The intestinal microbiota affect central levels of brain-derived neurotropic factor and behavior in mice.

BACKGROUND & AIMS: Alterations in the microbial composition of the gastrointestinal tract (dysbiosis) are believed to contribute to inflammatory and functional bowel disorders and psychiatric comorbidities. We examined whether the intestinal microbiota affects behavior and brain biochemistry in mice. METHODS: Specific pathogen-free (SPF) BALB/c mice, with or without subdiaphragmatic vagotomy or chemical sympathectomy, or germ-free BALB/c mice received a mixture of nonabsorbable antimicrobials (neomycin, bacitracin, and pimaricin) in their drinking water for 7 days. Germ-free BALB/c and NIH Swiss mice were colonized with microbiota from SPF NIH Swiss or BALB/c mice. Behavior was evaluated using step-down and light preference tests. Gastrointestinal microbiota were assessed using denaturing gradient gel electrophoresis and sequencing. Gut samples were analyzed by histologic, myeloperoxidase, and cytokine analyses; levels of serotonin, noradrenaline, dopamine, and brain-derived neurotropic factor (BDNF) were assessed by enzyme-linked immunosorbent assay. RESULTS: Administration of oral antimicrobials to SPF mice transiently altered the composition of the microbiota and increased exploratory behavior and hippocampal expression of BDNF. These changes were independent of inflammatory activity, changes in levels of gastrointestinal neurotransmitters, and vagal or sympathetic integrity. Intraperitoneal administration of antimicrobials to SPF mice or oral administration to germ-free mice did not affect behavior. Colonization of germ-free BALB/c mice with microbiota from NIH Swiss mice increased exploratory behavior and hippocampal levels of BDNF, whereas colonization of germ-free NIH Swiss mice with BALB/c microbiota reduced exploratory behavior. CONCLUSIONS: The intestinal microbiota influences brain chemistry and behavior independently of the autonomic nervous system, gastrointestinal-specific neurotransmitters, or inflammation. Intestinal dysbiosis might contribute to psychiatric disorders in patients with bowel disorders.

Impaired parasympathetic function increases susceptibility to inflammatory bowel disease in a mouse model of depression.

Clinical and experimental evidence indicates that intestinal inflammatory conditions can be exacerbated by behavioral conditions such as depression. The recent demonstration of a tonic counterinflammatory influence mediated by the vagus nerve in experimental colitis provides a potential link between behavior and gut inflammation. Here we show that experimental conditions that induced depressive-like behaviors in mice increased susceptibility to intestinal inflammation by interfering with the tonic vagal inhibition of proinflammatory macrophages and that tricyclic antidepressants restored vagal function and reduced intestinal inflammation. These results show that reserpine-induced monoamine depletion and maternal separation, 2 models for depression, produced a vulnerability to colitis by a mechanism involving parasympathetic transmission and the presence of gut macrophages. The tricyclic antidepressant desmethylimipramine protected against this vulnerability by a vagal-dependent mechanism. Together these results illustrate the critical role of the vagus in both the vulnerability to inflammation induced by depressive-like conditions and the protection afforded by tricyclic antidepressants and rationalize a clinical evaluation of both parasympathomimetics and tricyclic antidepressants in treatment of inflammatory bowel disease.

Chronic gastrointestinal inflammation induces anxiety-like behavior and alters central nervous system biochemistry in mice.

BACKGROUND & AIMS: Clinical and preclinical studies have associated gastrointestinal inflammation and infection with altered behavior. We investigated whether chronic gut inflammation alters behavior and brain biochemistry and examined underlying mechanisms. METHODS: AKR mice were infected with the noninvasive parasite Trichuris muris and given etanercept, budesonide, or specific probiotics. Subdiaphragmatic vagotomy was performed in a subgroup of mice before infection. Gastrointestinal inflammation was assessed by histology and quantification of myeloperoxidase activity. Serum proteins were measured by proteomic analysis, circulating cytokines were measured by fluorescence activated cell sorting array, and serum tryptophan and kynurenine were measured by liquid chromatography. Behavior was assessed using light/dark preference and step-down tests. In situ hybridization was used to assess brain-derived neurotrophic factor (BDNF) expression in the brain. RESULTS: T muris caused mild to moderate colonic inflammation and anxiety-like behavior that was associated with decreased hippocampal BDNF messenger RNA (mRNA). Circulating tumor necrosis factor-α and interferon-γ, as well as the kynurenine and kynurenine/tryptophan ratio, were increased. Proteomic analysis showed altered levels of several proteins related to inflammation and neural function. Administration of etanercept, and to a lesser degree of budesonide, normalized behavior, reduced cytokine and kynurenine levels, but did not influence BDNF expression. The probiotic Bifidobacterium longum normalized behavior and BDNF mRNA but did not affect cytokine or kynurenine levels. Anxiety-like behavior was present in infected mice after vagotomy. CONCLUSIONS: Chronic gastrointestinal inflammation induces anxiety-like behavior and alters central nervous system biochemistry, which can be normalized by inflammation-dependent and -independent mechanisms, neither of which requires the integrity of the vagus nerve.

