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Background: Berberine is a plant alkaloid that has multiple beneficial effects against intestine inflammation. In our previous study, we have found that berberine also possesses an antidiabetic effect. However, whether berberine is useful in the prevention of type 2 diabetes mellitus (T2DM) through its effect on intestine endocrine function and gut microbiota is unclear., Aim: To investigate the effects of berberine in the prevention of T2DM, as well as its effects on intestine GLP-2 secretion and gut microbiota in ZDF rats., Methods: Twenty Zucker Diabetic Fatty (ZDF) rats were fed a high-energy diet until they exhibited impaired glucose tolerance (IGT). The rats were then divided into two groups to receive berberine (100 mg/kg/d; berberine group) or vehicle (IGT group) by gavage for 3 weeks. Five Zucker Lean (ZL) rats were used as controls. Fasting blood glucose (FBG) was measured, an oral glucose tolerance test was performed, and the Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) was calculated. Intestinal expression of TLR-4, NF-κB, TNF-α, mucin, zona occludens-1 (ZO-1) and occludin were assessed (immunohistochemistry). Plasma levels and glutamine-induced intestinal secretion of glucagon-like peptide-1 (GLP-1) and GLP-2 were measured (enzyme-linked immunosorbent assay). The plasma lipopolysaccharide (LPS) level was measured. Fecal DNA extraction, pyrosequencing, and bioinformatics analysis were performed., Results: After 3 weeks of intervention, diabetes developed in all rats in the IGT group, but only 30% of rats in the berberine group. Treatment with berberine was associated with reductions in food intake, FBG level, insulin resistance, and plasma LPS level, as well as increases in fasting plasma GLP-2 level and glutamine-induced intestinal GLP-2 secretion. Berberine could increase the goblet cell number and villi length, and also reverse the suppressed expressions of mucin, occludin, ZO-1 and the upregulated expressions of TLR-4, NF-κB and TNF-α induced in IGT rats (P<0.05). Berberine also improved the structure of the gut microbiota and restored species diversity., Conclusion: Berberine may slow the progression of prediabetes to T2DM in ZDF rats by improving GLP-2 secretion, intestinal permeability, and the structure of the gut microbiota., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Wang, Liu, Zheng, Yang, Ren, Kong, Wang, Wang, Jiang, Yang and Shan.)

In this work, two different approaches have been developed to predict the food effect and the bioequivalence of marketed itraconazole (ITRA) formulations. Kinetic solubility and simultaneous dissolution-permeation tests of three (ITRA) formulations (Sporanox capsules and solution and SUBA-ITRA capsules) were carried out in simulated fasted and fed states. Fraction of dose absorbed ratios estimating food effect and bioequivalence were calculated based on these results and were compared to the in vivo study results published by Medicines Agencies. The comparison demonstrated that kinetic solubility and flux values could be used as input parameters for biopharmaceutics modeling and simulations to estimate food effect and bioequivalence. Both prediction methods were able to determine a slightly negative food effect in the case of the Sporanox solution and also a pronounced positive food effect for the Sporanox capsule. Superior bioavailability was predicted when the Sporanox solution was compared to the Sporanox capsule (in agreement with in vivo data).

Ethnopharmacological Relevance: Origanum majorana L. (Lamiaceae) is commonly used in Moroccan folk medicine to treat infantile colic, abdominal discomfort and diarrhea. Liquid stools and abdominal discomfort observed in acute infectious diarrhea are the consequences of imbalance between intestinal water secretion and absorption in the lumen, and relaxation of smooth muscle surrounding the intestinal mucosa., Aim of the Study: The objective of our study was to see if aqueous extract of Origanum majorana L. (AEOM) may exhibit an effect on those deleterious mechanisms., Materials and Methods: The effect of AEOM on electrogenic Cl - secretion and Na + absorption, the two main mechanisms underlying water movement in the intestine, was assessed on intestinal pieces of mice intestine mounted, in vitro, in Ussing chambers. AEOM effect on muscle relaxation was measured on rat intestinal smooth muscle mounted in an isotonic transducer., Results: 1) AEOM placed on the serosal (i.e. blood) side of the piece of jejunum entirely inhibited in a concentration-dependent manner the Forskolin-induced electrogenic chloride secretion, with an IC 50 = 654 ± 8 µg/mL. 2) AEOM placed on the mucosal (i.e. luminal) side stimulated in a concentration-dependent manner an electrogenic Na + absorption, with an IC 50 = 476.9 ± 1 µg/mL. 3) AEOM (1 mg/mL) inhibition of Forskolin-induced electrogenic secretion was almost entirely prevented by prior exposure to Ca ++ channels or neurotransmitters inhibitors. 4) AEOM (1 mg/mL) proabsorptive effect was greater in the ileum and progressively declined in the jejunum, distal colon and proximal colon (minimal). 5) AEOM inhibited in a concentration-dependent manner smooth muscle Carbachol or KCl induced contraction, with an IC 50 = 1.64 ± 0.2 mg/mL or 1.92 ± 0.8 mg/mL, respectively., Conclusion: the present results indicate that aqueous extract of Origanum majorana L. exhibit positive cooperative effects on the main mechanisms that are involved in acute infectious diarrhea., (Copyright © 2018. Published by Elsevier B.V.)

Secretory diarrhea is one of the most common causes of death world-wide especially in children under 5 years old. Isoliquiritigenin (ISLQ), a plant-derived chalcone, has previously been shown to exert anti-secretory action in vitro and in vivo by inhibiting CFTR Cl - channels. However, its CFTR inhibition potency is considerably low (IC 50  > 10 μM) with unknown mechanism of action. This study aimed to identify novel chalcone derivatives with improved potency and explore their mechanism of action. Screening of 27 chalcone derivatives identified CHAL-025 as the most potent chalcone analog that reversibly inhibited CFTR-mediated Cl - secretion in T84 cells with an IC 50 of ∼1.5 μM. As analyzed by electrophysiological and biochemical analyses, the mechanism of CFTR inhibition by CHAL-025 is through AMP-activated protein kinase (AMPK), a negative regulator of CFTR activity. Furthermore, Western blot analyses and molecular dynamics (MD) results suggest that CHAL-025 activates AMPK by binding at the allosteric site of an upstream kinase calcium/calmodulin-dependent protein kinase kinase β (CaMKKβ). Interestingly, CHAL-025 inhibited both cholera toxin (CT) and bile acid-induced Cl - secretion in T84 cells and prevented CT-induced intestinal fluid secretion in mice. Therefore, CHAL-025 represents a promising anti-diarrheal agent that inhibits CFTR Cl - channel activity via CaMKKβ-AMPK pathways., (Copyright © 2019 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Short-chain fatty acids (SCFAs), such as acetate, propionate, and butyrate, are synthesized from dietary carbohydrates by colonic bacterial fermentation. These SCFAs supply energy, suppress cancer, and affect ion transport. However, their roles in ion transport and regulation in the intracellular environment remain unknown. In order to elucidate the roles of SCFAs, we measured short-circuit currents (I SC ) and performed RT-PCR and immunohistochemical analyses of ion transporters in rat rectal colon. The application of 30 mM butyrate shifted I SC in a negative direction, but did not attenuate the activity of epithelial Na + channels (ENaC). The application of bumetanide, a Na + -K + -2Cl - cotransporter inhibitor, to the basolateral side reduced the negative I SC shift induced by butyrate. The application of XE991, a KCNQ-type K + channel inhibitor, to the apical side decreased the I SC shift induced by butyrate in a dose-dependent manner. The I SC shift was independent of HCO 3 - and insensitive to ibuprofen, an SMCT1 inhibitor. The mucosa from rat rectal colon expressed mRNAs of H + -coupled monocarboxylate transporters (MCT1, MCT4, and MCT5, also referred to as SLC16A1, SLC16A3, and SLC16A4, respectively). RT-PCR and immunofluorescence analyses demonstrated that KCNQ2 and KCNQ4 localized to the apical membrane of surface cells in rat rectal colon. These results indicate that butyrate, which may be transported by H + -coupled monocarboxylate transporters, activates K + secretion through KCNQ-type K + channels on the apical membrane in rat rectal colon. KCNQ-type K + channels may play a role in intestinal secretion and defense mechanisms in the gastrointestinal tract.

