Maternal high-fat diet prevents developmental programming by early-life stress.

Anxiety disorders and depression are well-documented in subjects exposed to adverse childhood events. Recently, maternal obesity and/or maternal consumption of high-fat diets (HFD) have been also proposed as risk factors for offspring mental health. Here using an animal model in rats, we explored the combinatorial effects of a maternal HFD (40% of energy from fat without impact on maternal weight; during gestation and lactation) and maternal separation (MS) in offspring. In the prefrontal cortex (PFC) of pups, MS led to changes in the expression of several genes such as Bdnf (brain derived neurotrophic factor), 5HT-r1a (serotonin receptor 1a) and Rest4 (neuron-restrictive silencer element, repressor element 1, silencing transcription factor (Rest), splicing variant 4). Surprisingly, perinatal HFD strongly attenuated the developmental alterations induced by MS. Furthermore, maternal HFD totally prevented the endophenotypes (anxiety, spatial memory, social behavior, hypothalamic-pituitary-adrenal (HPA) axis response to stress, hippocampal neurogenesis and visceral pain) associated with MS at adulthood. Finally, we also demonstrated that HFD intake reduced anxiety and enhanced maternal care in stressed dams. Overall, our data suggest that a HFD restricted to gestation and lactation, which did not lead to overweight in dams, had limited effects in unstressed offspring, highlighting the role of maternal obesity, rather than fat exposure per se, on brain vulnerability during development.

The hypersensitivity to colonic distension of IBS patients can be transferred to rats through their fecal microbiota.

BACKGROUND: Alterations of intestinal microbiota and hypersensitivity to colonic distension are two features of the irritable bowel syndrome (IBS). However, the role of intestinal microbiota in visceral hypersensitivity of IBS patients is far to be established. The aim of our study was to determine whether the intestinal microbiota is involved in the visceral hypersensitivity in IBS. METHODS: The painful response to colorectal distension and colonic mucosal parameters were assessed in gnotobiotic rats. Germfree (GF) rats were inoculated with the fecal microbiota from IBS patients characterized by hypersensitivity to colorectal distension (IBS HMA rats) or from non-hypersensitive healthy volunteers (Healthy HMA rats). Conventional rats were studied as normosensitivity control. Fecal microbial analyses were carried out in human and HMA rats fecal samples using cultural and molecular approaches. KEY RESULTS: The microbial dysbiosis of the IBS gut microbiota (more sulfate-reducing bacteria and Enterobacteriaceae and less bifidobacteria) could be maintained in gnotobiotic rats. The number of abdominal contractions in response to colorectal distensions was significantly higher in IBS HMA rats than in healthy HMA rats. No difference was observed between healthy HMA and conventional rats. Colorectal compliance, epithelial paracellular permeability, and density of colonic mucosal mast cells were similar in the three groups of rats. CONCLUSIONS & INFERENCES: We herein showed that sensitivity to colonic distension of IBS patients can be transferred to rats by the fecal microbiota. Mucosal alterations associated with microbiota transfer are not involved in this hypersensitivity. The altered IBS microbiota may have important role in the hypersensitivity characterizing IBS patients through specific bacterial metabolites.

Effects of octreotide on jejunal hypersensitivity triggered by Cryptosporidium parvum intestinal infection in an immunocompetent suckling rat model.

BACKGROUND: Similar to other bacterial or protozoan infections, human cryptosporidiosis may trigger postinfectious irritable bowel syndrome (IBS)-like symptoms, a condition in which enhanced visceral perception of pain during intestinal distension plays a pivotal role. In an immunocompetent suckling rat model which mimicks features of postinfectious IBS, Cryptosporidium parvum infection induces long-lasting jejunal hypersensitivity to distension in association with intestinal activated mast cell accumulation. The aim of the present study was to explore in this model whether octreotide, a somatostatin agonist analog, could prevent the development of jejunal hypersensitivity and intestinal mast cell/nerve fiber accumulation. METHODS: Five-day-old Sprague-Dawley rats were infected with C. parvum and treated 10 days later with octreotide (50 g kg(-1) day(-1), i.p.) for 7 days. KEY RESULTS: Compared with untreated infected rats, octreotide treatment of infected rats resulted in increased weight gain [day 23 postinfection (PI)], decreased food intake (day 16 PI), and a reduction in jejunal villus alterations (day 14 PI), CD3(+) IEL (day 37 PI) and mast cell (days 37 and 50 PI) accumulations, nerve fiber densities (day 50 PI), and hypersensitivity to distension (day 120 PI). In uninfected rats, the effects of octreotide treatment were limited to higher weight gain (days 16 and 23 PI) and decreased food intake (day 23 PI) compared with uninfected-untreated rats. CONCLUSIONS & INFERENCES: Data confirms the relevance of the present rat model to postinfectious IBS studies and prompt further investigation of somatostatin-dependent regulatory interactions in cryptosporidiosis.