Reactivation of inflammatory bowel disease in a mouse model of depression.

BACKGROUND & AIMS: Patients with inflammatory bowel disease (IBD) frequently also have depression, yet little is known of its role in IBD pathogenesis. We investigated whether the development of depression after the establishment of chronic inflammation reactivates an acute relapse of IBD and underlying pharmacologic mechanisms in mouse models. METHODS: Colitis was induced by administration of dextran sulfate sodium (DSS) or dinitrobenzenesulfonic acid to C57BL/6 mice. Depression was induced by olfactory bulbectomy or chronic intracerebroventricular injection of reserpine. Colitis was reactivated by subsequent exposure to DSS or dinitrobenzenesulfonic acid. Some mice were given the antidepressant desmethylimipramine. Acute DSS-colitis was induced in mice lacking the alpha 7 subunit of the nicotinic acetylcholine receptor (alpha 7nAchR), and vagotomy was performed. Disease severity and colon tissue histology and inflammation were evaluated. Levels of C-reactive protein and proinflammatory cytokines were determined by enzyme-linked immunosorbent assay analysis of colon samples and macrophage culture. RESULTS: Induction of depression reactivated inflammation in mice in which colitis had been established and become quiescent. The induction was associated with impaired cholinergic inhibition of proinflammatory cytokine secretion by macrophages and mediated by alpha 7nAchR on these cells; macrophages isolated from depressed mice showed increased proinflammatory cytokine secretion. Depression-induced reactivation of colitis was prevented by desmethylimipramine and accompanied by a normalization of proinflammatory cytokine secretion. CONCLUSIONS: Depression reactivates dormant chronic colitis via the alpha 7nAchR. These findings encourage closer monitoring of behavior for signs of depression in IBD patients because treatment might prevent inflammatory conditions. Furthermore, alpha 7nAchR agonists might achieve this effect without the need for psychotropic medication.

Adoptive transfer of macrophage from mice with depression-like behavior enhances susceptibility to colitis.

BACKGROUND: Depression is common in patients with inflammatory bowel disease (IBD) but the pathway is not well understood. We examined whether the locus of susceptibility to colitis in mice with depression-like behavior (DLB) resides with the macrophage and implicates the vagus nerve. METHODS: Chronic colitis mimicking ulcerative colitis (UC) was induced by dextran sulfate sodium administered to C57BL/6-mice. Depression was induced by intracerebroventricular infusion of reserpine in healthy or vagotomized mice treated with antidepressant desmethylimipramine (DMI). Colitis was assessed macroscopically, histologically, and by C-reactive protein measurement in serum and by cytokines in colonic samples. Cytokine release was measured on macrophages isolated from these models. Naive macrophage colony-stimulating factor-deficient mice (op/op) were injected with peritoneal macrophages obtained from the different groups and acute colitis was induced. RESULTS: Vagotomy reactivated inflammation in mice with chronic colitis. DLB reactivated colitis and this was prevented by DMI only in mice with intact vagi. Macrophages isolated from vagotomized or DLB-mice showed a selective increase of proinflammatory cytokine release and this was not seen in macrophages isolated from DLB-DMI-treated mice; moreover, vagotomy abolished this beneficial effect. In op/op, adoptive transfer of macrophages from non-DLB mice significantly increased the inflammatory markers. These parameters were significantly increased when transferred with macrophages isolated from DLB or VXP mice. Op/op mice that received macrophages from DLB-DMI-treated mice showed a significant decrease of all parameters and vagotomy abolished this effect. CONCLUSIONS: These data identify the critical role of macrophage in linking depression and susceptibility to intestinal inflammation via the vagus nerve. The results provide a basis for developing new approaches to the management of UC patients with coexisting depression by rebalancing cytokine production by the cell.

