disulfonic acid, inhibited HCO3- secretion by 88%, 51%, and 30%, respectively. However, the Na+/H+ exchange inhibitors, amiloride (1 mmol/L), dimethylamiloride (0.1 mmol/L), ethylisopropylamiloride (0.1 mmol/L), failed to affect HCO3- secretion. Acetazolamide (1 mmol/L) blocked HCO3- secretion by approximately 60% when in the serosal solution but had little effect when in the mucosal solution. Ion substitution studies showed that HCO3- secretion required Na+ in the serosal solution (K0.5 approximately 12 mmol/L). HCO3- secretion was unaffected by depolarizing the basolateral membrane potential with K(+)-rich medium. CONCLUSIONS: These data are consistent with Na+ linked HCO3- transport across the colonocyte basolateral membrane, which appears to be electroneutral.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

Protective effect of pentoxifylline on gastric mucosa.

Pentoxifylline (PF) has been shown to increase tissue oxygen tension. This study was performed to determine if PF has a protective effect on the gastric mucosa against alcohol (EtOH)-induced injury. Fasted Sprague-Dawley rats were pretreated with randomized test solution (control, normal saline, or PF, 75 mg/kg) intraperitoneally (ip). At 30 min, 100% EtOH (pH 8.5) was given intragastric. At 90 min, laparotomy was performed and gastric serosal stomach surface oxygen tensions (pO2) were measured. Stomachs were excised and opened and pH was measured. Photographs were taken and sections were obtained for histologic analysis to determine mucosal injury. The PF-pretreated rats had significantly higher serosal pO2 and significantly lower intragastric pH than control animals. There was significantly less gross and histologic mucosal injury in PF-treated animals. We conclude that PF is protective against EtOH gastric mucosal injury. This effect correlates with increased gastric serosal pO2 and is likely due to improved microcirculatory blood flow following PF administration.

Animals↗

Effect of a smooth muscle antagonist on contraction of patched intestinal defects.

Growing new intestinal mucosa on serosal patches may potentially increase intestinal surface area but is limited by contraction of the serosal patch. Since this might be related to smooth muscle contraction or altered collagen metabolism, our aim was to determine whether the smooth muscle antagonist thiphenamil inhibits contraction. Fifty rabbits had two 2 x 5-cm full-thickness intestinal defects patched with adjacent cecum. Group I (n = 25) received saline and Group II (n = 25) 0.02 M thiphenamil at 10 cc/hr intraluminally. Animals were sacrificed at 1, 3, 5, 7, and 10 days. Group II had significantly less contraction of the proximal patch until the 10th day after patching (84 +/- 8 vs 66 +/- 20% Day 1, 67 +/- 4 vs 52 +/- 9% Day 5, 42 +/- 14 vs 42 +/- 7% Day 10). Epithelialization of patches was significantly less in Group II animals at 10 days (88 +/- 8 and 86 +/- 11% vs 47 +/- 20 and 50 +/- 16%, P less than 0.05) but crypt cell production rate and villus height were similar. The hydroxyproline content of regenerating tissue increased significantly 7 and 10 days after patching but was similar in the two groups (30.8 +/- 5.9 micrograms/mg tissue Day 10 vs 12.8 +/- 2.8 Day 1). Smooth muscle antagonism by thiphenamil inhibited contraction of serosal patches but had a deleterious effect on epithelialization and mucosal enzyme activity. The transient effect of thiphenamil and the associated increase of hydroxyproline content suggest that collagen may have the predominant role in contraction.

Animals↗

Basement membrane components enhance isolated enterocyte growth.