This study investigated the effects of the ethanolic extract from the bark of Combretum leprosum (ECL) on intestinal transit and castor-oil induced diarrhea in mice. The oral administration of ECL (750 and 1000 mg/kg) slowed intestinal transit (ID50 of 455 mg/kg). The ECL (250-1000 mg/kg) reduced castor-oil induced diarrhea, in a time- and dose-dependent manner (p < 0.05). To determine if antidiarrheal effect of ECL involves α2-adrenergic or opioid receptor activation, the mice were pretreated with antagonists of these receptors, yohimbine or naloxone respectively. None of these drugs inhibited the antidiarrheal effect of ECL. To test if antidiarrheal effect of ECL is due to an antisecretory action, we realized the enteropooling assay on rats. The ECL increased bowel content and did not inhibit intestinal fluid secretion increase induced by misoprostol (100 µg/kg, s.c.). To determine if antimotility effect of ECL is due to a reduction on gastric motility, we realized the organ bath assay in the rat fundus stomach. Isotonic recordings show that the carbachol /KCl - induced contraction was not reversed by the addition of ECL. In conclusion, our results suggest that ECL contains antidiarrheal compounds and these compounds could induce a reduction of intestinal tract motility.

Context: Sweet taste receptors (STRs) involve in regulating the release of glucose-stimulated glucagon-like peptide-1 (GLP-1). Our in vivo and in vitro studies found that 3-deoxyglucosone (3DG) inhibited glucose-stimulated GLP-1 secretion., Objective: This study investigated the role of STRs in 3DG-induced inhibition of high glucose-stimulated GLP-1 secretion., Methods: STC-1 cells were incubated with lactisole or 3DG for 1 h under 25 mM glucose conditions. Western blotting was used to study the expression of STRs signaling molecules and ELISA was used to analyse GLP-1 and cyclic adenosine monophosphate (cAMP) levels., Results: Lactisole inhibited GLP-1 secretion. Exposure to 25 mM glucose increased the expressions of STRs subunits when compared with 5.6 mM glucose. 3DG decreased GLP-1 secretion and STRs subunits expressions, with affecting other components of STRs pathway, including the downregulation of transient receptor potential cation channel subfamily M member 5 (TRPM5) expression and the reduction of intracellular cAMP levels., Conclusion: 3DG attenuates high glucose-stimulated GLP-1 secretion by reducing STR subunit expression and downstream signaling components.

Background: Plant foods may stimulate intestinal secretion through chemicals designed to deter herbivores, including lactucins in lettuce and rhein in rhubarb. This may increase ileostomy output and induce diarrhoea in people with intact bowels., Objective: We aimed to determine the effect of food on intestinal water content using Magnetic Resonance Imaging (MRI)., Design: A three period crossover trial of isocaloric meals in adults without bowel disorders. Meals: 2 slices white bread with 10 g butter; 300 g rhubarb with 60 mL lactose free cream; 300 g lettuce with 30 mL mayonnaise., Primary Outcome: Area under curve (AUC) small bowel water content (SBWC) using MRI., Secondary Outcomes: ascending colon water content; T1 relaxation time of ascending colon (T1AC); gastric volume; visual analogue scales of bloating and satiety (0-100). MRI analysts were blinded. Scanned fasting and hourly to 180 min postprandial. Symptoms scored half-hourly., Results: 9 female and 6 male subjects completed the study. AUC SBWC fell after bread but rose after lettuce and even more after rhubarb, difference from baseline being (Bread AUC -5662 (1209) ml.min vs Lettuce 3194 (1574) ml.min and Rhubarb 10586 (1629) ml.min (P < 0.01). Rhubarb induced a rise in T1AC but differences at 3 hours were not significant (P = 0.06). Gastric volume at T = 0 significantly was higher for both lettuce and rhubarb (571 ± 92 and 558 ± 89 mls) respectively compared to bread (314 ± 108 mls) (p < 0.0001). Symptom scores were higher for lettuce > rhubarb > bread., Conclusion: Lettuce and rhubarb meals increased intestinal water content, demonstrating how different foods can alter ileal flow and stool consistency., (© 2018 American Society for Parenteral and Enteral Nutrition.)

Aim: To investigate the effect of Hemp seed soft capsule (HSCC) on colonic ion transport and its related mechanisms in constipation rats., Methods: Sprague-Dawley male rats were randomly divided into three groups: normal group, constipation group and HSSC group. Rats in the constipation and HSSC groups were administrated loperamide 3 mg/kg per day orally for 12 d to induce the constipation model. Then, the HSSC group was given HSSC 0.126 g/kg per day by gavage for 7 d. The normal and constipation groups were treated with distilled water. After the treatment, the fecal wet weight and water content were measured. The basal short-circuit current ( Isc ) and resistance were measured by an Ussing Chamber. Besides the in vivo drug delivery experiment above, an in vitro drug application experiment was also conducted. The accumulative concentrations of HSSC (0.1 mg/mL, 0.5 mg/mL, 1.0 mg/mL, 2.5 mg/mL, 5.0 mg/mL, 10.0 mg/mL and 25.0 mg/mL) were added to the normal isolated colonic mucosa and the Isc was recorded. Further, after the application of either ion (Cl - or HCO 3 - ) substitution, ion channel-related inhibitor (N-phenylanthranilic acid, glybenclamide, 4,4-diisothiocyano-2,2-stilbenedisulfonic acid or bumetanide) or neural pathway inhibitor [tetrodotoxin (TTX), atropine, or hexamethonium], the Isc induced by HSSC was also measured., Results: In the constipation group, the fecal wet weight and the water content were decreased in comparison with the normal group ( P < 0.01). After the treatment with HSSC, the fecal wet weight and the water content in the HSSC group were increased, compared with the constipation group ( P < 0.01). In the constipation group, the basal Isc was decreased and resistance was increased, in comparison with the normal group ( P < 0.01). After the treatment with HSSC, the basal Isc was increased ( P < 0.05) and resistance was decreased ( P < 0.01) in the HSSC group compared with the constipation group. In the in vitro experiment, beginning with the concentration of 1.0 mg/mL, differences in Isc were found between the experimental mucosa (with HSSC added) and control mucosa. The Isc of experimental mucosa was higher than that of control mucosa under the same concentration (1.0 mg/mL, P < 0.05; 2.5-25 mg/mL, P < 0.01). After the Cl - or HCO 3 - removal and pretreated with different inhibitors (cAMP-dependent and Ca 2+ -dependent Cl - channels, Na + -K + -2Cl - cotransporter (NKCC), Na + -HCO 3 - cotransporter or Cl - /HCO 3 - exchanger inhibitor), there were differences between experimental mucosa and control mucosa; the Isc of experimental mucosa was lower than that of control mucosa under the same concentration ( P < 0.05). Meanwhile, after pretreatment with neural pathway inhibitor (TTX, atropine, or hexamethonium), there were no differences between experimental mucosa and control mucosa under the same concentration ( P > 0.05)., Conclusion: HSSC ameliorates constipation by increasing colonic secretion, which is mediated via the coaction of cAMP-dependent and Ca 2+ -dependent Cl - channels, NKCC, Na + -HCO 3 - cotransporter or Cl - /HCO 3 - exchanger., Competing Interests: Conflict-of-interest statement: None of the authors declared any conflicts of interest related to this study.

Aim: To investigate the acute toxicity, phytochemical profile, antidiarrheal activity and mechanisms of action of Maytenus erythroxylon ( M. erythroxylon ) ethanol extract., Methods: A castor oil-induced diarrhea model was used to evaluate antidiarrheal activity. Intestinal transit and gastric emptying protocols were used to evaluate a possible antimotility effect. K ATP channels, nitric oxide, presynaptic α2-adrenergic and tissue adrenergic receptors were investigated to uncover antimotility mechanisms of action and castor oil-induced enteropooling to elucidate antisecretory mechanisms., Results: All tested doses of the extract (62.5, 125, 250 and 500 mg/kg) possessed antidiarrheal activity, with a significant decrease of the evacuation index. This activity is possibly related to a reduced gastric emptying (125, 250 and 500 mg/kg) and to a decreased percentage of intestinal transit for all tested doses. That last effect seems to be modulated by nitric oxide, K ATP channels and tissue adrenergic receptors. Besides, the extract also presented antisecretory effect due to a decrease of intestinal fluid accumulation., Conclusion: The antidiarrheal effect of M. erythroxylon found in this study involves antimotility and antisecretory mechanisms that may be attributed to the chemical compounds found in this species: saponins, flavonoids, tannins, triterpenes and steroids., Competing Interests: Conflict-of-interest statement: The authors declare that no conflict of interest exists.

Background: Diarrhea in piglets is one of the main causes of animal death after weaning; zinc oxide (ZnO) has been used in high doses for the control of this sickness. The aim of this study was to determine the physicochemical properties of ZnO nanoparticles synthesized and immobilized on a chitosan/alginate (CH/SA) complex and investigate the antimicrobial activity and in vitro release profile of zinc (Zn 2+ ) from these new compounds. The ZnO nanoparticles composites were prepared and combined with CH/SA or CH/SA and sodium tripolyphosphate (TPP). The structure and morphology of the composites were analyzed by characterization methods such as X-ray diffraction, FTIR spectroscopy, thermogravimetric analysis, atomic absorption spectrophotometry and scanning electron microscopy., Results: The crystallite size of ZnO nano was 17 nm and the novel ZnO composites were effective in protecting ZnO in simulated gastric fluid, where Zn 2+ reached a concentration six-fold higher than the levels obtained with the unprotected commercial-zinc oxide. In addition, the novel composites suggest effective antimicrobial activity against Escherichia coli and Staphylococcus aureus., Conclusions: The results described herein suggest that the novel nano composites may work as an alternative product for pig feeding as verified by the in vitro assays, and may also contribute to lower the zinc released in the environment by fecal excretion in animals waste.