Role of gut-brain axis in persistent abnormal feeding behavior in mice following eradication of Helicobacter pylori infection.

Bacterial infection can trigger the development of functional GI disease. Here, we investigate the role of the gut-brain axis in gastric dysfunction during and after chronic H. pylori infection. Control and chronically H. pylori-infected Balb/c mice were studied before and 2 mo after bacterial eradication. Gastric motility and emptying were investigated using videofluoroscopy image analysis. Gastric mechanical viscerosensitivity was assessed by cardioautonomic responses to distension. Feeding patterns were recorded by a computer-assisted system. Plasma leptin, ghrelin, and CCK levels were measured using ELISA. IL-1beta, TNF-alpha, proopiomelanocortin (POMC), and neuropeptide Y mRNAs were assessed by in situ hybridizations on frozen brain sections. Gastric inflammation was assessed by histology and immunohistochemistry. As shown previously, H. pylori-infected mice ate more frequently than controls but consumed less food per bout, maintaining normal body weight. Abnormal feeding behavior was accompanied by elevated plasma ghrelin and postprandial CCK, higher TNF-alpha (median eminence), and lower POMC (arcuate nucleus) mRNA. Infected mice displayed delayed gastric emptying and visceral hypersensitivity. Eradication therapy normalized gastric emptying and improved gastric sensitivity but had no effect on eating behavior. This was accompanied by persistently increased TNF-alpha in the brain and gastric CD3(+) T-cell counts. In conclusion, chronic H. pylori infection in mice alters gastric emptying and mechanosensitivity, which improve after bacterial eradication. A feeding pattern reminiscent of early satiety persists after H. pylori eradication and is accompanied by increased TNF-alpha in the brain. The results support a role for altered gut-brain pathways in the maintenance of postinfective gut dysfunction.

Vagus nerve integrity and experimental colitis.

Previous studies have identified a counterinflammatory vagal reflex in the context of endotoxic shock. We have extended this observation to show that the vagus confers protection against acute (5 days) colitis induced by dextran sodium sulfate (DSS) or by dinitrobenzene sulfonic acid (DNBS). We have shown that this is mediated via macrophages and involves the suppression of proinflammatory cytokines. In this study, we have examined whether the vagal integrity confers long-lasting protection by studying DNBS- and DSS-induced inflammatory responses in the colon at 9 to 61 days postvagotomy. The integrity of vagotomy was confirmed at all time points using CCK-induced satiety. As previously described in a DNBS and DSS model, vagotomy associated with the pyloroplasty increased all indices of inflammation. Vagotomy increased the disease activity index as well as the macroscopic and histological scores by 75 and 41%, respectively. In addition, myeloperoxidase (MPO) activity, serum levels of C-reactive protein (CRP), and colonic tissue levels of proinflammatory cytokine increased when colitis was induced 9 days postvagotomy. However, these increases in inflammatory indices were substantially diminished in mice with colitis induced 21, 33, and 61 days postvagotomy. This was accompanied by an increased production of interleukin-10, transforming growth factor-beta, Forkhead Box P3 (FOXP3) staining in colonic tissue, and serum corticosterone. These findings indicate that although vagal integrity is an important protective factor, other counterinflammatory mechanisms come into play if vagal integrity is compromised beyond 2 wk.

Cytokine modulation of muscarinic receptors in the murine intestine.

The extent to which gut motility and smooth muscle contractility are altered by intestinal inflammation depends on the nature of the underlying immune activation. The muscarinic receptor on smooth muscle plays a critical role in mediating acetylcholine-driven motor function. We examined the ability of cytokines to influence muscarinic receptor characteristics on intestinal longitudinal muscle and related the findings to studies on carbachol-induced contraction. Cells were isolated from longitudinal muscle myenteric plexus (LMMP). Cytokine receptor expression, muscle contractility, and muscarinic agonist receptor characteristics were examined by agonist displacement of [N-methyl-(3)H]scopolamine ([(3)H]NMS) binding. The TGF-beta1 receptor (543 bp) and the IFN-gamma receptor 1 (660 bp) were identified on smooth muscle cells. Scatchard analysis revealed dissociation constant and maximum binding values for [(3)H]NMS of 2.6 nM and 2.4 x 10(4) sites/cell, respectively, in control cells. Nematode infection was accompanied by a reduction in inhibitory constant of the high-affinity sites (K(H)), and this was independent of signal transduction and activator of transcription 6. Preincubation with TGF-beta1 enhanced longitudinal muscle contractility and decreased the K(H) to 2.2 pM (increased muscarinic receptor affinity), whereas preincubation with IFN-gamma increased the K(H) to 0.4 muM (decreased muscarinic receptor affinity) and decreased longitudinal muscle contractility. Preincubation of LMMP with IL-13 decreased the K(H) to 0.2 nM. Cytokines exert differential effects on the muscarinic receptor on intestinal longitudinal smooth muscle. These findings explain the basis for altered muscle contractility observed in Th1 and Th2 models of inflammation, as well as in the post-nematode-infected state.