Maternal deprivation alters epithelial secretory cell lineages in rat duodenum: role of CRF-related peptides.

OBJECTIVE: Chronic psychological stress is associated with development of intestinal barrier dysfunction and impairs host defence mechanisms. The intestinal epithelium, consisting of enterocytes, endocrine cells, goblet cells and Paneth cells, is an important component of this barrier. In the present study, the impact of maternal deprivation (MD) on secretory lineages of duodenal epithelium and the involvement of the peripheral corticotropin-releasing factor (CRF) pathway were investigated. METHODS: Rat pups were deprived of their dam for 3 h/day (days 5-20). Non-deprived pups served as controls. On days 8, 13, 20, 24, 34, 44 and 84, duodenal tissues were collected for quantitative real-time PCR and immunohistochemistry studies. RESULTS: MD induced a sustained decrease in the number of Paneth and goblet cells but hyperplasia of endocrine cells. These alterations were associated with a duodenal increase of CRF, urocortin 2 and CRF receptor subtype 2 (CRFR(2)) mRNA, whereas CRFR(1) expression was decreased. The effects of MD on intestinal epithelium were inhibited by the CRFR(1)/R(2) antagonist astressin injected daily before MD. Studies using specific receptor antagonists in rats subjected to MD revealed that CRFR(1) was involved in the hyperplasia of endocrine cells and CRFR(2) in the depletion of Paneth cells. Conversely, daily injection of CRF and of the CRFR(2) agonist urocortin 2 in control rats resulted in changes in epithelial differentiation similar to MD. CONCLUSIONS: The activation of CRFR(1) and CRFR(2) induced by MD markedly altered the quantitative distribution of secretory cells of the intestinal epithelium. These alterations, in particular the depletion of Paneth and goblet cells, may create conditions leading to the development of an epithelial barrier defect.

Guanylate cyclase C-mediated antinociceptive effects of linaclotide in rodent models of visceral pain.

BACKGROUND Linaclotide is a novel, orally administered investigational drug currently in clinical development for the treatment of constipation-predominant irritable bowel syndrome (IBS-C) and chronic idiopathic constipation. Visceral hyperalgesia is a major pathophysiological mechanism in IBS-C. Therefore, we investigated the anti-nociceptive properties of linaclotide in rodent models of inflammatory and non-inflammatory visceral pain and determined whether these pharmacological effects are linked to the activation of guanylate cyclase C (GC-C). METHODS Orally administered linaclotide was evaluated in non-inflammatory acute partial restraint stress (PRS) and acute water avoidance stress (WAS) models in Wistar rats, and in a trinitrobenzene sulfonic acid (TNBS)-induced inflammatory model in Wistar rats and GC-C null mice. KEY RESULTS In TNBS-induced colonic allodynia, linaclotide significantly decreased the number of abdominal contractions in response to colorectal distension without affecting the colonic wall elasticity change in response to distending pressures after TNBS. However, linaclotide had no effect on visceral sensitivity under basal conditions. In addition, linaclotide significantly decreased colonic hypersensitivity in the PRS and WAS models. In wild type (wt) and GC-C null mice, the instillation of TNBS induced similar hyperalgesia and allodynia. However, in post-inflammatory conditions linaclotide significantly reduced hypersensitivity only in wt mice, but not in GC-C null mice. CONCLUSIONS & INFERENCES These findings indicate that linaclotide has potent anti-nociceptive effects in several mechanistically different rodent models of visceral hypersensitivity and that these pharmacological properties of linaclotide are exerted through the activation of the GC-C receptor. Therefore, linaclotide may be capable of decreasing abdominal pain in patients suffering from IBS-C.