Intestinal Cl - secretion is involved in the pathogenesis of secretory diarrheas including cholera. We recently demonstrated that flufenamic acid (FFA) suppressed Vibrio cholerae El Tor variant-induced intestinal fluid secretion via mechanisms involving AMPK activation and NF-κB-suppression. The present study aimed to investigate the effect of FFA on transepithelial Cl - secretion in human intestinal epithelial (T84) cells. FFA inhibited cAMP-dependent Cl - secretion in T84 cell monolayers with IC 50 of ∼8 μM. Other fenamate drugs including tolfenamic acid, meclofenamic acid and mefenamic acid exhibited the same effect albeit with lower potency. FFA also inhibited activities of CFTR, a cAMP-activated apical Cl - channel, and KCNQ1/KCNE3, a cAMP-activated basolateral K + channel. Mechanisms of CFTR inhibition by FFA did not involve activation of its negative regulators. Interestingly, FFA inhibited Ca 2+ -dependent Cl - secretion with IC 50 of ∼10 μM. FFA inhibited activities of Ca 2+ -activated Cl - channels and K Ca 3.1, a Ca 2+ -activated basolateral K + channels, but had no effect on activities of Na + -K + -Cl - cotransporters and Na + -K + ATPases. These results indicate that FFA inhibits both cAMP and Ca 2+ -dependent Cl - secretion by suppressing activities of both apical Cl - channels and basolateral K + channels. FFA and other fenamate drugs may be useful in the treatment of secretory diarrheas., (Copyright © 2017 The Authors. Production and hosting by Elsevier B.V. All rights reserved.)

Bile acids (BAs) play a complex role in colonic fluid secretion. We showed that dihydroxy BAs, but not the monohydroxy BA lithocholic acid (LCA), stimulate Cl(-) secretion in human colonic T84 cells (Ao M, Sarathy J, Domingue J, Alrefai WA, Rao MC. Am J Physiol Cell Physiol 305: C447-C456, 2013). In this study, we explored the effect of LCA on the action of other secretagogues in T84 cells. While LCA (50 μM, 15 min) drastically (>90%) inhibited FSK-stimulated short-circuit current (Isc), it did not alter carbachol-stimulated Isc LCA did not alter basal Isc, transepithelial resistance, cell viability, or cytotoxicity. LCA's inhibitory effect was dose dependent, acted faster from the apical membrane, rapid, and not immediately reversible. LCA also prevented the Isc stimulated by the cAMP-dependent secretagogues 8-bromo-cAMP, lubiprostone, or chenodeoxycholic acid (CDCA). The LCA inhibitory effect was BA specific, since CDCA, cholic acid, or taurodeoxycholic acid did not alter FSK or carbachol action. While LCA alone had no effect on intracellular cAMP concentration ([cAMP]i), it decreased FSK-stimulated [cAMP]i by 90%. Although LCA caused a small increase in intracellular Ca(2+) concentration ([Ca(2+)]i), chelation by BAPTA-AM did not reverse LCA's effect on Isc LCA action does not appear to involve known BA receptors, farnesoid X receptor, vitamin D receptor, muscarinic acetylcholine receptor M3, or bile acid-specific transmembrane G protein-coupled receptor 5. LCA significantly increased ERK1/2 phosphorylation, which was completely abolished by the MEK inhibitor PD-98059. Surprisingly PD-98059 did not reverse LCA's effect on Isc Finally, although LCA had no effect on basal Isc, nystatin permeabilization studies showed that LCA both stimulates an apical cystic fibrosis transmembrane conductance regulator Cl(-) current and inhibits a basolateral K(+) current. In summary, 50 μM LCA greatly inhibits cAMP-stimulated Cl(-) secretion, making low doses of LCA of potential therapeutic interest for diarrheal diseases., (Copyright © 2016 the American Physiological Society.)

Guanylin (GN) action on seawater eel intestine was examined under simulated in vivo conditions, where isotonic luminal fluid has low NaCl and high MgSO4 (MgSO4 Ringer). In Ussing chamber, MgSO4 Ringer induced serosa-negative potential difference (PD) even after bumetanide treatment, which is due to the higher paracellular Na(+) permeability over Cl(-), as confirmed by the replacement by MgCl2 (no Cl(-) gradient) or Na2SO4 Ringer (no Na(+) gradient). Luminal GN reversed serosa-negative PD, probably by enhancing Cl(-) secretion into the lumen, as the GN effect was blocked by apical Cl(-) channel blockers [diphenylamine-2-carboxylic acid (DPC), 5-nitro-2-(3-phenylpropylamino) benzoic acid, glibenclamide but not cystic fibrosis transmembrane regulator (CFTR)inh-172] or replacement of luminal fluid by MgCl2 Ringer. The blockers' effect was undetectable when normal Ringer was on both sides. In the sac preparation, NaCl secretion occurred into the lumen (Na(+) > Cl(-)), and GN further enhanced Cl(-) secretion (Cl(-) > Na(+)), resulting in water secretion. These GN effects were also blocked by DPC. Quantitative analyses showed that isotonic NaCl is absorbed when luminal fluid is normal Ringer, but, when luminal fluid is MgSO4 Ringer, hypertonic NaCl, almost equivalent to seawater, is secreted into the lumen after GN. These results indicate that GN stimulates the secretion of hypertonic NaCl into the lumen of seawater eel intestine, like rectal gland of marine elasmobranchs, to get rid of excess NaCl although marine teleost intestine is thought to have only absorptive-type cells with a unique Na-K-Cl cotransport system. The secreted NaCl may activate the cotransport system and further help absorb water in the final segment of seawater eel intestine., (Copyright © 2015 the American Physiological Society.)

Regulation of intestinal secretion is important for body fluid homeostasis. We investigated the role of three MAP kinases (MAPKs) as negative regulators in muscarinic cholinoceptor (mAChR)-mediated intestinal secretion in mice. Electrophysiological analyses revealed that mAChR stimulation enhanced intestinal chloride secretion, which was further augmented by the inhibition of JNK but not by that of ERK or p38 with specific inhibitors SP600125, U0126 or SB203580, respectively. Immunoblot analyses in colonic mucosa showed that mAChR stimulation increased MAPKs phosphorylation that was suppressed by the specific inhibitor for each MAPK. This suggests that JNK is a major negative regulator in mAChR-induced intestinal secretion., (Copyright © 2014 Japanese Pharmacological Society. Production and hosting by Elsevier B.V. All rights reserved.)

Proteases play a diverse role in health and disease. An excessive concentration of proteases has been found in the feces of patients with inflammatory bowel disease or irritable bowel syndrome and been implicated in the pathogenesis of such disorders. This study examined the effect of the serine protease, trypsin, on intestinal epithelial anion secretion when added to the luminal side. A mucosal-submucosal sheet of the mouse cecum was mounted in Ussing chambers, and the short-circuit current (I sc) was measured. Trypsin added to the mucosal (luminal) side increased I sc with an ED50 value of approximately 10 μM. This I sc increase was suppressed by removing Cl(-) from the bathing solution. The I sc increase induced by 10-100 μM trypsin was substantially suppressed by tetrodotoxin, and partially inhibited by a neurokinin-1 receptor antagonist, but not by a muscarinic or nicotinic ACh-receptor antagonist. The trypsin-induced I sc increase was also significantly inhibited by a 5-hydroxytryptamine-3 receptor (5-HT3) antagonist and substantially suppressed by the simultaneous addition of both 5-HT3 and 5-HT4 receptor antagonists. We conclude that luminal trypsin activates the enteric reflex to induce anion secretion, 5-HT and substance P playing important mediating roles in this secreto-motor reflex. Luminal proteases may contribute to the cause of diarrhea occurring with some intestinal disorders.

