Association between jejunal remodeling in fasting rats and hypersensitivity of intestinal afferent nerves to mechanical stimulation.

Remodeling of Intestinal properties and hypersensitivity of intestinal afferents to mechanical stimulation were previously demonstrated in a fasting rat model. Other studies investigated the association between mechanical and histological remodeling during fasting. This study aimed to further explore the relationship between the jejunal remodeling and intestinal afferent hypersensitivity by combining afferent nerve recordings with histological and mechanical data. Eight male Sprague Dawley rats had no access to food for 7 days (Fasting group). Seven male rats served as controls (Control group). Jejunal segments were studied in vitro in an organ bath for analysis of afferent signaling and for analysis of mechanical and histomorphological parameters. Correlation analyses were done to analyze association between nerve activity (spike rate increase ratio, SRIR) at distension levels of 20, 40 and 80 cmH2O and mechanical stress and histomorphological changes of the jejunal segments. Compared with the Control group, the main findings in jejunal segments in the Fasting group were (1) Most histomorphometry parameters were reduced (P < 0.05-P < 0.001), (2) SRIR values were higher (P < 0.001), (3) The relative numbers of intermuscular and submucosal neurons were increased (P < 0.05-P < 0.01), and (4) SRIR was negatively correlated with intestinal wall thickness, circumferential muscle layer thickness and positively correlated with the inner residual strain, the number of neurons, and the mechanical stress. In conclusion, the fasting-induced histomorphological remodeling (reduced wall thickness and increased relative number of neurons) and biomechanical remodeling (residual strain changes and high stress level) of the intestine in fasting rats were associated with hypersensitivity of intestinal afferents. Afferent hypersensitivity appears to be dependent on stress rather than on strain.

Effect of NSAIDs Supplementation on the PACAP-, SP- and GAL-Immunoreactive Neurons in the Porcine Jejunum.

Side effects associated with nonsteroidal anti-inflammatory drugs (NSAIDs) treatment are a serious limitation of their use in anti-inflammatory therapy. The negative effects of taking NSAIDs include abdominal pain, indigestion nausea as well as serious complications such as bleeding and perforation. The enteric nervous system is involved in regulation of gastrointestinal functions through the release of neurotransmitters. The present study was designed to determine, for the first time, the changes in pituitary adenylate cyclase-activating polypeptide (PACAP), substance P (SP) and galanin (GAL) expression in porcine jejunum after long-term treatment with aspirin, indomethacin and naproxen. The study was performed on 16 immature pigs. The animals were randomly divided into four experimental groups: control, aspirin, indomethacin and naproxen. Control animals were given empty gelatin capsules, while animals in the test groups received selected NSAIDs for 28 days. Next, animals from each group were euthanized. Frozen sections were prepared from collected jejunum and subjected to double immunofluorescence staining. NSAIDs supplementation caused a significant increase in the population of PACAP-, SP- and GAL-containing enteric neurons in the porcine jejunum. Our results suggest the participation of the selected neurotransmitters in regulatory processes of the gastrointestinal function and may indicate the direct toxic effect of NSAIDs on the ENS neurons.

Distribution of α-synuclein in normal human jejunum and its relations with the chemosensory and neuroendocrine system.

Alpha-synuclein (α-syn) is a presynaptic neuronal protein and its structural alterations play an important role in the pathogenesis of neurodegenerative diseases, such as Parkinson's disease (PD). It has been originally described in the brain and aggregated α-syn has also been found in the peripheral nerves including the enteric nervous system (ENS) of PD patients. ENS is a network of neurons and glia found in the gut wall which controls gastrointestinal function independently from the central nervous system. Moreover, two types of epithelial cells are crucial in the creation of an interface between the lumen and the ENS: they are the tuft cells and the enteroendocrine cells (EECs). In addition, the abundant enteric glial cells (EGCs) in the intestinal mucosa play a key role in controlling the intestinal epithelial barrier. Our aim was to localize and characterize the presence of α-syn in the normal human jejunal wall. Surgical specimens of proximal jejunum were collected from patients submitted to pancreaticoduodenectomy and intestinal sections underwent immunohistochemical procedure. Alpha-syn has been found both at the level of ENS and the epithelial cells. To characterize α-syn immunoreactive epithelial cells, we used markers such as choline acetyltransferase (ChAT), useful for the identification of tuft cells. Then we evaluated the co-presence of α-syn with serotonin (5-HT), expressed in EECs. Finally, we used the low-affinity nerve growth factor receptor (p75NTR), to detect peripheral EGCs. The presence of α-syn has been demonstrated in EECs, but not in the tuft cells. Additionally, p75NTR has been highlighted in EECs of the mucosal layer and co-localized with α-syn in EECs but not with ChAT-positive cells. These findings suggest that α-syn could play a possible role in synaptic transmission of the ENS and may contribute to maintain the integrity of the epithelial barrier of the small intestine through EECs.