Lactobacillus farciminis treatment attenuates stress-induced overexpression of Fos protein in spinal and supraspinal sites after colorectal distension in rats.

Abstract Irritable bowel syndrome (IBS), frequently associated with psychological distress, is characterized by hypersensitivity to gut wall distension. Some probiotics are able to alleviate IBS symptoms and reduce visceromotor response to mechanical stimuli in animals. Moreover, we have previously shown that Lactobacillus farciminis treatment abolished the hyperalgesia to colorectal distension (CRD) induced by acute stress. The aims of the present study were to determine whether (i) stress-induced visceral hyperalgesia modifies the expression of Fos, a marker of general neuronal activation, induced by CRD, (ii) this activation can be modulated by L. farciminis treatment. Female rats were treated by L. farciminis and CRD was performed after partial restraint stress (PRS) or sham-PRS. The expression of Fos protein was measured by immunohistochemistry. After CRD or PRS, Fos expression was increased in spinal cord section (S1), nucleus tractus solitarius (NTS), paraventricular nucleus (PVN) of the hypothalamus, and in the medial nucleus of the amygdala (MeA). The combination of both stimuli, PRS and CRD, markedly increased this Fos overexpression in the sacral spinal cord section, PVN and MeA, but not in NTS. By contrast, a pretreatment with L. farciminis significantly reduced the number of Fos positive cells in these area. This study shows that PRS enhances Fos protein expression induced by CRD at the spinal and supraspinal levels in rats. Lactobacillus farciminis treatment inhibited this enhancing effect, suggesting that the antinociceptive effect of this probiotic strain results from a decrease of the stress-induced activation/sensitization of sensory neurons at the spinal and supraspinal level.

Protease-activated receptor 2 and gut permeability: a review.

Digestive tract proteases are best known for their proteolytic activity in the digestion of alimentary proteins. However, during the last decade, a possible role of proteases as signalling molecules has been emphasized with the discovery of a novel class of G-protein coupled receptors located on cell membranes that may be activated by proteolytic cleavage of their N-terminal extracellular domain. Type 2 protease-activated receptors (PAR-2) are cleaved by serine-proteases such as trypsin and tryptase. PAR-2 is present in many intestinal cell types and particularly on epithelial cells. Multiple functions have been demonstrated in the gut for PAR-2, including epithelial permeability, mainly the intercellular permeability that is of paramount importance in the equilibrium between the external milieu (digestive contents) and the submucosal immune system. Alterations of both tissue and luminal levels of proteases or serine-protease activity may affect gut permeability and subsequently the immune status of the mucosa. Activation of PAR-2 on epithelial cells may directly affect cytoskeleton contraction by triggering phosphorylation of myosin light chain with subsequent changes in tight junction permeability. Enhanced fecal protease level has been recently reported in both organic (ulcerative colitis) and functional (irritable bowel syndrome) intestinal disorders and may play a role in the pathogenesis of such diseases.

Increased faecal serine protease activity in diarrhoeic IBS patients: a colonic lumenal factor impairing colonic permeability and sensitivity.

OBJECTIVES: Diarrhoea-predominant irritable bowel syndrome (IBS-D) is characterised by elevated colonic lumenal serine protease activity. The aims of this study were (1) to investigate the origin of this elevated serine protease activity, (2) to evaluate if it may be sufficient to trigger alterations in colonic paracellular permeability (CPP) and sensitivity, and (3) to examine the role of the proteinase-activated receptor-2 (PAR-2) activation and signalling cascade in this process. PATIENTS AND METHODS: Faecal enzymatic activities were assayed in healthy subjects and patients with IBS, ulcerative colitis and acute infectious diarrhoea. Following mucosal exposure to supernatants from control subjects and IBS-D patients, electromyographic response to colorectal balloon distension was recorded in wild-type and PAR-2(-/-) mice, and CPP was evaluated on colonic strips in Ussing chambers. Zonula occludens-1 (ZO-1) and phosphorylated myosin light chain were detected by immunohistochemistry. RESULTS: The threefold increase in faecal serine protease activity seen in IBS-D patients compared with constipation-predominant IBS (IBS-C) or infectious diarrhoea is of neither epithelial nor inflammatory cell origin, nor is it coupled with antiprotease activity of endogenous origin. Mucosal application of faecal supernatants from IBS-D patients in mice evoked allodynia and increased CPP by 92%, both of which effects were prevented by serine protease inhibitors and dependent on PAR-2 expression. In mice, colonic exposure to supernatants from IBS-D patients resulted in a rapid increase in the phosphorylation of myosin light chain and delayed redistribution of ZO-1 in colonocytes. CONCLUSIONS: Elevated colonic lumenal serine protease activity of IBS-D patients evokes a PAR-2-mediated colonic epithelial barrier dysfunction and subsequent allodynia in mice, suggesting a novel organic background in the pathogenesis of IBS.