Objectives: The aim of this study was to establish a minimally invasive and simple screening test for detection of pancreatic ductal adenocarcinoma (PDAC) using duodenal juice (DJ)., Methods: Duodenal juice was collected prospectively before endoscopic retrograde cholangiopancreatography in 46 patients. A protease inhibitor was not added to the samples collected during the initial 2.5 minutes but was added in the latter 2.5 minutes. Thereafter, secretin was administered intravenously, and DJ was subsequently collected for additional 10 minutes. The sensitivities of carcinoembryonic antigen (CEA), S100 calcium-binding protein P (S100P), and interleukin 8 in DJ and pancreatic juice were assessed., Results: There were 30 patients with PDAC and 16 with benign lesions. It was possible to collect an adequate amount of DJ without secretin administration. In the PDAC group, CEA concentrations in DJ were significantly higher than those in the benign group, even without the use of a protease inhibitor. S100P levels in DJ in the PDAC group were significantly higher than those in the benign group in the presence of the protease inhibitor., Conclusions: Duodenal juice collection during routine upper endoscopy and assessments of CEA and S100P in DJ might become a useful screening test for detection of PDAC.

Objectives: The aims were: firstly, to review the definition and diagnosis of irritable bowel syndrome with constipation (IBS-C, a subtype of IBS); secondly, to critically assess current therapies for IBS-C with a focus on effectiveness for abdominal pain; and thirdly, to review clinical studies evaluating the efficacy of linaclotide, a therapy recently approved by the US Food and Drug Administration for the treatment of adults with IBS-C and chronic idiopathic constipation and the European Medicines Agency for the symptomatic treatment of moderate to severe IBS-C in adults, and in development for treatment of IBS-C worldwide., Methods: A comprehensive literature review was performed to summarize IBS-C and current treatments. MEDLINE and gastrointestinal society congress proceedings were searched to identify data from linaclotide clinical studies in adults with IBS-C published between January 2010 and August 2012., Results: IBS-C patients have chronic, relapsing symptoms. Rome III diagnostic criteria define the presence of chronic abdominal pain that improves with defecation and has onset associated with changes in stool frequency or form as a key element of IBS-C and other IBS subtypes. IBS-C patients generally are not completely satisfied with existing therapies. A therapy that treats bowel and abdominal symptoms effectively and can be taken safely on a chronic basis is a current unmet need for IBS-C patients. The guanylate cyclase-C agonist linaclotide has been shown to reduce visceral hypersensitivity in preclinical studies and to improve abdominal pain and constipation symptoms in phase 2 and 3 clinical trials of IBS-C patients., Conclusions: IBS-C is a functional gastrointestinal disorder with chronic, relapsing abdominal and constipation symptoms. By virtue of its effects in relieving abdominal pain by reducing visceral hypersensitivity and improving constipation symptoms by increasing intestinal secretion and accelerating transit, linaclotide may be uniquely positioned for a role in the management of IBS-C patients.

Objectives: Zinc is a useful addition to oral rehydration therapy for acute diarrhoea. We have assessed the mechanism of its epithelial antisecretory action when intestinal epithelial tight junctions were pharmacologically opened., Methods: Rat isolated ileal and colonic mucosae were mounted in Ussing chambers and exposed to ZnSO(4) (Zn(2+) ) in the presence of secretagogues and inhibition of short circuit current (I(sc) ) was measured., Key Findings: Pre-incubation with basolateral but not apical Zn(2+) reduced I(sc) stimulated by forskolin, carbachol and A23187. In the presence of the tight junction-opener, cytochalasin D, antisecretory effects of apically-applied Zn(2+) were enabled in colon and ileum. The apparent permeability coefficient (P(app) ) of Zn(2+) was increased 1.4- and 2.4-fold across rat ileum and colon, respectively, by cytochalasin D. Basolateral addition of Zn(2+) also reduced the I(sc) stimulated by nystatin in rat colon, confirming K channel inhibition. In comparison with other inhibitors, Zn(2+) was a relatively weak blocker of basolateral K(ATP) and K (Ca2+) channels. Exposure of ileum and colon to Zn(2+) for 60 min had minimal effects on epithelial histology., Conclusions: Antisecretory effects of Zn(2+) on intestinal epithelia arose in part through nonselective blockade of basolateral K channels, which was enabled when tight junctions were open., (© 2012 The Authors. JPP © 2012 Royal Pharmaceutical Society.)

Background: The purpose of this investigation was to evaluate the efficacy and tolerability of a tannic acid-based medical food, Cesinex(®), in the treatment of diarrhea and to investigate the mechanisms underlying its antidiarrheal effect., Methods: Cesinex(®) was prescribed to six children and four adults with diarrhea. Patient records were retrospectively reviewed for the primary outcome. Cesinex(®) and its major component, tannic acid, were tested for their effects on cholera toxin-induced intestinal fluid secretion in mice. Polarized human gut epithelial cells (HT29-CL19A cells) were used to investigate the effects of tannic acid on epithelial barrier properties, transepithelial chloride secretion, and cell viability., Results: Successful resolution of diarrheal symptoms was reported in nine of ten patients receiving Cesinex(®). The treatment of HT29-CL19A cells with clinically relevant concentrations of tannic acid (0.01-1 mg/ml) significantly increased transepithelial resistance (TER) and inhibited the cystic fibrosis transmembrane conductance regulator (CFTR)-dependent or the calcium-activated Cl(-) secretion. Tannic acid could also improve the impaired epithelial barrier function induced by tumor necrosis factor alpha (TNFα) and inhibited the disrupting effect of TNFα on the epithelial barrier function in these cells. Cholera toxin (CTX)-induced mouse intestinal fluid secretion was significantly reduced by the administration of Cesinex(®) or tannic acid. Cesinex(®) has high antioxidant capacity., Conclusions: Cesinex(®) demonstrates efficacy and a good safety profile in the treatment of diarrhea. The broad-spectrum antidiarrheal effect of Cesinex(®) can be attributed to a combination of factors: its ability to improve the epithelial barrier properties, to inhibit intestinal fluid secretion, and the high antioxidant capacity.

Background/aims: Daily subcutaneous injections with the phytoestrogen genistein, 600 mg/ kg genistein/day (600G) significantly increased intestinal chloride (Cl(-)) secretion (I(sc), µA/cm(2)) in C57BL/6J female and male murine jejunum after 1-2-weeks treatment., Methods and Results: In 600G females, basolateral application of the adenylate cyclase inhibitor MDL-12330A (10 µM) significantly reduced basal and total I(sc) in the presence of forskolin (27 and 40% respectively, P < 0.05), with no effect in 600G males, suggesting that 600G-mediated increases in I(sc) in females are due to an adenylate cyclase-dependent mechanism. Concomitant injections with the non-selective estrogen receptor (ER) antagonist ICI-182780 (25 mg/kg/day) resulted in a significant inhibition of basal I(sc) in males (38%, P < 0.05), but was without effect in females (further reinforcing an ER-independent mechanism of action). The ERα-selective antagonist (MPP, 25 mg/kg/day) similarly significantly inhibited the basal I(sc) (37%, P < 0.05) in males, whereas the ERβ-selective antagonist (PHTPP, 25 mg/kg/day) was without effect, suggesting that 600G-mediated increases in I (sc) in male mice are due to an ERα-dependent mechanism. Jejunum ERα/actin expression was significantly increased by 600G in males. Compared to intact mice, orchiectomy has differing effects on 600G-mediated basal Isc; castration (CAST) abolished the 600G-mediated increases in I(sc), and ovariectomy (OVX) had no effect on the 600G-stimulated increases in I(sc). Daily estradiol injections (10-20 mg/kg body weight estradiol (10E2 or 20E2) had no effect in intact females, whereas 10E2 significantly increased basal I(sc) in OVX females., Conclusion: These data suggest that daily estradiol and genistein injections have differential sex-dependent mechanisms of action on murine intestinal Cl(-) secretion., (Copyright © 2012 S. Karger AG, Basel.)

Aim: To clarify whether CFTR is a molecular target of intestinal fluid secretion caused by the anthraquinone compounds from laxative herbal plants., Methods: A cell-based fluorescent assay to measure I(-) influx through CFTR chloride channel. A short-circuit current assay to measure transcellular Cl(-) current across single layer FRT cells and freshly isolated colon mucosa. A closed loop experiment to measure colon fluid secretion in vivo., Results: Anthraquinone compounds rhein, aloe-emodin and 1,8-dihydroxyanthraquinone (DHAN) stimulated I(-) influx through CFTR chloride channel in a dose-dependent manner in the presence of physiological concentration of cAMP. In the short-circuit current assay, the three compound enhanced Cl(-) currents in epithelia formed by CFTR-expressing FRT cells with EC(50) values of 73 ± 1.4, 56 ± 1.7, and 50 ± 0.5 μmol/L, respectively, and Rhein also enhanced Cl(-) current in freshly isolated rat colonic mucosa with a similar potency. These effects were completely reversed by the CFTR selective blocker CFTR(inh)-172. In in vivo closed loop experiments, rhein 2 mmol/L stimulated colonic fluid accumulation that was largely blocked by CFTR(inh)-172. The anthraquinone compounds did not elevate cAMP level in cultured FRT cells and rat colonic mucosa, suggesting a direct effect on CFTR activity., Conclusion: Natural anthraquinone compounds in vegetable laxative drugs are CFTR potentiators that stimulated colonic chloride and fluid secretion. Thus CFTR chloride channel is a molecular target of vegetable laxative drugs.