Small intestine remodeling in male Goto-Kakizaki rats.

BACKGROUND: Obesity is associated with the development of insulin resistance (IR) and type-2 diabetes mellitus (T2DM); however, not all patients with T2DM are obese. The Goto-Kakizaki (GK) rat is an experimental model of spontaneous and non-obese T2DM. There is evidence that the intestine contributes to IR development in GK animals. This information prompted us to investigate small intestine remodeling in this animal model. METHODS: Four-month-old male Wistar (control) and GK rats were utilized for the present study. After removing the small intestine, the duodenum, proximal jejunum, and distal ileum were separated. We then measured villi and muscular and mucosa layer histomorphometry, goblet cells abundance, total myenteric and submucosal neuron populations, and inflammatory marker expression in the small intestinal segments and intestinal transit of both groups of animals. KEY RESULTS: We found that the GK rats exhibited decreased intestinal area (p < 0.0001), decreased crypt depth in the duodenum (p = 0.01) and ileum (p < 0.0001), increased crypt depth in the jejunum (p < 0.0001), longer villi in the jejunum and ileum (p < 0.0001), thicker villi in the duodenum (p < 0.01) and ileum (p < 0.0001), thicker muscular layers in the duodenum, jejunum, and ileum (p < 0.0001), increased IL-1ß concentrations in the duodenum and jejunum (p < 0.05), and increased concentrations of NF-κB p65 in the duodenum (p < 0.01), jejunum and ileum (p < 0.05). We observed high IL-1ß reactivity in the muscle layer, myenteric neurons, and glial cells of the experimental group. GK rats also exhibited a significant reduction in submucosal neuron density in the jejunum and ileum, ganglionic hypertrophy in all intestinal segments studied (p < 0.0001), and a slower intestinal transit (about 25%) compared to controls. CONCLUSIONS: The development of IR and T2DM in GK rats is associated with small intestine remodeling that includes marked alterations in small intestine morphology, local inflammation, and reduced intestinal transit.

Squalamine Restores the Function of the Enteric Nervous System in Mouse Models of Parkinson's Disease.

BACKGROUND: Parkinson's disease (PD) is a progressive neurodegenerative disorder thought to be caused by accumulation of α-synuclein (α-syn) within the brain, autonomic nerves, and the enteric nervous system (ENS). Involvement of the ENS in PD often precedes the onset of the classic motor signs of PD by many years at a time when severe constipation represents a major morbidity. Studies conducted in vitro and in vivo, have shown that squalamine, a zwitterionic amphipathic aminosterol, originally isolated from the liver of the dogfish shark, effectively displaces membrane-bound α-syn. OBJECTIVE: Here we explore the electrophysiological effect of squalamine on the gastrointestinal (GI) tract of mouse models of PD engineered to express the highly aggregating A53T human α-syn mutant. METHODS: GI motility and in vivo response to oral squalamine in PD model mice and controls were assessed using an in vitro tissue motility protocol and via fecal pellet output. Vagal afferent response to squalamine was measured using extracellular mesenteric nerve recordings from the jejunum. Whole cell patch clamp was performed to measure response to squalamine in the myenteric plexus. RESULTS: Squalamine effectively restores disordered colonic motility in vivo and within minutes of local application to the bowel. We show that topical squalamine exposure to intrinsic primary afferent neurons (IPANs) of the ENS rapidly restores excitability. CONCLUSION: These observations may help to explain how squalamine may promote gut propulsive activity through local effects on IPANs in the ENS, and further support its possible utility in the treatment of constipation in patients with PD.