Isolated enterocyte transplantation may have a potential role in increasing intestinal surface area. However, enterocytes are notoriously difficult to grow. Basement membrane components (BMC) promote adherence, migration, and differentiation of enterocytes. Our aim was to determine if BMC enhance enterocyte growth. Twenty-nine rabbits had 5-cm ileal segments resected and serosal pouches (n = 22) created from the serosal surface of the colon. Harvesting of enterocytes by warm trypsinization resulted in 92 +/- 6% cell viability and yielded 5.0 +/- 2.4 10(6) enterocytes/cm intestine. Enterocytes (10(5) were cultured in vitro (n = 7) in 10 ml growth media in plain flasks and flasks coated with laminin alone or Matrigel (an extract of mouse basement membrane containing laminin plus Type IV collagen and heparan sulfate). The cells were subcultured at 2 weeks and examined after Geimsa staining at 4 weeks. Epithelial growth was confirmed by light microscopy and staining for cytokeratin and quantitated by image analysis (JAVA). Epithelial coverage of the flasks was greater with Matrigel (80 +/- 15%) than laminin (66 +/- 15%) which was greater than control (44 +/- 20%) (P less than 0.01). For in vivo studies 10(5) harvested cells were infused into the serosal pouches either in growth media (n = 9) or media + Matrigel (n = 13). Epithelial growth in the pouch was evaluated by qualitative scoring of cytokeratin staining. Cytokeratin staining was similar on control colon serosa (n = 5) and serosa after infusion of cells in media alone. However, Matrigel significantly enhanced the area, depth, and intensity of staining.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

The effect of immunization with protein-hapten conjugate on the intestinal uptake of p-aminobenzoic acid as a hapten.

Intestinal uptake of p-aminobenzoic acid was examined by means of an in vitro everted sac technique in rats immunized with ovalbumin-p-aminobenzoic acid conjugate. A dose-dependent and antigen-specific decrease in the serosal transfer of p-aminobenzoic acid was observed in rats immunized 6 times with protein-hapten conjugate compared with the control. There was a significant increase in the recovery of p-acetamidobenzoic acid, a metabolite of p-aminobenzoic acid, in mucosal fluid, tissue, and serosal fluid in the jejunum. In the case of ileum, increase of p-acetamidobenzoic acid was observed in mucosal fluid. However, there was no significant effect in the ileal p-acetamidobenzoic acid in tissue and serosal fluid between immunized and non-immunized rats. To examine the increased metabolism of immunized rats, N-acetyltransferase activity of the small intestinal mucosa was examined. There was a significant increase in mucosal N-acetyltransferase activity in immunized rats compared with the control animals. These observations suggested that the mucosal immune system may play an important role in regulating the intestinal uptake of the low molecular weight compounds.

4-Aminobenzoic Acid↗

Acid-base balance of pleural liquid in dogs.

Acid-base balance and electrolyte concentrations were measured in dogs on small artificial hydrothoraces and in vitro on bicarbonate buffered Ringer solution on serosal and interstitial side of specimens of parietal pleura. Under steady conditions, pleural liquid PCO 2 was similar to and pH higher (delta = 0.022 +/- 0.006 SE) than that in mixed venous blood. Computed pleural liquid [HCO-3] was similar to that in venous plasma and hence less than that set by the Donnan effect, with which Na+ and Cl- approximately complied. In vitro, pH, [Na+], [Cl-], and computed [HCO-3] were significantly lower (delta = 0.030 +/- 0.004; -2.6 +/- 0.5; -1.2 +/- 0.5 and -1.7 +/- 0.2 meq/L, respectively) on the serosal than on interstitial side of pleural specimens, PCO2 being 42 mm Hg on both sides . HCO-3 and Na+ were not distributed according to transpleural potential (-0.4 +/- 0.1 mV on serosal side), suggesting an active transport of Na+ and HCO-3 from pleural liquid to blood. This, however, does not seem to add to the absorption pressure of plasma proteins in setting pleural liquid pressure.

Acid-Base Equilibrium↗

Triphasic effect of prostaglandins E1, E2 and F2alpha on the fluid transport of isolated gall-bladder of guinea-pigs.