New therapies are being developed for irritable bowel syndrome (IBS). These advances are based on understanding pathophysiology or the development of medications with greater selectivity in classes of agents with known efficacy. Prucalopride, the newest European Medicines Agency-approved 5-hydroxytryptamine receptor 4 (5-HT(4)) agonist, is effective in the treatment of chronic constipation with improved cardiovascular safety relative to older 5-HT(4) drugs; similarly, ramosetron, the 5-hydroxytryptamine receptor 3 (5-HT(3)) antagonist, appears efficacious in diarrhea-predominant IBS. Secretagogues with different mechanisms of action target apical domains in enterocytes that are involved in chloride secretion, such as chloride channels, the cystic fibrosis transmembrane regulator, and guanylate cyclase C. As a class, such secretagogues have high efficacy and safety for constipation. With more data obtained from phase 2 and 3 trials, we expect other classes of medications, including bile acid modulators, anti-inflammatory agents, visceral analgesics, and newer centrally acting agents to be efficacious and enter the armamentarium for the treatment of IBS in the future.

The aim of this study was to evaluate the ability of TMC, with different degrees of quaternization, to increase insulin absorption in vivo following nasal and rectal administration in rats. Two batches of TMC with different degrees of quaternization (TMC-L, 12.3% quaternized and TMC-H, 61.2% quaternized) and chitosan hydrochloride were administered intranasally (0.25 and 0.5% w/v) and rectally (0.5% w/v) with insulin (4 IU/kg body weight), at a pH of 4.40 and 7.40, in rats. Blood samples were taken over a period of 2 h for measurement of blood glucose levels and plasma insulin levels. Local toxicity evaluation was done by histological examination of the nasal and rectal epithelia. At pH 4.40 all these polymers were able to increase nasal and rectal insulin absorption, compared to the control groups. However, at a pH of 7.40, only TMC-H was able to increase the nasal and rectal absorption of insulin. These results relate to the insolubility of chitosan hydrochloride at neutral pH values, while the charge density of TMC-L is still too low for any significant interaction at pH 7.40. Histological evaluation of the nasal and rectal eptihelia shows no changes in the morphology of the cells after exposure to these polymers. Only slight congestion of the nasal submucosa was observed and all these polymers led to a mild increase in mucus secretion at pH 4.40. Highly quaternized TMC proves to be a potent absorption enhancer in vivo, especially at neutral pH values where chitosan salts are ineffective.

Hermannia incana Cav. is a prostrate herb used to treat diarrhea, stomach ache, nausea and vomiting, by the people of Eastern Cape Province, South Africa. The phytochemical screening as well as the antidiarrheal activity of H. incana leaf extract at 200, 400 and 600 mg/kg body weight was evaluated in rats. Phytochemical screening revealed the presence of bioactive agents such as alkaloids, tannins, saponins, phenolics, triterpenes, cardiac glycosides, flavonoids, cardenolides and dienolides. The extract significantly prolonged the time of induction of diarrhea, reduced the frequency of diarrheal episodes and water content of the feces, and inhibited castor oil-induced enteropooling. The extract also suppressed intestinal propulsive movement of a charcoal meal through the gastrointestinal tract. These results demonstrate the antidiarrheal properties of the extract, thereby supporting the folkloric use of the plant as an antidiarrheal agent in the Eastern Cape of South Africa., (Copyright 2010 Prous Science, S.A.U. or its licensors. All rights reserved.)

TMT (trimethyltin chloride), an organotin, is ubiquitous in the environment. The consumption of contaminated food may cause exposure of the human diet to this toxic compound. The present study was to investigate the effects of TMT on the regulation of ion transport across the rat distal colon. The rat colonic mucosa was mounted in Ussing chambers. The effects of TMT were assessed using the Isc (short-circuit current). Both apical and basolateral TMT induced, dose-dependently, an increase in Isc, which was due to a stimulation of Cl- secretion as measured using ion substitution experiments and pharmacological manoeuvres. The secretion was also inhibited by several K+ channel blockers administrated at the basolateral side. When the apical side was permeabilized by nystatin, the TMT-induced K+ conductance was effectively blocked by tetrapentylammonium, a Ca2+-sensitive K+ channel blocker. The response of TMT was sensitive to the basolateral Ca2+ and the intracellular Ca2+ store, which could be disclosed by applying the inhibitors of ryanodine receptors and inositol 1,4,5-trisphosphate receptors. In conclusion, TMT led to Cl- secretion, which was essentially regulated by basolateral Ca2+-sensitive K+ channels. These results suggest the importance of K+ channels in the toxicity hazard of TMT.

A Na(+)/HCO(3)(-) cotransporter (NBC) is located in the basolateral membrane of the gastrointestinal epithelium, where it imports HCO(3)(-) during stimulated anion secretion. Having previously demonstrated secretagogue activation of NBC in murine colonic crypts, we now asked whether vesicle traffic and exocytosis are involved in this process. Electrogenic NBCe1-B was expressed at significantly higher levels than electroneutral NBCn1 in colonic crypts as determined by QRT-PCR. In cell surface biotinylation experiments, a time-dependent increase in biotinylated NBCe1 was observed, which occurred with a peak of +54.8% after 20 min with forskolin (P < 0.05) and more rapidly with a peak of +59.8% after 10 min with carbachol (P < 0.05) and which corresponded well with the time course of secretagogue-stimulated colonic bicarbonate secretion in Ussing chamber experiments. Accordingly, in isolated colonic crypts pretreated with forskolin and carbachol for 10 min, respectively, and subjected to immunohistochemistry, the NBCe1 signal showed a markedly stronger colocalization with the E-cadherin signal, which was used as a membrane marker, compared with the untreated control. Cytochalasin D did not change the observed increase in membrane abundance, whereas colchicine alone enhanced NBCe1 membrane expression without an additional increase after carbachol or forskolin, and LY294002 had a marked inhibitory effect. Taken together, our results demonstrate a secretagogue-induced increase of NBCe1 membrane expression. Vesicle traffic and exocytosis might thus represent a novel mechanism of intestinal NBC activation by secretagogues.

The thiazolidinedione (TZD) drugs rosiglitazone (Ro) and pioglitazone (Po) are PPARgamma agonists in widespread clinical use as insulin-sensitizing agents in Type 2 diabetes. On the basis of recent evidence implicating PPARgamma as a positive modulator of intestinal epithelial differentiation, we hypothesized that TZD drugs might attenuate intestinal secretory function. To evaluate this possibility, we examined the effects of Ro and Po on electrogenic Cl- secretion [short-circuit current (I(sc))] in mouse intestinal segments and in cultured human intestinal epithelial cells (HT29-Cl.19A). As hypothesized, oral administration of Ro (20 mg.kg(-1).day(-1)) to mice for 8 days markedly reduced intestinal I(sc) responses to cAMP (forskolin)- and Ca2+ (carbachol)-dependent stimuli. In these Ro-treated mice, cholera toxin-induced intestinal fluid accumulation was reduced 65%. With continued Ro treatment, the I(sc) response to carbachol recovered significantly, whereas that to forskolin remained attenuated. Treatment of HT29 cells for 5 days with 10 muM Ro or Po in vitro brought about a similar hyposecretory state. In HT29 cells, the loss of cAMP-dependent Cl- secretion was attributable to a reduced expression of CFTR Cl- channel, KCNQ1 K+ channel, and Na-K-2Cl cotransporter-1 proteins. The transient loss of Ca2+-dependent Cl- secretion involved an impairment of basolateral Ca2+-stimulated K+ channel activity without a detectable loss of K(Ca)3.1 channel protein. Our results establish TZD drugs as important modulators of intestinal Cl- secretory function.

Chronic constipation is a common condition that affects up to 27% of the population. Dietary and lifestyle measures are usually the first-line therapy, but if these fail to have an effect then a variety of prescription and consumer laxatives are available. Traditional laxatives include bulking agents, osmotic agents, stool softeners, and stimulants of the gastrointestinal tract. All have been found to be more effective than placebo at relieving symptoms of constipation, but these results have been obtained primarily in short (4-week) trials and no class of laxative has been shown to be superior to another. Traditional laxatives work in many, but not all, patients and some patients cannot cope with the side effects, unpleasant taste, the requirements of the dosing regimen, or the notion of dose increase. New enterokinetic agents that affect peristalsis through selective interaction with 5-hydroxytryptamine-4 receptors and novel agents acting on intestinal secretion could offer an alternative option for patients with chronic constipation who cannot get adequate relief from current laxatives.