Botulinum toxin as a treatment for short bowel syndrome in rats.

PURPOSE: The denervation of the intestine with benzalkonium chloride (BAC) reduces mortality and improves weight gain in rats with short bowel syndrome (SBS). Nevertheless, translating these promising findings from bench to bedside is not feasible because BAC promotes peritonitis and irreversible denervation which may be followed by an uncontrolled dilatation of the viscera. The use of botulinum toxin (BT) instead of BAC to achieve the denervation of the remaining small intestine in SBS could be an interesting option because it leads to a mild and transient denervation of the intestine. METHODS: Here we evaluated the effects of the ileal denervation with BT in rats with SBS by verifying the body weight variation and intestinal morphological parameters. Four groups with 6 animals each were submitted to enterectomy with an ileal injection of saline (group E) or BT (group EBT). Control groups were submitted to simulated surgery with an ileal injection of BT (group BT) or saline (group C - control). RESULTS: We observed that the treatment of the remaining ileum with BT completely reversed the weight loss associated to extensive small bowel resection. CONCLUSION: This may provide a new promising approach to the surgical treatment of SBS.

Ethyl Acetate Fraction from Trichilia catigua Confers Partial Neuroprotection in Components of the Enteric Innervation of the Jejunum in Diabetic Rats.

BACKGROUND/AIMS: Diabetes causes damage to the enteric nervous system. The enteric nervous system consists of neurons and enteric glial cells (EGCs). The present study evaluated the effects of an ethyl-acetate fraction (EAF) from Trichilia catigua (T. catigua; 200 mg/kg) on the total population of enteric neurons (HuC/D-immunoreactive [IR]) and EGCs (S100-IR and glial fibrillary acidic protein [GFAP]-IR) in the total preparation and jejunal mucosa in diabetic rats. METHODS: The animals were distributed into four groups: normoglycemic rats (N), diabetic rats (D), normoglycemic rats that received the EAF (NC), and diabetic rats that received the EAF (DC). The jejunum was processed for immunohistochemistry to evaluate HuC/D, S100, and GFAP immunoreactivity. The expression of S100 and GFAP proteins was also quantified by Western blot. RESULTS: The D group exhibited a decrease in the number of neurons and EGCs, an increase in the area of cell bodies, an increase in S100 protein expression, a decrease in GFAP protein expression, and a decrease in S100-IR and GFAP-IR EGCs in the jejunal mucosa. The DC group exhibited a decrease in the number of neurons and EGCs, a decrease in the area of cell bodies, a decrease in S100 and GFAP protein expression, and a decrease in S100-IR and GFAP-IR EGCs in the jejunal mucosa. The NC group exhibited maintenance of the number of neurons and EGCs, an increase in the area of cell bodies, and a decrease in S100 and GFAP protein expression. CONCLUSION: The EAF from T. catigua partially conferred protection against diabetic neuropathy in the enteric nervous system.

Interactions Between Commensal Bacteria and Enteric Neurons, via FPR1 Induction of ROS, Increase Gastrointestinal Motility in Mice.

BACKGROUND & AIMS: Reduced gastrointestinal (GI) motility is a feature of disorders associated with intestinal dysbiosis and loss of beneficial microbes. It is not clear how consumption of beneficial commensal microbes, marketed as probiotics, affects the enteric nervous system (ENS). We studied the effects of the widely used probiotic and the commensal Lactobacillus rhamnosus GG (LGG) on ENS and GI motility in mice. METHODS: Conventional and germ free C57B6 mice were gavaged with LGG and intestinal tissues were collected; changes in the enteric neuronal subtypes were assessed by real-time polymerase chain reaction, immunoblots, and immunostaining. Production of reactive oxygen species (ROS) in the jejunal myenteric plexi and phosphorylation (p) of mitogen-activated protein kinase 1 (MAPK1) in the enteric ganglia were assessed by immunoblots and immunostaining. Fluorescence in situ hybridization was performed on jejunal cryosections with probes to detect formyl peptide receptor 1 (FPR1). GI motility in conventional mice was assessed after daily gavage of LGG for 1 week. RESULTS: Feeding of LGG to mice stimulated myenteric production of ROS, increased levels of phosphorylated MAPK1, and increased expression of choline acetyl transferase by neurons (P < .001). These effects were not observed in mice given N-acetyl cysteine (a ROS inhibitor) or LGGΩSpaC (an adhesion-mutant strain of LGG) or FPR1-knockout mice. Gavage of mice with LGG for 1 week significantly increased stool frequency, reduced total GI transit time, and increased contractions of ileal circular muscle strips in ex vivo experiments (P < .05). CONCLUSIONS: Using mouse models, we found that LGG-mediated signaling in the ENS requires bacterial adhesion, redox mechanisms, and FPR1. This pathway might be activated to increase GI motility in patients.