Prostaglandins (PGs) F2alpha, E1 and E2 exerted a triphasic influence on the fluid transport of isolated guinea-pig gall-bladders, when applied to the serosal side. PGE1 and PGE2 produced these effects in lower concentrations than F2alpha. Directly after PG addition to the serosal side a short stimulation of fluid transport to between 200 and 400% was observed. The stimulatory effect of PGs was most distinct in gall-bladders from female guinea-pigs, less pronounced in male and nearly absent in pregnant animals. Since PGs increased intraluminal hydrostatic pressure in gall-bladders by contraction of the smooth muscle, experiments were performed in which hydrostatic pressure was increased by different procedures. These included the addition of imidazole (10- minus 2 M), raising of K+ in the bathing solution and an increase in intraluminal pressure by addition of Ringer's solution into the lumen. All three procedures stimulated fluid reabsorption temporarily in the same way as PGs, hence increase of intraluminal pressure is thought to be the reason for the observed temporary stimulation of fluid transport. Direct evidence for this thesis was obtained when the gall-bladder was mounted as a flat sheet over a chamber; in this preparation no stimulation of fluid transport was obtained. The second phase of the PG influence was characterized by a concentration-related inhibition of fluid reabsorption followed by a significant but small reverse of fluid transport (secretion of fluid). When PGs were applied to the mucosal side, only an inhibition of fluid transport was observed, which was much weaker compared to the addition to the serosal side.

Animals↗

Absorption of intact morphiceptin by diisopropylfluorophosphate-treated rabbit ileum.

The amidated beta-casomorphin morphiceptin Tyr-Pro-Phe-Pro-NH2 is an opioid peptide isolated from bovine milk beta-casein digests whose physiological significance remains unclear. Opiates are known to modify intestinal electrolyte transport by acting on receptors located on the serosal side of the intestine. The aim of the present study was to determine under what conditions morphiceptin can act from the luminal side. When added to the serosal side of untreated rabbit ileum in an Ussing chamber in vitro, 10(-3) M morphiceptin acted through an opiate mechanism to reduce simultaneously short-circuit current (delta Isc = 0.33 +/- 0.07 muEq.hr-1.cm-2) and stimulate net Na and Cl absorption (delta JnetNa = 1.62 +/- 0.11 and delta JnetCl = 2.07 +/- 0.08 muEg.hr-1.cm-2). After mucosal addition under the same conditions, morphiceptin was degraded without any opiate action on electrolyte transport. Pretreatment of the ileum by 10(-3) M diisopropylfluorophosphate, which inhibited brush-border dipeptidylpeptidase IV, prevented mucosal degradation of morphiceptin. Under these conditions, morphiceptin was able, when added mucosally, to cross the epithelium intact (Jm----s = 1.8 +/- 0.16 nmole.hr-1.cm-2) and to stimulate electrolyte absorption by means of an opioid mechanism (delta Isc = 0.22 +/- 0.02 muEq.hr-1.cm-2). These results showed that the action of morphiceptin from the lumen depends on its transfer intact to the serosal side of the intestine where the opiate receptors are located. The limiting step in this transfer is at the brush-border membrane, where dipeptidylpeptidase IV in particular seems to play a major role.

Analgesics↗

In vitro studies on the intestinal absorption of curcumin in rats.

When everted sacs of rat intestines were incubated with 50-750- micrograms of curcumin in 10 ml incubation medium, 30-80% of the added curcumin disappeared from the mucosal side. No curcumin was however detectable in the serosal fluid. Less than 3% of the added curcumin was found in the tissue at the highest concentration. In experiments with [3H[ curcumin, 5-6% of added radioactivity was found in the serosal side. TLC examination of the mucosal extract showed the presence of 2 compounds, 1 corresponding to curcumin and the other to a less polar, colourless compound. The serosal fluid had no curcumin but a compound whose RF was identical with the colourless compound present in the mucosal side. These experiments indicated that curcumin undergoes transformation during absorption from the intestine.

Animals↗

Intestinal glucose and galactose transport in the cultured gilthead bream (Sparus aurata).