Taste transduction molecules, such as Galpha(gust), and taste receptor families for bitter [taste receptor type 2 (T2R)], sweet, and umami, have previously been identified in taste buds and the gastrointestinal (GI) tract; however, their physiological functions in GI tissues are still unclear. Here, we investigated the physiological function and expression of T2R in human and rat large intestine using various physiological and molecular biological techniques. To study the physiological function of T2R, the effect of a bitter compound, 6-n-propyl-2-thiouracil (6-PTU), on transepithelial ion transport was investigated using the Ussing chamber technique. In mucosal-submucosal preparations, mucosal 6-PTU evoked Cl(-) and HCO(3)(-) secretions in a concentration-dependent manner. In rat middle colon, levels of 6-PTU-evoked anion secretion were higher than in distal colon, but there was no such difference in human large intestine. The response to 6-PTU was greatly reduced by piroxicam, but not by tetrodotoxin. Additionally, prostaglandin E(2) concentration-dependently potentiated the response to 6-PTU. Transcripts of multiple T2Rs (putative 6-PTU receptors) were detected in both human and rat colonic mucosa by RT-PCR. In conclusion, these results suggest that the T2R ligand, 6-PTU, evokes anion secretion, and such response is regulated by prostaglandins. This luminal bitter sensing mechanism may be important for host defense in the GI tract.

Background and Purpose: The aim of this study was the identification of the mechanism of oxidant-induced intestinal secretion., Experimental Approach: The action of H2O2 on ion transport across rat distal colon was evaluated in Ussing chambers. Changes in cytosolic Ca2+ concentration were measured using fura-2., Key Results: H2O2 concentration-dependently induced an increase in short-circuit current (Isc), which was due to a stimulation of Cl(-) secretion. The effect of H2O2 was dependent on the presence of serosal Ca2+. It was inhibited after emptying of intracellular Ca2+ stores by cyclopiazonic acid or blockade of ryanodine receptors by ruthenium red, whereas a blocker of inositol 1,4,5-trisphosphate receptors was less effective. Fura 2-experiments confirmed an increase in the cytosolic Ca2+ concentration in the presence of H2O2. Measurements of Cl- currents across the apical membrane at basolaterally depolarized epithelia revealed the activation of a glibenclamide-sensitive, SITS-resistant Cl- conductance by the oxidant. The activation of this conductance was inhibited after blockade of protein kinases with staurosporine. When the apical membrane was permeabilized with nystatin, two sites of action of H2O2 were identified at the basolateral membrane. The oxidant stimulated a basolateral tetrapentylammonium-sensitive K+ conductance and increased the current generated by the Na+-K+ pump. Pretreatment of the tissues with H2O2 reduced the action of subsequently administered Ca2+-, cAMP- and cGMP-dependent secretagogues demonstrating a long-term downregulation after the initial secretory response evoked by the oxidant., Conclusions and Implications: H2O2 affects colonic anion secretion by action sites at both the apical, as well as the basolateral membrane.

Marine teleosts have extracellular fluids less concentrated than their environment, resulting in continual water loss, which is compensated for by drinking, with intestinal water absorption driven by NaCl uptake. Absorption of Cl(-) occurs in part by apical Cl(-)/HCO(3)(-) exchange, with HCO(3)(-) provided by transepithelial transport and/or by carbonic anhydrase-mediated hydration of endogenous epithelial CO(2). Hydration of CO(2) also liberates H(+), which is transported across the basolateral membrane. In this study, gulf toadfish (Opsanus beta) were acclimated to 9, 35 and 50 ppt. Intestinal HCO(3)(-) secretion, water and salt absorption, and the ensuing effects on acid-base balance were examined. Rectal fluid excretion greatly increased with increasing salinity from 0.17+/-0.05 ml kg(-1) h(-1) in 9 ppt to 0.70+/-0.19 ml kg(-1) h(-1) in 35 ppt and 1.46+/-0.22 ml kg(-1) h(-1) in 50 ppt. Rectal fluid composition and excretion rates allowed for estimation of drinking rates, which increased with salinity from 1.38+/-0.30 to 2.60+/-0.92 and 3.82+/-0.58 ml kg(-1) h(-1) in 9, 35 and 50 ppt, respectively. By contrast, the fraction of imbibed water absorbed decreased from 85.9+/-3.8% in 9 ppt to 68.8+/-3.2% in 35 ppt and 61.4+/-1.0% in 50 ppt. Despite large changes in rectal base excretion from 9.3+/-2.7 to 68.2+/-20.4 and 193.2+/-64.9 mumol kg(-1) h(-1) in 9, 35 and 50 ppt, respectively, acute or prolonged exposure to altered salinities was associated with only modest acid-base balance disturbances. Extra-intestinal, presumably branchial, net acid excretion increased with salinity (62.0+/-21.0, 229.7+/-38.5 and 403.1+/-32.9 mumol kg(-1) h(-1) at 9, 35 and 50 ppt, respectively), demonstrating a compensatory response to altered intestinal base secretion associated with osmoregulatory demand.

Stevioside and its major metabolite, steviol, have been reported to affect ion transport in many types of tissues, such as the kidney, pancreas, and intestine. The effect of stevioside, steviol, and its analogs on intestinal Cl(-) secretion was investigated in a human T84 epithelial cell line. Short-circuit current measurements showed that steviol and analogs isosteviol, dihydroisosteviol, and isosteviol 16-oxime inhibited in a dose-dependent manner forskolin-induced Cl(-) secretion with IC(50) values of 101, 100, 9.6, and 50 microM, respectively, whereas the parent compound stevioside had no effect. Apical Cl(-) current measurement indicated that dihydroisosteviol targeted the cystic fibrosis transmembrane regulator (CFTR). The inhibitory action of dihydroisosteviol was reversible and was not associated with changes in the intracellular cAMP level. In addition, dihydroisosteviol did not affect calcium-activated chloride secretion and T84 cell viability. In vivo studies using a mouse closed-loop model of cholera toxin-induced intestinal fluid secretion showed that intraluminal injection of 50 microM dihydroisosteviol reduced intestinal fluid secretion by 88.2% without altering fluid absorption. These results indicate that dihydroisosteviol and similar compounds could be a new class of CFTR inhibitors that may be useful for further development as antidiarrheal agents.

The reflex response to distension of the small intestine in vivo is complex and not well understood. The aim of this study was to characterize the neural mechanisms contributing to the complex time course of the intestinal secretory response to distension. Transmucosal potential difference (PD) was used as a marker for mucosal chloride secretion, which reflects the activity of the secretomotor neurons. Graded distensions (5, 10, and 20 mmHg) of distal rat duodenum with saline for 5 min induced a biphasic PD response with an initial peak (rapid response) followed by a plateau (sustained response). The rapid response was significantly reduced by the neural blockers tetrodotoxin and lidocaine (given serosally) and by intravenous (iv) administration of the ganglionic blocker hexamethonium and the NK(1) receptor antagonist SR-140333. Serosal TTX and iv SR-140333 significantly reduced the sustained response, which was also reduced by the NK(3) receptor antagonist talnetant and by the vasoactive intestinal polypeptide (VPAC) receptor antagonist [4Cl-d-Phe(6), Leu(17)]-VIP. Serosal lidocaine and iv hexamethonium had no significant effect on this component. Inhibition of nitric oxide synthase had no effect on any of the components of the PD response to distension. The PD response to distension thus seems to consist of two components, a rapidly activating and adapting component operating via nicotinic transmission and NK(1) receptors, and a slow component operating via VIP-ergic transmission and involving both NK(1) and NK(3) receptors.

The majority of prostaglandins (PGs) are known to induce intestinal fluid secretion (enteropooling). In contrast, PGD(2) has been demonstrated to inhibit fluid secretion induced by other PGs. This study was aimed to investigate, by the use of selective agonists/antagonists, which type of PGD(2) receptor mediates this inhibitory effect. The DP1 agonist BW245C dose-dependently inhibited the enteropooling effect of 16,16-dimethyl-PGE(2). This inhibition was counteracted by the DP1 antagonist BWA868C. In contrast, the CRTH2 receptor does not seem to be involved in the anti-enteropooling effect of PGD(2), since the selective agonists 13,14-dihydro-15-keto-PGD(2) and 15(R)-15-methyl-PGD(2) were without effect. Therefore, our results suggest that the inhibitory effect of PGD(2) in the small intestine is mediated via activation of the DP1 receptor., (Copyright 2008 S. Karger AG, Basel.)