Long-term alterations of colonic nerve-mast cell interactions induced by neonatal maternal deprivation in rats.

BACKGROUND: Neonatal maternal deprivation induces colonic alterations in adult rats, such as hypersensitivity to distension or an increase in paracellular permeability, characteristics of irritable bowel syndrome (IBS) patients. Recent studies described neuroimmune alterations in the colonic mucosa of IBS patients. METHODS: Male Wistar rats were submitted to maternal deprivation for 3 h daily during postnatal days 2-14, and were sacrificed at 4 or 12 weeks of age. Control pups were left undisturbed with their dam. RESULTS: Colonic mast cell hyperplasia was observed at 4 and 12 weeks in maternally deprived rats, and was associated with an increase in protease content. Mucosal nerve fibre density assessed by protein gene product (PGP) 9.5 immunoreactivity was increased at 12 weeks but not at 4 weeks, while calcitonin gene-related protein (CGRP)-immunoreactive fibres remain constant. Synaptogenesis assessed by synaptophysin immunostaining was increased at 4 weeks but not at 12 weeks. The number of mast cells in close proximity to PGP 9.5- or CGRP-immunoreactive fibres was greater at both 4 and 12 weeks. Expression of neurokinin NK(1) receptors in the spinal cord was enhanced at 12 weeks. No significant change in total mast cell number, PGP 9.5 immunoreactivity and mast cells associated with PGP 9.5-immunoreactive fibres was observed in the jejunum. Treatment of pups with anti-nerve growth factor (NGF) antibodies abolished the increases in synaptogenesis and in the number of mast cells in close proximity to nerve fibres observed 4 weeks after maternal deprivation. CONCLUSIONS: Neonatal maternal deprivation induces closer association of colonic mast cells with nerves, which is similar to that seen in IBS patients. NGF is a possible mediator of this effect.

In vivo and transgenic animal models used to study visceral hypersensitivity.

Measurement of visceral sensitivity in animals is mainly based on 'pseudoaffective' responses, which are brain stem reflexes. For example, in female, but not male rats, acute partial restraint stress induces hypersensitivity to colorectal distension. Mucosal mast cell density increases in rats after nematode infection or maternal deprivation, and both also induce colon hypersensitivity. Significantly, the proximity between nerves and mast cells has been found to be increased in adult rats submitted to maternal deprivation. Protease activation of the proteinase-activated receptor-2 also increases visceral nociception in rats, suggesting that an increase in paracellular permeability may be the primum movens in several animal models of visceral hypersensitivity. Accumulating evidence suggests that sensitization of visceral afferents is not restricted to the presumed nociceptor population, suggesting that most of the mechanosensitive afferent population can contribute to visceral discomfort and pain. Other inflammation-produced changes (e.g. subunit composition of purine-gated P2X channels) in visceral sensory neurones may also contribute to visceral hypersensitivity. This article discusses use of in vivo strategies (and transgenic mouse models) to reveal putative roles in mechanosensitivity and sensitization for molecules not previously considered to have mechanosensory functions.

Colonic luminal proteases activate colonocyte proteinase-activated receptor-2 and regulate paracellular permeability in mice.