The protective effect of the vagus nerve in a murine model of chronic relapsing colitis.

The vagus nerve inhibits the response to systemic administration of endotoxin, and we have recently extended this observation to show that the vagus attenuates acute experimental colitis in mice. The purpose of the present study was to determine whether there is a tonic counterinflammatory influence of the vagus on colitis maintained over several weeks. We assessed disease activity index, macroscopic and histological damage, myeloperoxidase (MPO) activity, and Th1 and Th2 cytokine profiles in chronic colitis induced by administration of dextran sodium sulfate (DSS) in drinking water for three cycles during 5 days with 11 days of rest between each cycle (DSS 3, 2, 2%) in healthy and vagotomized C57BL/6 mice and in mice deficient in macrophage-colony stimulating factor (M-CSF). A pyloroplasty was performed in vagotomized mice. Vagotomy accelerated the onset and the severity of inflammation during the first and second but not the third cycle. Although macroscopic scores were not significantly changed, histological scores as well as MPO activity and colonic tissue levels of IL-1alpha, TNF-alpha, IFN-gamma, and IL-18 but not IL-4 were significantly increased in vagotomized mice compared with sham-operated mice that received DSS. In control mice (without colitis), vagotomy per se did not affect any inflammatory marker. Vagotomy had no effect on the colitis in M-CSF-derived macrophage-deficient mice. These results indicate that the vagus protects against acute relapses on a background of chronic inflammation. Identification of the molecular mechanisms underlying the protective role of parasympathetic nerves opens a new therapeutic avenue for the treatment of acute relapses of chronic inflammatory bowel disease.

Visceral pain perception is determined by the duration of colitis and associated neuropeptide expression in the mouse.

BACKGROUND: Even though inflammation is a traditional tool for the induction of hyperalgesia in many tissues, recent observations suggest that not all inflammatory processes produce this change. Tolerance to colorectal distension (CRD) is reduced in patients with acute ulcerative colitis but is increased in patients with chronic inflammatory bowel disease. This suggests that the nature of the inflammatory infiltrate influences visceral perception. AIM: To test this hypothesis by assessing responses to CRD in mice with mild, acute or chronic colitis. METHODS: CRD responses were measured in mice with mild non-specific colitis, and dextran sodium sulphate (DSS)-induced acute and chronic colitis. Responses were compared with tissue infiltrate and damage, interleukin (IL)1beta and myeloperoxidase (MPO) activity and substance P, beta-endorphin and micro opioid receptor (MOR) expression. RESULTS: Mild and acute colitis were associated with increased responsiveness to CRD. In contrast, CRD responses were not increased in mice with chronic colitis and this difference was not due to altered colonic wall compliance. MPO and IL1beta levels were greater in acute than in chronic colitis. Larger increases in tissue substance P were seen in acute than in chronic DSS, whereas CD4 T cells, beta-endorphin and MOR expression were evident only in chronic colitis. An inverse correlation was seen between substance P and MOR in these tissues. CONCLUSIONS: Acute colitis increased responsiveness to CRD and is accompanied by an acute inflammatory infiltrate and increased tissue substance P. Chronic DSS is accompanied by an increase in beta-endorphin and MOR expression, and CD4 T cells, but no change in compliance or CRD responses. We conclude that acute inflammation generates hyperalgesia, whereas chronic inflammation involves infiltration by lymphocytes accompanied by MOR and beta-endorphin up regulation, and this provides an antinociceptive input that restores normal visceral perception.