Inducible nitric oxide synthase-derived nitric oxide reduces vagal satiety signalling in obese mice.

KEY POINTS: Obesity is associated with disrupted satiety regulation. Mice with diet-induced obesity have reduced vagal afferent neuronal excitability and a decreased afferent response to satiety signals. A low grade inflammation occurs in obesity with increased expression of inducible nitric oxide synthase (iNOS). Inhibition of iNOS in diet-induced obese mice restored vagal afferent neuronal excitability, increased the afferent response to satiety mediators and distention of the gut, and reduced short-term energy intake. A prolonged inhibition of iNOS reduced energy intake and body weight gain during the first week, and reduced amounts of epididymal fat after 3 weeks. We identified a novel pathway underlying disrupted satiety regulation in obesity. Blocking of this pathway might be clinically useful for the management of obesity. ABSTRACT: Vagal afferents regulate feeding by transmitting satiety signals to the brain. Mice with diet-induced obesity have reduced vagal afferent sensitivity to satiety signals. We investigated whether inducible nitric oxide synthase (iNOS)-derived NO contributed to this reduction. C57BL/6J mice were fed a high- or low-fat diet for 6-8 weeks. Nodose ganglia and jejunum were analysed by immunoblotting for iNOS expression; NO production was measured using a fluorometric assay. Nodose neuron excitability and intestinal afferent sensitivity were evaluated by whole-cell patch clamp and in vitro afferent recording, respectively. Expression of iNOS and production of NO were increased in nodose ganglia and the small intestine in obese mice. Inhibition of iNOS in obese mice by pre-treatment with an iNOS inhibitor increased nodose neuron excitability via 2-pore-domain K+ channel leak currents, restored afferent sensitivity to satiety signals and reduced short-term energy intake. Obese mice given the iNOS inhibitor daily for 3 weeks had reduced energy intake and decreased body weight gain during the first week, compared to mice given saline, and lower amounts of epididymal fat at the end of 3 weeks. Inhibition of iNOS or blocking the action of iNOS-derived NO on vagal afferent pathways might comprise therapeutic strategies for hyperphagia and obesity.

Botulinum toxin as a treatment for short bowel syndrome in rats

Abstract Purpose: The denervation of the intestine with benzalkonium chloride (BAC) reduces mortality and improves weight gain in rats with short bowel syndrome (SBS). Nevertheless, translating these promising findings from bench to bedside is not feasible because BAC promotes peritonitis and irreversible denervation which may be followed by an uncontrolled dilatation of the viscera. The use of botulinum toxin (BT) instead of BAC to achieve the denervation of the remaining small intestine in SBS could be an interesting option because it leads to a mild and transient denervation of the intestine. Methods: Here we evaluated the effects of the ileal denervation with BT in rats with SBS by verifying the body weight variation and intestinal morphological parameters. Four groups with 6 animals each were submitted to enterectomy with an ileal injection of saline (group E) or BT (group EBT). Control groups were submitted to simulated surgery with an ileal injection of BT (group BT) or saline (group C - control). Results: We observed that the treatment of the remaining ileum with BT completely reversed the weight loss associated to extensive small bowel resection. Conclusion: This may provide a new promising approach to the surgical treatment of SBS.

Effect of absorption-modifying excipients, hypotonicity, and enteric neural activity in an in vivo model for small intestinal transport.