Luminal activation of protease-activated receptors-2 (PAR(2)) on colonocytes by trypsin or PAR(2)-activating peptide increases colonic paracellular permeability (CPP). The aim of this study was to evaluate the role of proteases from endogenous and bacterial origin in the modulation of CPP and colonocyte PAR(2) expression in mice. CPP was assessed with (51)Cr-EDTA after intracolonic administration of different protease inhibitors. After 12 days of oral antibiotic treatment, measurements of colonic luminal serine protease activity (CLSPA), CPP, mucosal mouse mast cell proteinase-1 (MMCP-1) content, immunochemistry of PAR(2) and assessment of effects of PAR(2) agonist (SLIGRL) and mast cell degranulator (C48/80) on CPP in Ussing chambers were performed. Immunochemistry was repeated after intracolonic trypsin administration. Colonic infusion of protease inhibitors significantly reduced CPP. In antibiotic-treated mice, CLSPA was reduced coupled with a decrease in PAR(2) expression, but with no change in CPP and MMCP-1 content. Trypsin administration restored PAR(2) expression. The increase in CPP induced by SLIGRL and C48/80 was reduced after antibiotic treatment. Protease activity of colonic content plays an important role in the regulation of mucosal barrier through activation of PAR(2).

Involvement of cannabinoid receptors in inflammatory hypersensitivity to colonic distension in rats.

Activation of cannabinoid CB1 and CB2 receptors is known to attenuate nociception and hyperalgesia in somatic inflammatory conditions. The aim of this study was to determine whether cannabinoids modulate colonic sensitivity in basal and inflammatory conditions. The effects of CB1 and CB2 receptor agonists and antagonists on the abdominal contractile response to colorectal distension (CRD) in basal conditions and after 2,4,6-trinitrobenzenesulphonic acid-induced colitis were investigated. As previously described, colitis triggered a hypersensitivity to CRD. In basal conditions, both CB1 (WIN 55212-2) and CB2 (JWH 015) agonists reduced the abdominal response to CRD at a dose of 1 mg kg(-1), i.p. Both compounds were active at a lower dose (0.1 mg kg(-1)) abolishing the hypersensitivity induced by colitis. Administered alone, CB1 (Rimonabant) and CB2 (SR 144528) receptor antagonists (10 mg kg(-1)) had no effect on basal sensitivity. In contrast, the CB1, but not the CB2, receptor antagonist enhanced colitis-induced hyperalgesia. It is concluded that colonic inflammation enhances the antinociceptive action of CB1 and CB2 receptor agonists, and activates an endogenous, CB1 receptor mediated, antinociceptive pathway.

Lactobacillus farciminis treatment suppresses stress induced visceral hypersensitivity: a possible action through interaction with epithelial cell cytoskeleton contraction.

BACKGROUND: Stress induced increase in colonic paracellular permeability results from epithelial cell cytoskeleton contraction and is responsible for stress induced hypersensitivity to colorectal distension (CRD). The probiotic Lactobacillus farciminis releases spontaneously nitric oxide (NO) in the colonic lumen in vivo and exerts anti-inflammatory effects. This study aimed: (i) to evaluate the effects of L farciminis on stress induced hypersensitivity to CRD and increase in colonic paracellular permeability; and (ii) to ascertain whether these effects are NO mediated and related to changes in colonocyte myosin light chain phosphorylation (p-MLC). METHODS: Female Wistar rats received either 10(11) CFU/day of L farciminis or saline orally over 15 days before partial restraint stress (PRS) or sham-PRS application. Visceral sensitivity to CRD and colonic paracellular permeability was assessed after PRS or sham-PRS. Haemoglobin was used as an NO scavenger. Western blotting for MLC kinase, MLC, and p-MLC were performed in colonic mucosa from L farciminis treated and control rats after PRS or sham-PRS. RESULTS: PRS significantly increased the number of spike bursts for CRD pressures of 30-60 mm Hg as well as colonic paracellular permeability. L farciminis treatment prevented both effects, while haemoglobin reversed the protective effects of L farciminis. p-MLC expression increased significantly from 15 to 45 minutes after PRS, and L farciminis treatment prevented this increase. CONCLUSION: L farciminis treatment prevents stress induced hypersensitivity, increase in colonic paracellular permeability, and colonocyte MLC phosphorylation. This antinociceptive effect occurs via inhibition of contraction of colonic epithelial cell cytoskeleton and the subsequent tight junction opening, and may also involve direct or indirect effects of NO produced by this probiotic.