1. Electrical parameters and transepithelial glucose and galactose transport were determined in vitro across anterior and posterior intestine of the culture fish Sparus aurata. 2. Electrical potential difference (PD) and short-circuit current (Isc) were serosa-positive in anterior intestine, while they were serosa-negative or near zero in posterior intestine. 3. Tissue conductance (Gt) was higher in posterior than in anterior intestine. In both parts it was decreased when the Na ion was omitted in mucosal and serosal reservoirs. 4. Addition of glucose or galactose to the mucosal side of intestine caused an increase in PD and Isc in posterior intestine but did not significantly change PD and Isc in anterior intestine. 5. Isotopic flux of glucose and galactose measurements in short-circuit conditions showed a net active glucose and galactose absorption in posterior intestine, while in anterior intestine active transport of glucose or galactose was not observed. 6. The net transport of glucose and galactose in posterior intestine was decreased to zero in the absence of Na in mucosal and serosal reservoirs or in the presence of ouabain (1 mM) in serosal solution.

Animals↗

The beta-anomeric and glucose preferences of glucose transport carrier for intestinal active absorption of monosaccharide conjugates.

The anomeric preference for intestinal absorption of glucosides and galactosides (1- and 2-naphthyl glycosides) was studied by the everted rat intestine method. After the absorption of beta-glucoside and beta- and alpha-galactosides, the glycosides itself appeared on the serosal side, whereas after the alpha-glucoside absorption, the glucoside itself was not detected on the serosal side, but a large amount of aglycone appeared instead. This indicates an alpha-anomeric preference of desglucosylation through the intestinal membrane. A significant decrease of the total (glycoside + aglycone + glucuronide metabolites) amount transported to the serosal side in the absence of Na+, a cosubstrate of the glucose transport carrier (GTC), was observed in alpha- and beta-glucosides and beta-galactoside, but not in alpha-galactoside. This indicates the poor contribution of GTC to the alpha-galactoside absorption. The Na(+)-dependent absorption of the glycosides by the GTC were beta-glucoside > alpha-glucoside > beta-galactoside (for 2-naphthyl glycosides), and beta-galactoside > alpha-galactoside (for 1-naphthhyl glycosides). These results and those of a previous study led to the conclusion that the intestinal glucose transport carrier prefers beta-anomer to alpha-anomer, and glucose to galactose for monosaccharide conjugates absorption.

Animals↗

Polychlorinated biphenyls (phenoclor and pyralene) and intestinal transport of hexoses and amino acids in mice.

1. Using mouse everted whole small intestine 7 days after a single dose of Phenoclor DP6 (hexachlorobiphenyl) at 1.0 g kg-1 body wt, absorption of D-glucose decreased significantly, probably owing to toxic traces of pentodicholobenzofuran in the PCB preparation and not its high chlorine content. D-glucose tissue accumulation decreased. The serosal fluid transfer also fell but not the gut fluid uptake. D-glucose absorption and fluid transfer remained unchanged following doses of other Phenoclor and Pyralene (PCB) compounds. 2. Using DP6, malabsorption of D-glucose and impaired serosal fluid transfer were not evident 7 days after a single dose of 0.1, 0.25 and 0.50 g kg-1 body wt. One to 21 days after a single dose (1.0 g kg-1 body wt) of DP6, D-glucose absorption and serosal fluid transfer decreased from 3 to 14 days but thereafter became normal. 3. Although absorption of D-galactose, D-glucose following membrane hydrolysis of D-maltose, and L-arginine, L-histidine, L-ornithine and L-proline decreased slightly 7 days after a single dose of Phenoclor DP6,the results were not significant, nor were changes in fluid transfer. 4. Following Phenoclor DP6-treatment, D-glucose malabsorption was abolished by an exogenous energy supply (D-mannose). DP6 affected intracellular metabolism and not the glucose carrier at the membrane brush border. 5. The body-weights and fresh small intestinal weights of mice treated with different PCBs remained unchanged after seven days. The histology of the small intestine showed minimal changes.

Amino Acids↗

A new model for regulation of sodium transport in high resistance epithelia.