Indigenous uses of Azadirachta indica A. juss (Maliaceae) (locally known as neem) leaves in different parts of India for curing gastrointestinal disorder such as diarrhea and cholera is wide spread. The objective of the present study was to evaluate the antibacterial and antisecretory activity of neem extract against Vibrio cholerae, a causative agent of watery diarrhea such as cholera. The methanol extract of neem leaf was tested for its antibacterial, antisecretory and antihemorrhagic activity against Vibrio cholerae. Azadirachta indica extract had significant antibacterial activity against the multi-drug-resistant Vibrio cholerae of serotypes O1, O139 and non-O1, non-O139. The minimum inhibitory concentration reached by 50% (MIC50) and 90% (MIC90), and minimum bactericidal concentration for the extract were 2.5, > 5, and 10 mg/ml, respectively. Neem extract showed antisecretory activity on Vibrio cholerae induced fluid secretion in mouse intestine with inhibition values of 27.7%, 41.1%, 43.3%, 57.0%, and 77.9% at doses of 100, 200, 300, 450 and 1800 mg/kg, respectively. Oral administration of the extract inhibited hemorrhage induced by Vibrio cholerae in mouse intestine at a dose > or = 300 mg/kg. The results obtained in this study give some scientific support to the uses of neem employed by the indigenous people in India employed for the treatment of diarrhea and dreadful disease cholera.

Heme induces Cl(-) secretion in intestinal epithelial cells, most likely via carbon monoxide (CO) generation. The major source of endogenous CO comes from the degradation of heme via heme oxygenase (HO). We hypothesized that an inhibitor of HO activity, tin protoporphyrin (SnPP), may inhibit the stimulatory effect of heme on Cl(-) secretion. To test this hypothesis, we treated an intestinal epithelial cell line (Caco-2 cells) with SnPP. In contrast to our expectations, Caco-2 cells treated with SnPP had an increase in their short-circuit currents (I(sc)) in Ussing chambers. This effect was observed only when the system was exposed to ambient light. SnPP-induced I(sc) was caused by Cl(-) secretion because it was inhibited in Cl(-)-free medium, with ouabain or 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB). The Cl(-) secretion was not via activation of the CFTR, because a specific inhibitor had no effect. Likewise, inhibitors of adenylate cyclase and guanylate cyclase had no effect on the enhanced I(sc). SnPP-induced I(sc) was inhibited by the antioxidant vitamins, alpha-tocopherol and ascorbic acid. Electron paramagnetic resonance experiments confirmed that oxidative reactions were initiated with light in cells loaded with SnPP. These data suggest that SnPP-induced effects may not be entirely due to the inhibition of HO activity but rather to light-induced oxidative processes. These novel effects of SnPP-photosensitized oxidation may also lead to a new understanding of how intestinal Cl(-) secretion can be regulated by the redox environment of the cell.

Ethanol extracts of the fresh aerial parts of various Epilobium species were tested to elucidate the mechanism of their gastrointestinal activity in animals. The methods of charcoal meal, castor oil-induced diarrhoea, and enteropooling assay were used to evaluate their effect on mouse gut at various dose levels. The extracts were found to have a significant activity in all models. Moreover, the extracts resulted to possess very little toxicity. Thus, it can be concluded that Epilobium possesses anti-diarrhoeal, anti-motility, and anti-secretory activities and can prove beneficial in the treatment of gastrointestinal disorders.

The calcium-sensing receptor (CaSR) provides a fundamental mechanism for diverse cells to detect and respond to modulations in the ionic and nutrient compositions of their extracellular milieu. The roles for this receptor are largely unknown in the intestinal tract, where epithelial cells are normally exposed to large variations in extracellular solutes. Here, we show that colonic CaSR signaling stimulates the degradation of cyclic nucleotides by phosphodiesterases and describe the ability of receptor activation to reverse the fluid and electrolyte secretory actions of cAMP- and cGMP-generating secretagogues, including cholera toxin and heat stable Escherichia coli enterotoxin STa. Our results suggest a paradigm for regulation of intestinal fluid transport where fine tuning is accomplished by the counterbalancing effects of solute activation of the CaSR on neuronal and hormonal secretagogue actions. The reversal of cholera toxin- and STa endotoxin-induced fluid secretion by a small-molecule CaSR agonist suggests that these compounds may provide a unique therapy for secretory diarrheas.

Intestinal infection with Vibrio cholerae results in secretory diarrhea with potentially massive fluid losses and volume depletion. Morbidity and mortality associated with cholera remain a major problem in the developing world despite the success of oral rehydration therapy. New research aiming to inhibit cholera toxin binding to receptors in the intestine provides an attractive strategy for cholera therapy. Together with anti-secretory agents, including inhibitors of enkephalinase and of the cystic fibrosis transmembrane conductance regulator, new treatment options for managing severe diarrhea in cholera could soon be available.

We showed previously that exposure to microcystin-LR causes renal toxic effects in isolated perfused rat kidney, and that inflammatory mediators from supernatants of macrophages stimulated by microcystin-LR are involved in this process. The aim of this research was to examine water and electrolytes secretion in vivo, induced by microcystin-LR and supernatant of macrophages stimulated for this toxin (SUP.MphiS + MCLR), using perfused rat ileal segment and ligated intestinal loop models. We found microcystin-LR at 1 microg/ml (0.09 +/- 0.003* vs. control 0.07 +/- 0.001 g of secretion/2 cm of loop; P < 0.05*) and the SUP.MphiS + MCLR after 18 h postinoculation (0.10 +/- 0.003 vs. control 0.03 +/- 0.002 g/cm) caused intestinal secretion. In addition, microcystin-LR caused significant sodium secretion (-2.18 +/- 0.72* vs. control 2.18 +/- 0.50 microEq g(-1) min(-1)), potassium (-0.26 +/- 0.04* vs. control 0.32 +/- 0.03 microEq g(-1) min(-1)), chloride (MCLR = -3.29 +/- 1.93* vs. control 0.88 +/- 1.25 microEq g(-1) min(-1)) and water (-0.012 +/- 0.004* vs. control 0.002 +/- 0.002 ml g(-1) min(-1)). We also demonstrated SUP.MphiS + MCLR to induce intestinal secretion of electrolytes (sodium, potassium, chloride) and water. These findings suggested that microcystin-LR and lamina propria macrophages-derived mediators are able to induce intestinal secretion in vivo, probably via inhibition of protein phosphatase.

Precipitation of basic drugs within oral prolonged release systems, at the higher pH values of the intestine, would affect drug release. Coevaporates of a model basic drug verapamil HCl, in single or mixed polymer systems, containing Eudragit L100 (L100) and ethyl cellulose or Eudragit RS100, were prepared from ethanolic solution. XRD and DSC indicated loss of crystallinity of the drug in the coevaporates. The presence of the enterosoluble polymer in the system was found to aid in faster dissolution of the drug at higher pH values. This was affected by the presence and type of retarding polymer present in the system. Compression of the coevaporates resulted in either very slow release of the drug or undesirable changes in the release profile. Pelletization of a coevaporate containing drug and L100 yielded systems, which released the drug uniformly when studied by the buffer change method in simulated gastric (SGF) and intestinal (SIF) fluids. The presence of L100 in intimate contact with the drug was found to be essential for the desirable drug release properties of the system. The drug release occurred predominantly by diffusion in SGF and by a combination of diffusion and polymer dissolution/erosion in SIF. Appropriate choice of release modifiers and formulation variables and development of suitable formulations can yield systems which compensate for the reduced solubility of the drug in the higher pH environments of the intestine.

Acute infectious diarrhoea continues to cause high morbidity and mortality worldwide. Although oral rehydration therapy has reduced the mortality associated with acute diarrhoea, stool volume often increases during the rehydration process. Therefore, for > 20 years there has been a search for agents that will directly inhibit intestinal secretory mechanisms and thereby reduce stool volume. The most obvious target for antisecretory therapy has been the chloride channel and second messengers within the enterocyte. So far, this search has been largely unrewarding, although recent evidence suggests that a new class of chloride channel blocker is effective in vitro but further evaluation in humans is required. In addition, research during the past decade has highlighted the importance of neurohumoral mechanisms in the pathogenesis of diarrhoea, notably the role of 5-hydroxtryptamine, substance P, vasoactive intestinal polypeptide and neural reflexes within the enteric nervous system. This new dimension of intestinal pathophysiology has already exposed possible novel targets for antisecretory therapy; namely, 5-hydroxytryptamine receptor antagonists, substance P antagonists and sigma-receptor agonists. There is also the possibility for potentiating the proabsorptive effects of endogenous enkephalins by using enkephalinase inhibitors. There now seems to be a real possibility that antisecretory therapy will become more widely available in the future.