The small intestine mucosal barrier is physiologically regulated by the luminal conditions, where intestinal factors, such as diet and luminal tonicity, can affect mucosal permeability. The intestinal barrier may also be affected by absorption-modifying excipients (AME) in oral drug delivery systems. Currently, there is a gap in the understanding of how AMEs interact with the physiological regulation of intestinal electrolyte transport and fluid flux, and epithelial permeability. Therefore, the objective of this single-pass perfusion study in rat was to investigate the effect of three AMEs on the intestinal mucosal permeability at different luminal tonicities (100, 170, and 290 mOsm). The effect was also evaluated following luminal administration of a nicotinic receptor antagonist, mecamylamine, and after intravenous administration of a COX-2 inhibitor, parecoxib, both of which affect the enteric neural activity involved in physiological regulation of intestinal functions. The effect was evaluated by changes in intestinal lumen-to-blood transport of six model compounds, and blood-to-lumen clearance of 51Cr-EDTA (a mucosal barrier marker). Luminal hypotonicity alone increased the intestinal epithelial transport of 51Cr-EDTA. This effect was potentiated by two AMEs (SDS and caprate) and by parecoxib, while it was reduced by mecamylamine. Consequently, the impact of enteric neural activity and luminal conditions may affect nonclinical determinations of intestinal permeability. In vivo predictions based on animal intestinal perfusion models can be improved by considering these effects. The in vivo relevance can be increased by treating rats with a COX-2 inhibitor prior to surgery. This decreases the risk of surgery-induced ileus, which may affect the physiological regulation of mucosal permeability.

Imanaga's First Method for Reconstruction with Preservation of Mesojejunal Autonomic Nerves During Pylorus-Preserving Pancreatoduodenectomy.

BACKGROUND Pancreatic surgeries have undergone substantial development. Pancreaticoduodenectomy and pylorus-preserving pancreatoduodenectomy inherently require reconstruction. In 1960, Professor Imanaga introduced a reconstructive technique performed in the order of the gastric remnant, pancreatic duct, and biliary tree from the viewpoint of physiologic function after pancreaticoduodenectomy. We herein report our experience with Imanaga's first method during pylorus-preserving pancreatoduodenectomy and retrospectively evaluate the short- and long-term outcomes. Technicalities and pitfalls are also discussed. CASE REPORT Eight patients were evaluated (mean follow-up period, 16.7 ± 1.0 years). Mesojejunal autonomic nerves were preserved without tension to the greatest extent possible for reconstruction. Intentional dissection of regional lymph nodes and nerves was performed in five and two patients, respectively. During the short-term postoperative period, one patient developed pancreatic leakage resulting in an intraperitoneal abscess, and endoscopic transgastric drainage was required. Two patients developed delayed gastric emptying. In three patients, passage from the duodenojejunostomy to pancreaticojejunostomy was mechanically disturbed, and endoscopic dilations with a balloon bougie were repeated. Repeated cholangitis was observed in three patients. During the long-term postoperative period, neither cachexia nor sarcopenia was observed, although two patients had diabetes. Two patients were free from all medications. Three patients who did not undergo intentional dissection of lymph nodes and nerves showed acceptable short- and long-term outcomes, although one each developed repeated cholangitis and adhesive ileus during the short-term period. CONCLUSIONS Imanaga's first reconstruction may have potential benefits, especially for diseases that do not require intentional dissection. Adequate mobilization of the pancreatic remnant is important for successful reconstruction.

Assemblages A and B of Giardia duodenalis reduce enteric glial cells in the small intestine in mice.

Infection of Giardia duodenalis is one of the most common human parasitic disease worldwide. This infection may be related to important changes in the enteric nervous system. The objective of this study was to evaluate the myenteric and submucosal plexuses, the intestinal muscle layer, and gastrointestinal transit in mice infected with assemblages A and B of G. duodenalis. Swiss albino mice (Mus musculus) were infected with assemblages A and B of G. duodenalis for 15 days. Gastrointestinal transit time was evaluated before euthanasia. Duodenum and jejunum were removed for histological and immunohistochemical analyses. It was observed a reduction in the enteric glial cell count and a decrease in the ratio of enteric glial cells to neurons. The number of neurons did not change, but morphological changes were observed in the duodenum and jejunum in both plexuses, including an increase in the nuclear area and a reduction of cell bodies in the myenteric plexus and a decrease in the nuclear area in the submucosal plexus. A reduction of the thickness of the muscle layer was observed in the duodenum, with no significant differences in the gastrointestinal transit times. Assemblages A and B of G. duodenalis decrease the number of enteric glial cells in the myenteric and submucosal plexuses, decrease the thickness of the muscle layer, and change the morphology of neurons. Graphical abstract ᅟ.