Phenotypic changes in colonocytes following acute stress or activation of mast cells in mice: implications for delayed epithelial barrier dysfunction.

BACKGROUND AND AIMS: Stressful life events are known to modulate the development or relapse of disease in both inflammatory bowel disease and irritable bowel disease patients but underlying mechanisms remain unclear. Stress is known to effect mast cells, interferon gamma (IFN-gamma), and myosin light chain phosphorylation to trigger colonic epithelial barrier dysfunction. The aim of this study was to investigate whether acute stress induced or chemical mast cell activation impaired expression and function of epithelial tight junctions, and altered colonocyte differentiation in mice. METHODS: Colonic paracellular permeability was assessed as the in vivo lumen to blood ratio of 51Cr-EDTA in different groups of mice (controls, stressed, mast cell degranulator BrX-537A treated), pretreated or not with the mast cell stabiliser doxantrazole. Involvement of mast cells and IFN-gamma was evaluated in wild-type and IFN-gamma deficient mice. Tight junction alteration was assessed by histology, transmission electron microscopy, and real time reverse transcription-polymerase chain reaction. Colonocyte differentiation was determined by protein kinase C zeta (PKCzeta) immunofluorescence and western blotting, and alkaline phosphatase activity assay. RESULTS: Acute stress induced a three day delayed increase in colonic paracellular permeability which involved mast cell degranulation and overproduction of IFN-gamma. The colonic epithelial barrier was morphologically altered and expression of mRNA encoding tight junction proteins ZO-2 and occludin was decreased. Moreover, three days after acute stress, colonocyte differentiation was reduced, as shown by decreased expression of both PKCzeta isotype and alkaline phosphatase. CONCLUSION: These data highlight new mechanisms whereby an acute stress acts on the gastrointestinal tract by inducing alterations in colonocyte differentiation and decreased expression of mRNA encoding tight junction proteins. Thus phenotypic changes in colonocytes could pave the way for stress related intestinal disorders.

Intracerebroventricular administration of PD 140.548 N-methyl-D-glucamine attenuates the release of cortisol and catecholamines induced by duodenal distension in the sheep.

The aim of this study was to determine the influence of mechanically induced duodenal distension (DD) and PD 140.548 N-methyl-D-glucamine (a specific peptide antagonist of a CCK1 receptor) premedication on mechanographical reticulo-ruminal activity, animal general behaviour, catecholamines (CA) and the blood plasma cortisol levels, as well as the clinical symptoms of visceral pain induced by DD in sheep. After 24 h fasting, 6 animals, Polish merino sheep were praeanaesthetised by i.m. injection of ketamine (20 mg x kg(-1) b.w.) and anaesthetised with i.v. infusion of pentobarbital (20 mg x kg(-1) b.w.) and a permanent stainless steel cannula (gate cannula) was inserted inside the lateral cerebral ventricle (controlled by cerebrospinal fluid efflux) 10 mm above the bregma and 5 mm laterally from the midline suture using stereotaxic method. Under the same general anaesthesia and analgesia a T-shaped silicon cannula, was inserted into the duodenum (12 cm from pylorus) and a second one was inserted into the dorsal sac of the rumen. During 7 consecutive days after surgery each animal was treated i.m. with procaine penicillin (300000 I.U..kg(-1) b.w.), dihydrostreptomycine (DHS, 10 microg x kg(-1) b.w.), prednisolone acetate 1.2 mg x kg(-1) b.w.) together and i.m. injection of ketamine (20 mg x kg(-1) b.w.), separetely. The influence of PD 140.548 N-methyl-D-glucamine on the unfavourable effects of duodenal distension using a 10 cm long balloon filled with 40 and 80 ml (DD40 and DD80) water at animal body temperature was investigated in this study. Five minutes DD40 and DD80 caused an immediate and compete inhibition of the reticulo-ruminal frequency, a significant increase in plasma CA and cortisol levels, an increase in the heart rate, hyperventilation and other symptoms of pain, proportionally to the degree of intestinal distension. Intracerebroventricular (i.c.v.) administration of PD 140.548 alone at a dose of 0.25, 0.5, 1 or 2 mg in toto did not significantly change the reticulo-ruminal motility, CA and cortisol concentrations, but 10 min after the i.c.v. infusion (or 10 min before DD) at a dose 1 and 2 mg in toto , it completely blocked the increase of blood plasma cortisol, epinephrine (E), norepinephrine (NE) and dopamine (DA) concentrations for 20 min. In the some time it prevented the reticulo-ruminal atony provocked by DD. It is concluded that PD 140.548 N-methyl-D-glucamine--an antagonist of the central CCK1 receptor can be an effective analgesic agent in duodenal pain. This action is due to the inhibition of peripheral CCK1 type receptor in the central descending nerve pathway, facilitating pain transmission in sheep perhaps in the hypothalamic-pituitary-adrenal axis.