The vagus nerve: a tonic inhibitory influence associated with inflammatory bowel disease in a murine model.

BACKGROUND & AIMS: The recently proposed Inflammatory Reflex describes an interaction between the vagus nerve and peripheral macrophages, resulting in attenuation of proinflammatory cytokine release in response to systemic exposure to bacterial endotoxin. The purpose of this study was to determine whether a similar vagus/macrophage axis modulates the inflammatory responses in the colon in mice. METHODS: We assessed the Disease Activity Index (DAI), macroscopic and histologic damage, serum amyloid-P level, and myeloperoxidase activity in colitis induced by administration of dextran sodium sulfate (DSS) in healthy and vagotomized C57BL/6 and in mice deficient in macrophage-colony stimulating factor (M-CSF)-induced and in hapten-induced colitis. A pyloroplasty was performed in vagotomized mice. RESULTS: DAI, macroscopic and histologic scores, myeloperoxidase activity, levels of serum amyloid-P, and colonic tissue levels of interleukin (IL)-1beta, IL-6, and tumor necrosis factor-alpha were increased significantly in vagotomized mice 5 days post-DSS and 3 days after hapten-induced colitis compared with sham-operated mice that received DSS or the hapten. Pretreatment with nicotine significantly decreased each of these markers in vagotomized mice with DSS colitis, and all markers except DAI and IL-6 in sham-operated DSS-treated mice. Conversely, hexamethonium treatment significantly increased each of these markers in the sham-operated DSS-treated mice. Vagotomy had no effect on the colitis in M-CSF-deficient mice. CONCLUSIONS: The vagus nerve plays a counterinflammatory role in acute colitis via a macrophage-dependent mechanism, involving hexamethonium-sensitive nicotinic receptors. The identification of a counterinflammatory neural pathway would open new therapeutic avenues for treating acute exacerbations of inflammatory bowel disease.

Antidepressants attenuate increased susceptibility to colitis in a murine model of depression.

BACKGROUND & AIMS: Psychiatric factors may determine gastrointestinal health outcomes. Here, we used a model of depression based on neonatal maternal separation (MS) to identify alterations in gut physiology and to assess its association with increased sensitivity to experimental colitis in adulthood. We also examined whether antidepressant therapy attenuates the increased susceptibility to colitis. METHODS: C57BL/6 mouse pups were separated from mothers for 3 hours per day at 1-21 days of age. Maternally unseparated (US) litters served as controls. At 8 weeks of age mice were examined for changes in behavior, intestinal permeability, and sensitivity to colitis. Separate sets of MS and US mice were given either saline or the antidepressant desipramine 15 mg/kg once daily at 23-36 days of age. Testing of mice occurred at 8 weeks of age. RESULTS: Adult MS mice showed evidence of depressive-like behavior and enhanced intestinal permeability but showed no evidence of spontaneous inflammation. A more severe colitis was seen in MS compared with US mice. Antidepressant therapy improved parameters of depressive-like behavior and reduced the vulnerability to dextran sulphate sodium colitis in MS mice but had no effect on colitis in US mice. CONCLUSIONS: MS may lead to depression and increased responsiveness to stress, to impaired intestinal barrier function, and to enhanced vulnerability to colitis in adulthood. This vulnerability is reversed by antidepressant therapy. Depression increases vulnerability to intestinal inflammation. We speculate that pre-existing depression may facilitate the expression of inflammatory bowel diseases.

CD4+ T-cell modulation of visceral nociception in mice.

BACKGROUND & AIMS: Although inflammatory and immune cells are present in the gut in the absence of pathology, their presence does not result in sensitization of sensory nerves, implying the existence of a local antinociceptive influence. We hypothesized that a component of the immune system exerts an antinociceptive influence, thus enabling the gut to function in the absence of undue pain or discomfort. METHODS: Visceromotor responses to colorectal distention were measured in mice with severe combined immune deficiency (SCID) and their wild-type controls. RESULTS: SCID mice exhibited significantly lower pain thresholds. Transfer of CD4(+) T, but not B lymphocytes, normalized visceral pain in these mice. The restoration of normal visceral nociception following T-cell reconstitution in SCID mice was blocked by naloxone methiodide. Using an enzyme immunoassay and immunohistochemistry for beta-endorphin, we showed that in vitro stimulation of T lymphocytes induced the synthesis and release of beta-endorphin and that transfer of T cells into SCID mice increased the expression of beta-endorphin in the enteric nervous system. CONCLUSIONS: These findings indicate that the immune system is a critical determinant of visceral nociception and that T lymphocytes provide an important opioid-mediated antinociceptive influence in the gut.