Small bowel capsule endoscopy revealing neuromuscular and vascular hamartoma of the jejunum: A case report.

RATIONALE: Digestive hemorrhage is a life-threatening and represents for both clinicians and patient a challenger problematic condition with the urgencies to discover the origin for correct the cause and safe the life of patient. PATIENT CONCERNS: We report the case of a 58 -year-old man with extremely rare hamartomatous neurovascular lesion. Following recurrent episode of intestinal hemorrhage the patient underwent small bowel capsule endoscopy. DIAGNOSES: Diagnosed with small intestine neoplasia. INTERVENTIONS: The patient underwent curative small bowel resection. Histologic diagnosis was neuromuscular and vascular hamartoma (NMVH). In the small intestine, neoplastic lesions are very rare (2%) and mostly malformative while the more frequent cause of cryptic digestive hemorrhage remains angiodysplasia (50%) . The preexisting NMVH was exacerbated by the use of non-steroidal anti-inflammatory drugs, causing hemorrhage due to diffuse ulceration. OUTCOMES: The patient stay healthy after treatment. LESSONS: This is an hemorrhagic lesion with macroscopic "neoplastic" patterns due to abnormal mixing of normal indigenous tissue components. It poses a diagnostic challenge for clinicians and pathologists, but diagnosis is facilitated by capsule endoscopy and surgical treatment should provide definitive resolution.

Unique expression pattern of KIBRA in the enteric nervous system of APP/PS1 mice.

KIBRA has been recognized as a memory-related gene, which is abundant in the brain and kidney of mammals. However, the expression pattern of KIBRA in the "second brain"-enteric nervous system (ENS) is still unknown, especially in neurodegenerative disorders. In this study, we aimed to investigate the detailed expression pattern of KIBRA in the intestinal myenteric nerve plexus of APP/PS1 and wild type mice by whole mount staining technology. The deposition of Aß and increased levels of phosphorylated Tau (p-Tau) and total Tau (T-Tau) protein were observed in the intestinal myenteric nerve plexus of APP/PS1 mice. Interestingly, the amount of Tuj+ cells remained unchanged between these two groups. Compared to the control mice, the protein levels of KIBRA significantly increased in the jejunal myenteric plexus of APP/PS1 mice, and the proportion of KIBRA+ GABAergic neurons in both the jejunal myenteric nerve plexus and the cortex was much higher in the APP/PS1 mice. But there was no significant difference in the number of KIBRA+ cholinergic neurons and KIBRA+ nitrergic neurons between APP/PS1 and wild type mice. In summary, our study further confirmed that typical pathology features of Alzheimer's disease (AD) not only existed in the central nervous system but also in the ENS.

Cholinergic signaling plasticity maintains viscerosensory responses during Aspiculuris tetraptera infection in mice small intestine.

Intestinal parasites alter gastrointestinal (GI) functions like the cholinergic function. Aspiculuris tetraptera is a pinworm frequently observed in laboratory facilities, which infests the mice cecum and proximal colon. However, little is known about the impact of this infection on the GI sensitivity. Here, we investigated possible changes in spontaneous mesenteric nerve activity and on the mechanosensitivity function of worm-free regions of naturally infected mice with A. tetraptera. Infection increased the basal firing of mesenteric afferent nerves in jejunum. Our findings indicate that nicotinic but not muscarinic receptors, similarly affect spontaneous nerve firing in control and infected animals; these axons are mainly vagal. No difference between groups was observed on spontaneous activity after nicotinic receptor inhibition. However, and contrary to the control group, during infection, the muscarinic signaling was shown to be elevated during mechanosensory experiments. In conclusion, we showed for the first time that alterations induced by infection of the basal afferent activity were independent of the cholinergic function but changes in mechanosensitivity were mediated by muscarinic, but not nicotinic, receptors and specifically by high threshold nerve fibers (activated above 20mmHg), known to play a role in nociception. These plastic changes within the muscarinic signaling would function as a compensatory mechanism to maintain a full mechanosensory response and the excitability of nociceptors during infection. These changes indicate that pinworm colonic infection can target other tissues away from the colon.