Review article: bifidobacteria as probiotic agents -- physiological effects and clinical benefits.

Bifidobacteria, naturally present in the dominant colonic microbiota, represent up to 25% of the cultivable faecal bacteria in adults and 80% in infants. As probiotic agents, bifidobacteria have been studied for their efficacy in the prevention and treatment of a broad spectrum of animal and/or human gastrointestinal disorders, such as colonic transit disorders, intestinal infections, and colonic adenomas and cancer. The aim of this review is to focus on the gastrointestinal effects of bifidobacteria as probiotic agents in animal models and man. The traditional use of bifidobacteria in fermented dairy products and the GRAS ('Generally Recognised As Safe') status of certain strains attest to their safety. Some strains, especially Bifidobacterium animalis strain DN-173 010 which has long been used in fermented dairy products, show high gastrointestinal survival capacity and exhibit probiotic properties in the colon. Bifidobacteria are able to prevent or alleviate infectious diarrhoea through their effects on the immune system and resistance to colonization by pathogens. There is some experimental evidence that certain bifidobacteria may actually protect the host from carcinogenic activity of intestinal flora. Bifidobacteria may exert protective intestinal actions through various mechanisms, and represent promising advances in the fields of prophylaxis and therapy.

Neonatal maternal deprivation triggers long term alterations in colonic epithelial barrier and mucosal immunity in rats.

BACKGROUND: Stressful events in the early period of life (for example, maternal deprivation) have been shown to modify adult immune and gastrointestinal tract functions. The present study aimed to establish whether maternal deprivation affects colonic epithelial barrier and the development of an experimental colitis in adult rats. METHODS: Male Wistar rat pups were separated during postnatal days 2-14 or left undisturbed with their dam. At 12 weeks of age, we assessed colonic paracellular permeability, bacterial translocation, myeloperoxidase (MPO) activity, mucosal mast cell density, cytokine (interleukin (IL)-1 beta, IL-2, IL-4, IL-10, and interferon gamma (IFN-gamma)) mRNA expression, and macroscopic damage. Total gut permeability, MPO activity, and macroscopic damage were also assessed four days after intracolonic administration of 2,4,6-trinitrobenzenesulphonic acid (TNBS). RESULTS: Maternal deprivation triggered a significant increase in colonic permeability associated with bacterial translocation into the mesenteric lymph nodes, liver, and spleen. These alterations were associated with some macroscopic damage and an increase in colonic MPO activity, mucosal mast cell density, and cytokine mRNA expression. Intracolonic infusion of TNBS induced a significantly higher inflammatory reaction in separated animals, as judged by enhanced MPO colonic levels, total gut permeability, and macroscopic lesions. CONCLUSIONS: Maternal deprivation promotes long term alterations in the colonic epithelial barrier associated with an exaggerated immune response to an external immune stimulus. This suggests a role for early psychological factors in the regulation of colonic mucosal barrier in later life.

Nitric oxide released by Lactobacillus farciminis improves TNBS-induced colitis in rats.

