Search

Background: Visceral hypersensitivity is considered the core pathophysiological mechanism that causes abdominal pain in patients with irritable bowel syndrome (IBS). Fungal dysbiosis has been proved to contribute to visceral hypersensitivity in IBS patients. However, the underlying mechanisms for Dectin-1, a major fungal recognition receptor, in visceral hypersensitivity are poorly understood. This study aimed to explore the role of Dectin-1 in visceral hypersensitivity and elucidate the impact of Dectin-1 activity on the function of transient receptor potential vanilloid type 1 (TRPV1)., Methods: Visceral hypersensitivity model was established by the intracolonic administration of 0.1 mL TNBS (130 μg/mL in 30% ethanol) in the male mice. Fluconazole and nystatin were used as fungicides. Laminarin, a Dectin-1 antagonist and gene knockout (Clec7a -/- ) mice were used to interrupt the function of Dectin-1. Colorectal distension-electromyogram recording was performed to assess visceral sensitivity. Immunostaining experiment was performed to determine the localization of Dectin-1 in dorsal root ganglion (DRG) neurons. Calcium imaging study was performed to assay TRPV1-mediated calcium influx in acutely dissociated DRG neurons., Results: Pretreatment with fungicides, administration of laminarin or genetic deletion of Clec7a alleviated TNBS-induced visceral hypersensitivity in male mice. The expression of Dectin-1 was upregulated in the DRG and colon of TNBS-treated mice. Colocalization of Dectin-1 and TRPV1 was observed in DRG neurons. Importantly, pretreatment with curdlan, a Dectin-1 agonist, increased TRPV1-mediated calcium influx., Conclusions: Dectin-1 contributes to visceral hypersensitivity in IBS or in inflammatory bowel disease in remission and activation of Dectin-1 induces TRPV1 sensitization., Significance Statement: This work provides direct evidence for the functional regulation of TRPV1 channel by Dectin-1 activity, proposing a new mechanism underlying TRPV1 sensitization. Control of intestinal fungi might be beneficial for the treatment of refractory abdominal pain in patients with IBS or IBD in remission., (© 2024 European Pain Federation ‐ EFIC ®.)

Visceral hypersensitivity, a hallmark of disorders of the gut-brain axis, is associated with exposure to early-life stress (ELS). Activation of neuronal β3-adrenoceptors (AR) has been shown to alter central and peripheral levels of tryptophan and reduce visceral hypersensitivity. In this study, we aimed to determine the potential of a β3-AR agonist in reducing ELS-induced visceral hypersensitivity and possible underlying mechanisms. Here, ELS was induced using the maternal separation (MS) model, where Sprague Dawley rat pups were separated from their mother in early life (postnatal day 2-12). Visceral hypersensitivity was confirmed in adult offspring using colorectal distension (CRD). CL-316243, a β3-AR agonist, was administered to determine anti-nociceptive effects against CRD. Distension-induced enteric neuronal activation as well as colonic secretomotor function were assessed. Tryptophan metabolism was determined both centrally and peripherally. For the first time, we showed that CL-316243 significantly ameliorated MS-induced visceral hypersensitivity. Furthermore, MS altered plasma tryptophan metabolism and colonic adrenergic tone, while CL-316243 reduced both central and peripheral levels of tryptophan and affected secretomotor activity in the presence of tetrodotoxin. This study supports the beneficial role of CL-316243 in reducing ELS-induced visceral hypersensitivity, and suggests that targeting the β3-AR can significantly influence gut-brain axis activity through modulation of enteric neuronal activation, tryptophan metabolism, and colonic secretomotor activity which may synergistically contribute to offsetting the effects of ELS., (© 2023 The Authors. Journal of Neurochemistry published by John Wiley & Sons Ltd on behalf of International Society for Neurochemistry.)