Quantitative morphometric analysis of the myenteric nervous plexus ganglion structures along the human digestive tract.

BACKGROUND/AIM: All the functions of the digestive system are controlled, guided and initiated by the autonomic nervous system. A special part of this system placed in the wall of the gastrointestinal tract is known as the enteric or metasympathetic nervous system. The aim of this study was to analyse myenteric nervous plexus in different parts of the digestive tract. METHODS: We examined the myenteric nervous plexus of the esophagus, stomach, duodenum, jejunum, ileum, transverse colon and rectum in tissue samples taken from 30 cadavers of persons aged 20-84 years. After standard histological processing sections were stained with hematoxylin-eosin, cresyl violet (CV) and AgNO3 method. Multipurpose test system M42 was used in morphometric analysis. The results were analyzed by t-test and analysis of variance. RESULTS: The number of neurons per cm² surface was the lowest in the esophagus (2.045 ± 310.30) and the largest in the duodenum (65,511 ± 5,639). The statistical processing showed significant differences (P < 0.001) in the number of neurons between the esophagus and all other parts of the digestive tract. The maximal value of the average surface of the myenteric nervous plexus neurons was observed in the esophagus (588.93 ± 30.45 µm²) and the lowest in the stomach (296.46 ± 22.53 µm²). CONCLUSION: There are differences in the number of ganglion cells among different parts of the human digestive tract. The differences range from a few to several tens of thousands of neuron/cm2. The myenteric nervous plexus of the esophagus was characterized by a significantly smaller number of neurons but their bodies and nuclei are significantly larger compared to other parts of the digestive tract.

Electroacupuncture at ST25 inhibits jejunal motility: Role of sympathetic pathways and TRPV1.

AIM: To investigate whether electroacupuncture (EA) at ST25 affects jejunal motility in vivo and if so, whether a sympathetic pathway is involved. METHODS: Jejunal motility was assessed using a manometric balloon placed in the jejunum approximately about 3-5 cm away from the suspensory ligament of the duodenum in anesthetized animals. The effects of EA at ST25 were measured in male Sprague-Dawley rats, some of which were treated with propranolol or clenbuterol (EA intensities: 1, 3, 5, 7, and 9 mA), and in male transient receptor potential vanilloid-1 (TRPV1) (capsaicin receptor) knockout mice (EA intensities: 1, 2, and 4 mA). RESULTS: Anesthetized rats exhibited three types of fasting jejunal motor patterns (types A, B, and C), and only type C rats responded to EA stimulation. In type C rats, EA at ST25 significantly suppressed the motor activity of the jejunum in an intensity-dependent manner. The inhibitory effect of EA was weakened by propranolol (ß adrenoceptor antagonist) and disappeared with clenbuterol (ß adrenoceptor agonist) induced inhibition of motility, suggesting that the effect of EA on motility is mediated via a sympathetic pathway. Compared with wild-type mice, EA at ST25 was less effective in TRPV1 knockout mice, suggesting that this multi-modal sensor channel participates in the mechanism. CONCLUSION: EA at ST25 was found to inhibit jejunal motility in an intensity-dependent manner, via a mechanism in which sympathetic nerves and TRPV1 receptors play an important role.

Ageing and gastrointestinal sensory function: altered colonic mechanosensory and chemosensory function in the aged mouse.