Mechanisms underlying the maintenance of muscle hypercontractility in a model of postinfective gut dysfunction.

BACKGROUND & AIMS: Acute gastroenteritis is a strong risk factor for the development of irritable bowel syndrome (IBS). We have developed an animal model in which transient acute infection leads to persistent muscle hypercontractility. Here, we investigate the mechanisms underlying the maintenance of this hypercontractility in the postinfective (PI) state. METHODS: Muscle contraction and messenger RNA (mRNA) or protein expression of cytokines were examined from jejunal longitudinal muscle cells of NIH Swiss mice infected with Trichinella spiralis or incubated with or without cytokines. RESULTS: During acute infection, interleukin (IL)-4 or IL-13, transforming growth factor (TGF)-beta1, and cyclooxygenase (COX)-2 were increased in the muscle layer ( P < .05). In the PI phase of the model, T helper (Th)2 cytokines returned to normal, but TGF-beta1 remained in the muscle ( P < .05). Exposure of muscle cells to IL-4 or IL-13 increased TGF-beta1 ( P < .01), COX-2 protein, and prostaglandin (PG)E 2 . Exposure of muscle cells to TGF-beta1 increased PGE 2 ( P < .05) and COX-2 protein. Incubation of tissue with IL-4, IL-13, TGF-beta1, or PGE 2 enhanced carbachol-induced muscle cell contractility ( P < .05). COX-2 inhibitor attenuated TGF-beta1-induced muscle hypercontractility ( P < .05). CONCLUSIONS: These results support the hypothesis that Th2 cytokines induce muscle hypercontractility during infection by a direct action on smooth muscle. The maintenance of hypercontractility results from Th2 cytokine-induced expression of TGF-beta1 and the subsequent up-regulation of COX-2 and PGE 2 at the level of the smooth muscle cell. We propose that PI gut dysfunction reflects mediator production in the neuromuscular tissues and that this may occur in PI-IBS.

Interleukin-4- and -13-induced hypercontractility of human intestinal muscle cells-implication for motility changes in Crohn's disease.

Crohn's disease is an idiopathic inflammatory condition. However, little is known about the changes that occur in the muscularis externa, despite the fact that this tissue contributes to motility changes and stricture formation. We characterized immune activity in the muscularis externa from intestinal segments of Crohn's disease patients and evaluated the role of IL-4 and -13 as well as signal transducer and activator of transcription (STAT)6 in the contractility of the cultured human intestinal smooth muscle cells. CD3+ve cells (P < 0.01) and IL-4 protein (P < 0.01) were significantly increased in the muscularis externa of Crohn's disease patients compared with noninflamed controls. Preincubation of human cultured smooth muscle cells with IL-4 (P < 0.001) or IL-13 (P < 0.05) significantly enhanced carbachol-induced contraction, and this was significantly inhibited by the STAT6 inhibitor leflunomide (P < 0.0001). A similar profile was observed in muscle cells isolated from Crohn's disease patients. Both IL-4 and IL-13 increased specific STAT6-DNA binding in control cells, and this was inhibited by anti-STAT6 Ab (P < 0.05) or leflunomide (P < 0.05). IL-4 and IL-13 mediate the hypercontractility of intestinal muscle via a STAT6 pathway at the level of the smooth muscle cell. The STAT6 pathway may contribute to the hypercontractility of intestinal muscle in Crohn's disease.

Role of IL-4, IL-13, and STAT6 in inflammation-induced hypercontractility of murine smooth muscle cells.