Gum arabic, a branched polysaccharide, was oxidized using periodate to generate reactive aldehyde groups on the biopolymer. Primaquine, an 8-aminoquinoline, was covalently coupled onto oxidized gum arabic via an imine bond and simultaneously fabricated into microspheres of less than 2 microm in size by heat denaturation in a reverse emulsion of 1:1 light paraffin oil and toluene stabilized by sorbitan sesquioleate as the surfactant. The covalent binding of primaquine to the polysaccharide using the clinically used water-soluble form of the drug primaquine phosphate was achieved in the presence of borate buffer of pH 11. Up to 35% of the drug could be bound to the polymer backbone depending on the concentration of the drug employed initially and the degree of oxidation of the polysaccharide. Interestingly, both the aliphatic and the hindered aromatic amino groups of primaquine were found to react with the aldehyde functions through Schiff base formation leading to cross-linking of the polysaccharide with the drug itself. In vitro release of the drug from microspheres into phosphate buffered saline (PBS, pH 7.4, 0.1 M) at 37 degrees C showed that the release of primaquine from the matrix was slow, although gradually increased with time. The maximum released was below 50% of the drug payload even after 10 days. Release into simulated gastric and intestinal fluids was faster compared to the release in PBS due to rapid hydrolysis of the Schiff's linkage in the gastric fluid. A possible reason for the poor hydrolytic susceptibility of the Schiff's linkage is suggested based on the unequal reactivity of the amino groups on primaquine and its relevance in possible therapeutic application of this polymer-drug conjugate discussed.

An experiment was conducted with the objective to enhance mucosal immunity against ovalbumin (OVA) by co-administration of OVA with an aqueous extract from the fruit of Solanum torvum (STE). Five groups of female ICR mice aged approximately 8 weeks at the commencement of the experiment were caged in groups of eight and received various treatments. The treatments included OVA alone, OVA with cholera toxin (CT), and OVA with various doses of STE. Mice were primed intraperitoneally with 500 microg of OVA alone or co-administered with 0.1 microg CT, or with 1 microg STE. All mice were boosted orally via gastric intubation 14 days after priming with 10 mg OVA alone, or co-administered with 10 microg CT or with 10 mg, 1 mg or 0.1 mg STE. One week later all mice were killed and organs obtained for analysis of the immune response. Intestinal, faecal and pulmonary OVA-specific sIgA concentration was significantly increased (p<0.05) in mice that received booster combinations of OVA/CT and OVA with all extract doses (p<0.05). Specific serum IgG titres did not differ significantly between groups. It is concluded that STE can significantly enhance secretory immunity in the intestine to OVA with mucosal homing to the lungs. The adjuvant effect of STE is comparable to that of CT.

Objectives: As visceral afferents from different regions of the gastrointestinal tract converge at the level of the spinal cord, we hypothesized that sensitization of one gut organ would induce visceral hypersensitivity in another gut organ, remote to the sensitizing stimulus., Methods: Protocol 1: Eight healthy male volunteers, age 30 +/- 8.2 yr, underwent three studies on different days. Esophageal pain thresholds (PT) were recorded at 10-min intervals prior to and for 2 h following a 30-min duodenal infusion of either 0.15 M hydrochloric acid (HCl), saline, or no infusion. Five subjects repeated the study to demonstrate reproducibility. Protocol 2: Esophageal evoked potentials (EEP) were studied in six subjects on two occasions prior to and 1 h after a 30-min duodenal infusion of 0.15 M HCl or saline., Results: Protocol 1: After acid infusion, there were reproducible reductions in esophageal PT (ICC = 0.88), which were maximal at 110 min (15.05 +/- 2.25 mA) (p < 0.002). Following saline infusion there was an increase in esophageal PT (ICC = 0.71), which was similar to the no-infusion condition (6.21 +/- 1.54 mA vs 8.5 + 7.6 mA; p > 0.05). Protocol 2: Esophageal sensation scores increased (p= 0.02) after acid, but not after saline infusion (p= 0.1). A comparison of the latencies of EEP components prior to and following acid and saline infusion revealed a reduction in the N1 (p= 0.02) and P2 components (p= 0.04)., Conclusion: This study provides the first objective evidence that duodenal acidification can induce esophageal hypersensitivity associated with changes in sensitivity of the central visceral pain pathway. As the esophagus was remote from the sensitizing stimulus, central sensitization of spinal dorsal horn neurons is likely to have contributed to these changes.

The beneficial effects of red, yellow and white onion extracts have been assessed by antioxidant activity and antimutagenic activity. And the effects compared to BHT and ascorbic acid. Total phenolic compounds and flavonoids in onion extracts were determined. Yellow onion extract had more organic acid and free sugar than those detected in the white and red onion extract. The scavenging activity of DPPH radical and H(2)O(2) were increased depending on the concentration. The antioxidant activities using beta-carotene-linoleate system and reducing power were increased but the effect was small to that of BHT and ascorbic acid. After digested, extracts showed antimutagenic activities, and it seems that they inhibit the mutagenicity for digesting. This study demonstrated that the antimutagenicities and antioxidant properties of ethyl acetate extract against mutagens were related to their phenols and flavonoids, which are heat stable and losses digestive juices are relatively low.

After oral administration several gut-binding lectins induce accumulation of liquor and amylase in the proximal small intestine. Orally administered Phaseolus vulgaris phytohaemagglutinin (PHA) was used to study the mediation of these effects in rats. The regulation of amylase secretion clearly differed from that of the liquor. The amylase activity was of pancreatic origin, in agreement with the known cholecystokinin-releasing effect of PHA. It appears that CCK exerts its effect both directly and by facilitating neural stimulatory pathways. Intestinal secretion was identified as the source of the liquor, without a contribution by other secretions. It was mediated by a local cholinergic reflex with the involvement of both muscarinic and nicotinic acetylcholine receptors. It is speculated that the observed enteric reflex may enable the gut to transport secreted antibacterial peptides or secretory antibodies from the crypts to adherent bacteria on adjacent villi.

Fluid transport was measured gravimetrically in vivo in the jejunum, ileum and colon of fed, fasting (four days) and undernourished (50 % of control food intake for 21 days) gerbils (Gerbillus cheesmani). The effects of luminal enterotoxin Escherichia coli STa (50 ng/ml) and luminal 8-bromo-cyclic GMP (cGMP 1 mM) on fluid transport across jejunum, ileum and colon were also assessed. Fasting and undernourishment reversed the normal basal fluid absorption measured in fed ileum and colon into secretion. Neither fasting nor undernourishment had any effect on jejunal basal fluid absorption. In jejuna, ilea and colons of fed animals as well as in jejuna from fasting and undernourished gerbils STa (50 ng) reversed the normal absorptive "tone" to secretion but it had no significant effects on fluid secretion in either the ileum or colon from fasted gerbils. STa increased significantly the fluid secretion in ileum from undernourished gerbils. Luminal cGMP had no effect on basal absorptive tone in the jejunum of fed and fasted gerbils, but reversed absorption into secretion in the jejuna from undernourished gerbils. In the ilea taken from fed animals the small basal absorption was reversed to secretion by luminal cGMP. Although cGMP produced no significant changes in fluid secretion in the ilea taken from fasted gerbils, yet it caused a significant increase in those from undernourished gerbils. In the colon taken from fed animals cGMP decreased the basal fluid absorption significantly, but it had no significant effect on fluid secretion in the colon of fasted or undernourished gerbils. We conclude that fasting and undernourishment have no significant effects on fluid transport across the gerbil jejunum but reversed basal absorption in the fed ileum and colon into secretion. cGMP mimic the effects of STa in the jejunum taken from undernourished gerbils, in the ileum obtained under the three nutritional states and in the colon taken from fasting animals.

Background: Cholera toxin (CT) acts on intestinal epithelial cells both directly and indirectly via activation of a secretory neural reflex. The reflex may release acetylcholine as one of its final neurotransmitters. This opens up the possibility of a third mechanism of action for CT, namely a synergistic interaction between two secretagogues acting on different second messenger systems within the epithelial cell., Aims: To establish evidence for cholinergic innervation to human ileal epithelial cells and to investigate whether CT potentiates the action of acetylcholine on human intestinal epithelial cells., Methods: Transverse sections of human ileum were examined for mucosal cholinergic nerves and M3 muscarinic receptors using antibodies raised to choline acetyltransferase and M3 receptors. Short circuit current (Isc) responses and ion flux movements were elicited from T84 epithelial cell monolayers set up in Ussing chambers., Results: Immunohistochemistry of native human ileal mucosa revealed the presence of both cholinergic nerves and muscarinic M3 receptors located to the basolateral domain of epithelial cells. Secretory responses of T84 cell monolayers to acetylcholine were greatly potentiated in the presence of CT. This effect, substituting forskolin for CT, was mirrored by increases in basolateral 86Rb and apical 125I efflux. Charybdotoxin plus apamin reduced both Isc and 86Rb efflux evoked by acetylcholine, in the presence of forskolin., Conclusions: Human ileal mucosa receives a direct cholinergic innervation to its epithelial cells. Secretory effects of acetylcholine on epithelial cells are augmented in the presence of CT. Such a synergistic response is dependent on optimum opening of basolateral potassium channels by acetylcholine and apical chloride channels by CT. The interaction may contribute to the mechanism of action of cholera toxin induced secretory diarrhoea.