KEY POINTS: Remarkably little is known about how age affects the sensory signalling pathways in the gastrointestinal tract despite age-related gastrointestinal dysfunction being a prime cause of morbidity amongst the elderly population High-threshold gastrointestinal sensory nerves play a key role in signalling distressing information from the gut to the brain. We found that ageing is associated with attenuated high-threshold afferent mechanosensitivity in the murine colon, and associated loss of TRPV1 channel function. These units have the capacity to sensitise in response to injurious events, and their loss in ageing may predispose the elderly to lower awareness of GI injury or disease. ABSTRACT: Ageing has a profound effect upon gastrointestinal function through mechanisms that are poorly understood. Here we investigated the effect of age upon gastrointestinal sensory signalling pathways in order to address the mechanisms underlying these changes. In vitro mouse colonic and jejunal preparations with attached splanchnic and mesenteric nerves were used to study mechanosensory and chemosensory afferent function in 3-, 12- and 24-month-old C57BL/6 animals. Quantitative RT-PCR was used to investigate mRNA expression in colonic tissue and dorsal root ganglion (DRG) cells isolated from 3- and 24-month animals, and immunohistochemistry was used to quantify the number of 5-HT-expressing enterochromaffin (EC) cells. Colonic and jejunal afferent mechanosensory function was attenuated with age and these effects appeared earlier in the colon compared to the jejunum. Colonic age-related loss of mechanosensory function was more pronounced in high-threshold afferents compared to low-threshold afferents. Chemosensory function was attenuated in the 24-month colon, affecting TRPV1 and serotonergic signalling pathways. High-threshold mechanosensory afferent fibres and small-diameter DRG neurons possessed lower functional TRPV1 receptor responses, which occurred without a change in TRPV1 mRNA expression. Serotonergic signalling was attenuated at 24 months, but TPH1 and TPH2 mRNA expression was elevated in colonic tissue. In conclusion, we saw an age-associated decrease in afferent mechanosensitivity in the mouse colon affecting HT units. These units have the capacity to sensitise in response to injurious events, and their loss in ageing may predispose the elderly to lower awareness of GI injury or disease.

The TRPV1 channel in rodents is a major target for antinociceptive effect of the probiotic Lactobacillus reuteri DSM 17938.

Certain probiotic bacteria have been shown to reduce distension-dependent gut pain, but the mechanisms involved remain obscure. Live luminal Lactobacillus reuteri (DSM 17938) and its conditioned medium dose dependently reduced jejunal spinal nerve firing evoked by distension or capsaicin, and 80% of this response was blocked by a specific TRPV1 channel antagonist or in TRPV1 knockout mice. The specificity of DSM action on TRPV1 was further confirmed by its inhibition of capsaicin-induced intracellular calcium increases in dorsal root ganglion neurons. Another lactobacillus with ability to reduce gut pain did not modify this response. Prior feeding of rats with DSM inhibited the bradycardia induced by painful gastric distension. These results offer a system for the screening of new and improved candidate bacteria that may be useful as novel therapeutic adjuncts in gut pain. Certain bacteria exert visceral antinociceptive activity, but the mechanisms involved are not determined. Lactobacillus reuteri DSM 17938 was examined since it may be antinociceptive in children. Since transient receptor potential vanilloid 1 (TRPV1) channel activity may mediate nociceptive signals, we hypothesized that TRPV1 current is inhibited by DSM. We tested this by examining the effect of DSM on the firing frequency of spinal nerve fibres in murine jejunal mesenteric nerve bundles following serosal application of capsaicin. We also measured the effects of DSM on capsaicin-evoked increase in intracellular Ca(2+) or ionic current in dorsal root ganglion (DRG) neurons. Furthermore, we tested the in vivo antinociceptive effects of oral DSM on gastric distension in rats. Live DSM reduced the response of capsaicin- and distension-evoked firing of spinal nerve action potentials (238 ± 27.5% vs. 129 ± 17%). DSM also reduced the capsaicin-evoked TRPV1 ionic current in DRG neuronal primary culture from 83 ± 11% to 41 ± 8% of the initial response to capsaicin only. Another lactobacillus (Lactobacillus rhamnosus JB-1) with known visceral anti-nociceptive activity did not have these effects. DSM also inhibited capsaicin-evoked Ca(2+) increase in DRG neurons; an increase in Ca(2+) fluorescence intensity ratio of 2.36 ± 0.31 evoked by capsaicin was reduced to 1.25 ± 0.04. DSM releasable products (conditioned medium) mimicked DSM inhibition of capsaicin-evoked excitability. The TRPV1 antagonist 6-iodonordihydrocapsaicin or the use of TRPV1 knock-out mice revealed that TRPV1 channels mediate about 80% of the inhibitory effect of DSM on mesenteric nerve response to high intensity gut distension. Finally, feeding with DSM inhibited perception in rats of painful gastric distension. Our results identify a specific target channel for a probiotic with potential therapeutic properties.