BACKGROUND: Beneficial effects of lactobacilli have been reported in experimental colitis. On the other hand, despite the controversial role of nitric oxide (NO) in the inflammatory gut process, a protective action of exogenous NO in inflammation has been suggested. Consequently, this study aimed to determine the effect of (i) sodium nitroprusside (SNP), a NO donor and (ii) treatment with Lactobacillus farciminis, which produces NO in vitro, on trinitrobenzene sulphonic acid (TNBS)-induced colitis in rats and to evaluate the role of exogenous NO in this effect. METHODS: Rats were divided into three groups receiving one of the following: (i) a continuous intracolonic (IC) infusion of SNP for 4 days, (ii) L. farciminis orally for 19 days, or (iii) saline. On day 1 and day 15, respectively, TNBS and saline were administrated IC, followed by a continuous IC infusion of saline or haemoglobin, a NO scavenger. At the end of treatments, the following parameters were evaluated: macroscopic damage of colonic mucosa, myeloperoxidase and nitric oxide synthase activities and colonic luminal NO production. RESULTS: In colitic rats, SNP and L. farciminis treatment significantly (P < 0.05) reduced macroscopic damage scores, myeloperoxidase and nitric oxide synthase activities compared to controls. Haemoglobin infusion abolished the anti-inflammatory effect of both NO donor treatments, but had no effect per se on colitis. CONCLUSION: NO released intraluminally by SNP infusion or by L. farciminis given orally improves TNBS-induced colitis in rats. These results indicate a protective role of NO donation in colonic inflammation and show for the first time a mechanism involving NO delivery by a bacterial strain reducing an experimental colitis.

Colonic responses to Lactobacillus farciminis treatment in trinitrobenzene sulphonic acid-induced colitis in rats.

BACKGROUND: It has recently been shown that Lactobacillus farciminis treatment exerts an anti-inflammatory effect in trinitrobenzene sulphonic acid (TNBS)-induced colitis partly through a nitric oxide release by this strain. The aim of this study was to evaluate whether L. farciminis treatment shares also the general mechanisms of action involved in the beneficial effect of probiotics in the colonic inflammatory process. METHODS: Rats received L. farciminis for 15 days before and 4 days after intracolonic administration of TNBS or vehicle. The following parameters were evaluated: macroscopic damage of colonic mucosa, myeloperoxidase activity, cytokine mucosal levels, bacterial profile in colonic content and mucosa, bacterial translocation and colonic paracellular permeability. RESULTS: In the absence of TNBS, L. farciminis treatment reduced colonic paracellular permeability and increased the IL-10 level in the colonic wall. TNBS administration induced colonic macroscopic damage, associated with an increase of myeloperoxidase activity, bacterial translocation, colonic paracellular permeability and IL-1beta mucosal level, and a decrease in IL-10 mucosal level. Moreover, the bacterial profile of colonic content and mucosa was modified. All these alterations were abolished or significantly reduced by L. farciminis treatment. CONCLUSIONS: As previously shown, L. farciminis treatment improves TNBS-induced colitis. This study indicates that, in addition to the nitric oxide released by this bacterial strain, the anti-inflammatory action of L. farciminis involves also normalization of colonic microflora, prevention of bacterial translocation, enhancement of barrier integrity and a decrease in the IL-1beta mucosal level.

Influence of sex and experimental protocol on the effect of maternal deprivation on rectal sensitivity to distension in the adult rat.

Neonatal maternal separation induces visceral hyperalgesia before and after stress in male rats. This study compares the effects on sensitivity to rectal distension in adult male and female rats, using two protocols of deprivation. Between postnatal days 1 and 14, maternal deprivation was performed for 2 h per day according to a protocol of type M (removal of all pups from home cage) or type P (separation of half of littermates). Visceral sensitivity was assessed at 12 weeks of age by the number of abdominal contractions induced by rectal distension before and after restraint stress. Calcitonin gene-related peptide (CGRP) was identified in the rectal wall by immunohistochemistry. In basal conditions, both separation protocols induced hyperalgesia, that was greater after type M than type P, and in females than in males for type P separation. Acute restraint stress induced hyperalgesia in control females only, and this effect was similarly enhanced by both type P and M separation. No difference was found between controls and deprived rats in rectal CGRP immunoreactivity which was greater in females and increased after rectal distension. These results indicate that long-term visceral hyperalgesia depends upon the type of maternal deprivation and that females are more sensitive than males.