A new three barrier, four compartment model for sodium transport in high resistance urinary epithelia is presented. This model provides a unified and simplified mechanistic explanation for sodium transport and its quantitative regulation. Sodium enters the epithelial cell by passive diffusion. Active extrusion occurs across the lateral cell membrane into the lateral intercellular space (LICS). Sodium movement from the LICS into the serosal compartment is not free and unobstructed as in the models for low resistance epithelia, but rather occurs through a regulatory channel of the LICS passing through desmosomes and the basilar slit. The exact configuration of this regulatory channel controls the rate of sodium movement from the LICS into the serosal compartment. Thus, the configuration of the regulatory channel controls the afterload on the sodium pump and thus ultimately controls the rate of transepithelial sodium transport. Antidiuretic hormone could act by increasing the effective width of this regulatory channel by contraction of intracellular microtubules or microfilaments. Present theories for regulation of transepithelial sodium transport in high resistance epithelia invoke a regulatory barrier at the apical cell membrane or at the active sodium pump located in the basolateral cell membrane. The hypothetical model presented here invokes a new alternative: regulation of the active pump rate by the sodium concentration in the LICS serving as an afterload on the pump; sodium escape from the LICS into the serosal compartment thus becomes the regulatory step for transepithelial transport.

Biological Transport↗

Effect of pH on aflatoxin B1 transfer in the everted rat jejunum.

The effect of varying mucosal pH (5, 7.4 and 8) on aflatoxin B1 (AFB1) transmural transfer was studied in the everted rat jejunum. The viability of the preparation was evaluated by: light microscopy; rate of glucose uptake by tissue from the mucosal fluid; content and distribution of water and electrolytes in the tissue; oxygen uptake by the tissue; and the intestinal permeability to [14C]sucrose. These parameters revealed the structural and functional integrity of the preparation, which was not affected by incubation in the presence of AFB1 or at different pH values in the mucosal fluid. The results obtained show that AFB1 was transferred to the serosal fluid and that the cumulative transfer was greater at pH 5 than at pH 7.4 or 8. In addition, AFB1 clearance, calculated from the slope of the plot of the cumulative amount of AFB1 transferred per unit of mucosal concentration vs. incubation time, indicated a very rapid accumulation by tissue and transfer of AFB1 to the serosal fluid. This process was also faster at pH 5. The integrity of the AFB1 molecule during incubation was shown by NMR spectra. The lack of metabolization of AFB1 during its translocation was demonstrated by TLC of the serosal fluid.

Aflatoxin B1↗

Intestinal SGLT1-mediated absorption and metabolism of benzyl beta-glucoside contained in Prunus mume: carrier-mediated transport increases intestinal availability.

The intestinal absorption of benzyl beta-glucoside (BNZ beta glc) contained in the fruit of Prunus mume SIEB. et ZUCC. (Rosaceae), which is traditionally used as a medicinal food in Japan, was studied in rat intestines. BNZ beta glc was absorbed from the mucosal to serosal sides. Its metabolite, benzyl alcohol (BAL), was also detected on both the mucosal and serosal sides. In the presence of phloridzin (Na(+)/glucose cotransporter (SGLT1) inhibitor) or in the absence of Na+ (driving force), BNZ beta glc absorption was significantly decreased. Transport clearance of BNZ beta glc across the brush border membrane decreased as its concentration increased. These results indicate that BNZ beta glc is transported by SGLT1. Metabolic clearance of BNZ beta glc also decreased as its concentration increased. The amount ratio of BNZ beta glc to BAL on the serosal side increased with the increase of BNZ beta glc concentration. The intestinal availability of BNZ beta glc was lower in the absence of Na+ than in the presence of Na+, indicating that the SGLT1-mediated transport of BNZ beta glc increases intestinal availability by decreasing the intestinal extraction ratio. This neutraceutical study concluded that intestinal carrier-mediated transport across the brush border membrane improves the intestinal availability of nutritionally, pharmacologically or physiologically active compounds that undergo intestinal metabolism (first-pass effect).

Animals↗