T helper 2 (Th2) cytokines interleukin (IL)-4 and IL-13, which activate signal transducer and activator of transcription 6 (STAT6) are expressed in the muscularis externa during nematode infection and are candidate mediators of the associated hypercontractility. To determine the locus of action of these cytokines, we examined the IL-4- and IL-13-induced hypercontractility of the isolated muscle cells from STAT6 +/+ and STAT6 -/- mice. We compared the results with cells isolated from Trichinella spiralis-infected STAT6 +/+ and STAT6 -/- mice. Carbamylcholine chloride (Carbachol) induced the contraction of jejunal muscle cells in a concentration-dependent manner maximal contraction (R(max) 26.7 +/- 1.9%). Cells from T. spiralis-infected STAT6 -/- mice showed the hypertrophy (cell lengths 41.4 +/- 0.8 to 89.0 +/- 8.7 microm) and hypercontractility (R(max) 37.5 +/- 1.3%) induced by infection. IL-4Ralpha mRNA was detected in dispersed smooth muscle cells. Incubation of longitudinal muscle-myenteric plexus (LMMP) with IL-4 and IL-13 enhanced Carbachol-induced muscle contraction (R(max) 35.5 +/- 1.9 and 32.4 +/- 2.9%, respectively). Incubation of LMMP from STAT6 -/- mice with IL-4 did not enhance the contraction. The hypercontractility in T. spiralis-infected mice was attenuated in STAT6 -/- mice (P < 0.02). These results indicate both IL-4 and IL-13 induce hypercontractility of muscle cells via the STAT6 pathway, and this is the basis for hypercontractility observed in T. spiralis-infected mice.

Immune-mediated neural dysfunction in a murine model of chronic Helicobacter pylori infection.

BACKGROUND & AIMS: Neuromuscular changes producing dysmotility and hyperalgesia may underlie symptom generation in functional gastrointestinal disorders. We investigated whether chronic Helicobacter pylori-induced gastritis causes neuromuscular dysfunction. METHODS: In vitro muscle contractility and acetylcholine release were evaluated in mice before and after H. pylori eradication. H. pylori colonization and gastritis were graded histologically. Substance P (SP)-, vasoactive intestinal polypeptide (VIP)-, and calcitonin gene-related peptide (CGRP) immunoreactivity (IR) and macrophages were studied by immunohistochemistry. RESULTS: In Balb/c mice, chronic H. pylori infection did not affect muscle function but augmented antral relaxation after nerve electric field stimulation. Infected mice had lower acetylcholine release by electric field stimulation and had higher density of SP-, CGRP-, and VIP-IR nerves in the stomach and of SP- and CGRP-IR in the spinal cord. Cholinergic nerve dysfunction worsened progressively and was associated with increasing macrophage and mononuclear but not polymorphonuclear infiltrate or bacterial colonization. SCID mice had unchanged acetylcholine release despite high H. pylori colonization and macrophage infiltration. Eradication of H. pylori normalized functional and morphologic abnormalities except for increased density of gastric SP- and CGRP-IR nerves. CONCLUSIONS: H. pylori infection induces functional and morphologic changes in the gastric neural circuitry that are progressive and lymphocyte dependent, and some persist after H. pylori eradication. The data have direct implications regarding the role of H. pylori infection in functional dyspepsia.

Visceral hyperalgesia and intestinal dysmotility in a mouse model of postinfective gut dysfunction.

BACKGROUND & AIMS: We established the concept that transient enteric infection may lead to persistent gut dysfunction, evident in vitro, in nematode-infected mice. The present study determined whether gut dysfunction in this model involves motor and sensory changes reminiscent of changes found in patients with postinfective irritable bowel syndrome (PI-IBS) and investigated underlying mechanisms. METHODS: Mice infected up to 70 days previously with Trichinella spiralis (Tsp) underwent videofluoroscopy with image analysis to assess upper gastrointestinal motility. Pseudoaffective responses to colorectal distention (CRD) were assessed using a barostat and validated by single fiber recordings from spinal nerves during CRD. Tissues were examined at different time points for histology, immunohistochemistry, and cytokine analysis. Some mice received dexamethasone intraperitoneally on days 23-25 PI or Tsp antigen orally on days 29, 43, and 57 PI. RESULTS: From day 28 PI, no discernible inflammation was present in the gut. Frequency and propagation velocity of intestinal contractions decreased, and retroperistalsis increased at days 28 to 42 PI. CRD induced an allodynic and hyperalgesic response in PI mice, which was accompanied by increased single unit discharge. Gavage of Tsp antigen induced T-cell responses and sustained gut dysfunction for 70 days PI. Administration of dexamethasone postinfection normalized dysmotility and visceral hyperalgesia. CONCLUSIONS: Long-lasting gut dysmotility and hyperalgesia develop in mice after transient intestinal inflammation. These changes are maintained by luminal exposure to antigen and reversed by corticosteroid treatment. The findings prompt consideration of this as a model of PI-IBS.