Mass spectrometry imaging (MSI) is a powerful scientific tool for understanding the spatial distribution of biochemical compounds in tissue structures. In this paper, we introduce three novel approaches in MSI data processing to perform the tasks of data augmentation, feature ranking, and image registration. We use these approaches in conjunction with non-negative matrix factorization (NMF) to resolve two of the biggest challenges in MSI data analysis, namely: 1) the large file sizes and associated computational resource requirements and 2) the complexity of interpreting the very high dimensional raw spectral data. There are many dimensionality reduction techniques that address the first challenge but do not necessarily result in readily interpretable features, leaving the second challenge unaddressed. We demonstrate that NMF is an effective dimensionality reduction algorithm that reduces the size of MSI datasets by three orders of magnitude with limited loss of information, yielding spatial and spectral components with meaningful correlation to tissue structure that may be used directly for subsequent data analysis without the need for additional clustering steps. This analysis is demonstrated on an MSI dataset from female Sprague-Dawley rats for an animal model of comorbid visceral pain hypersensitivity (CPH). We find that high-dimensional MSI data (∼ 100,000 ions per pixel) can be reduced to 20 spectral NMF components with < 20% loss in reconstruction accuracy. The resulting spatial NMF components are reproducible and correlate well with H&E-stained tissue images. These components may also be used to generate images with enhanced specificity for different tissue types. Small patches of NMF data (i.e., 20 spatial NMF components over 20 × 20 pixels) provide an accuracy of ∼ 87% in classifying CPH vs naïve control subjects. This paper presents the novel data processing methodologies that were used to produce these results, encompassing novel data processing pipelines for data augmentation to support training for classification, ranking of features according to their contribution to classification, and image registration to enhance tissue-specific imaging., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Pathirage et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Background: Transient receptor potential vanilloid 1 (TRPV1) cation channels, expressed on nociceptors, are well established as key contributors to abdominal pain in inflammatory bowel disease (IBD). Previous attempts at blocking these channels have been riddled with side effects. Here, we propose a novel treatment strategy, utilizing the large pore of TRPV1 channels as a drug delivery system to selectively inhibit visceral nociceptors., Methods: We induced colitis in rats using intrarectal dinitrobenzene sulfonic acid. Visceral hypersensitivity, spontaneous pain, and responsiveness of the hind paws to noxious heat stimuli were examined before and after the intrarectal application of membrane-impermeable sodium channel blocker (QX-314) alone or together with TRPV1 channel activators or blockers., Results: Intrarectal co-application of QX-314 with TRPV1 channel activator capsaicin significantly inhibited colitis-induced gut hypersensitivity. Furthermore, in the model of colitis, but not in naïve rats, QX-314 alone was sufficient to reverse gut hypersensitivity. The blockade of TRPV1 channels prevented this effect of QX-314. Finally, applying QX-314 alone to the inflamed gut inhibited colitis-induced ongoing pain., Conclusions: Selective silencing of gut nociceptors by a membrane-impermeable sodium channel blocker entering via exogenously or endogenously activated TRPV1 channels diminishes IBD-induced gut hypersensitivity. The lack of effect on naïve rats suggests a selective analgesic effect in the inflamed gut. Our results suggest that in the colitis model, TRPV1 channels are tonically active. Furthermore, our results emphasize the role of TRPV1-expressing nociceptive fibers in colitis-induced pain. These findings provide proof of concept for using charged activity blockers for the blockade of IBD-associated abdominal pain., (© The Author(s) 2024. Published by Oxford University Press on behalf of Crohn’s & Colitis Foundation. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Background: The aim of this study was to investigate the frequency dependence of electroacupuncture (EA) in alleviating chronic visceral pain in patients with irritable bowel syndrome (IBS) and the differences in the gut microbiota and metabolites as potential mechanisms to explain frequency dependence., Methods: A visceral hyperalgesia model was established by colorectal instillation of 2,4,6-trinitrobenzene sulfonic acid in rats, and EA treatment at 2/10 Hz, 2/50 Hz and 2/100 Hz was applied at ST25. Visceral sensation was quantified by the abdominal withdrawal reflex score and the area under the curve of the rectus abdominis electromyogram in response to colorectal distension. Ultrastructural morphological damage of colonic tissue of the rats was examined by transmission electron microscopy. 16S rRNA gene sequencing and 1H-nuclear magnetic resonance spectroscopy were applied to study the differences in the gut microbiota and to perform metabonomic profiling of the colonic tissue., Key Results: EA at ST25 at different frequencies attenuated chronic visceral pain, ultrastructural morphological damage to colonic tissue and disruption of the gut microbiota in IBS rats. The frequency of 2/100 Hz has more regulatory pathways than 2/10 Hz and 2/50 Hz. In addition, IBS rats exhibited colonic metabolic disorders, and pantothenate was significantly upregulated after EA treatment at different frequencies. Very low-density lipoprotein and 2-hydroxybutyrate were significantly increased in the 2/10 Hz group, while low density lipoprotein, very low-density lipoprotein, 2-hydroxybutyrate, methylmalonate and alpha-hydroxyisobutyric acid were significantly increased in the 2/100 Hz group., Conclusions and Inferences: EA at ST25 at different frequencies attenuated chronic visceral pain through different gut microbiota and metabolic pathways., (© 2024 John Wiley & Sons Ltd.)

Irritable bowel syndrome (IBS) is a common functional bowel disorder characterized by abdominal pain and visceral hypersensitivity. Reducing visceral hypersensitivity is the key to effectively relieving abdominal pain in IBS. Increasing evidence has confirmed that the thalamic nucleus reuniens (Re) and 5-hydroxytryptamine (5-HT) neurotransmitter system play an important role in the development of colorectal visceral pain, whereas the exact mechanisms remain largely unclear. In this study, we found that high expression of the 5-HT 2B receptors in the Re glutamatergic neurons promoted colorectal visceral pain. Specifically, we found that neonatal maternal deprivation (NMD) mice exhibited visceral hyperalgesia and enhanced spontaneous synaptic transmission in the Re brain region. Colorectal distension (CRD) stimulation induced a large amount of c-Fos expression in the Re brain region of NMD mice, predominantly in glutamatergic neurons. Furthermore, optogenetic manipulation of glutamatergic neuronal activity in the Re altered colorectal visceral pain responses in CON and NMD mice. In addition, we demonstrated that 5-HT 2B receptor expression on the Re glutamatergic neurons was upregulated and ultimately promoted colorectal visceral pain in NMD mice. These findings suggest a critical role of the 5HT 2B receptors on the Re glutamatergic neurons in the regulation of colorectal visceral pain., (© 2024. The Author(s).)

Irritable bowel syndrome (IBS) is a chronic gastrointestinal condition associated with altered bowel habits and recurrent abdominal pain, often triggered by food intake. Current treatments focus on improving stool pattern, but effective treatments for pain in IBS are still lacking due to our limited understanding of pathophysiological mechanisms. Visceral hypersensitivity (VHS), or abnormal visceral pain perception, underlies abdominal pain development in IBS, and mast cell activation has been shown to play an important role in the development of VHS. Our work recently revealed that abdominal pain in response to food intake is induced by the sensitization of colonic pain-sensing neurons by histamine produced by activated mast cells following a local IgE response to food. In this review, we summarize the current knowledge on abdominal pain and VHS pathophysiology in IBS, we outline the work leading to the discovery of the role of histamine in abdominal pain, and we introduce antihistamines as a novel treatment option to manage chronic abdominal pain in patients with IBS., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The aim of this study was to determine the anti-hypersensitivity activity of novel non-hallucinogenic compounds derived from iboga alkaloids (i.e., ibogalogs), including tabernanthalog (TBG), ibogainalog (IBG), and ibogaminalog (DM506), using mouse models of neuropathic (Chronic Constriction Injury; CCI) and visceral pain (dextrane sulfate sodium; DSS). Ibogalogs decreased mechanical hyperalgesia and allodynia induced by CCI in a dose- and timeframe-dependent manner, where IBG showed the longest anti-hyperalgesic activity at a comparatively lower dose, whereas DM506 displayed the quickest response. These compounds also decreased hypersensitivity induced by colitis, where DM506 showed the longest activity. To understand the mechanisms involved in these effects, two approaches were utilized: ibogalogs were challenged with the 5-HT 2A receptor antagonist ketanserin and the pharmacological activity of these compounds was assessed at the respective 5-HT 2A, 5-HT 6 , and 5-HT 7 receptor subtypes. The behavioral results clearly demonstrated that ketanserin abolishes the pain-relieving activity of ibogalogs without inducing any effect per se, supporting the concept that 5-HT 2A receptor activation, but not inhibition, is involved in this process. The functional results showed that ibogalogs potently activate the 5-HT 2A and 5-HT 6 receptor subtypes, whereas they behave as inverse agonists (except TBG) at the 5-HT 7 receptor. Considering previous studies showing that 5-HT 6 receptor inhibition, but not activation, and 5-HT 7 receptor activation, but not inhibition, relieved chronic pain, we can discard these two receptor subtypes as participating in the pain-relieving activity of ibogalogs. The potential involvement of 5-HT 2B/2 C receptor subtypes was also ruled out. In conclusion, the anti-hypersensitivity activity of ibogalogs in mice is mediated by a mechanism involving 5-HT 2A receptor activation., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Following colonic inflammation, the uninjured bladder afferent neurons are also activated. The mechanisms and pathways underlying this sensory neuron cross-activation (from injured neurons to uninjured neurons) are not fully understood. Colonic and bladder afferent neurons reside in the same spinal segments and are separated by satellite glial cells (SGCs) and extracellular matrix in dorsal root ganglia (DRG). SGCs communicate with sensory neurons in a bidirectional fashion. This review summarizes the differentially regulated genes/proteins in the injured and uninjured DRG neurons and explores the role of SGCs in regulation of sensory neuron crosstalk in visceral cross-organ sensitization. The review also highlights the paracrine pathways in mediating neuron-SGC and SGC-neuron coupling with an emphasis on the neurotrophins and purinergic systems. Finally, I discuss the results from recent RNAseq profiling of SGCs to reveal useful molecular markers for characterization, functional study, and therapeutic targets of SGCs. SIGNIFICANCE STATEMENT: Satellite glial cells (SGCs) are the largest glial subtypes in sensory ganglia and play a critical role in mediating sensory neuron crosstalk, an underlying mechanism in colon-bladder cross-sensitization. Identification of novel and unique molecular markers of SGCs can advance the discovery of therapeutic targets in treatment of chronic pain including visceral pain comorbidity., (Copyright © 2024 by The American Society for Pharmacology and Experimental Therapeutics.)

Slow transit constipation (STC) is one of the most common gastrointestinal disorders in children and adults worldwide. Paeoniflorin (PF), a monoterpene glycoside compound extracted from the dried root of Paeonia lactiflora, has been found to alleviate STC, but the mechanisms of its effect remain unclear. The present study aimed to investigate the effects and mechanisms of PF on intestinal fluid metabolism and visceral sensitization in rats with compound diphenoxylate-induced STC. Based on the evaluation of the laxative effect, the abdominal withdrawal reflex test, enzyme-linked immunosorbent assay, quantitative real-time polymerase chain reaction, western blot, and immunohistochemistry were used to detect the visceral sensitivity, fluid metabolism-related proteins, and acid-sensitive ion channel 3/extracellular signal-regulated kinase (ASIC3/ERK) pathway-related molecules. PF treatment not only attenuated compound diphenoxylate-induced constipation symptoms and colonic pathological damage in rats but also ameliorated colonic fluid metabolic disorders and visceral sensitization abnormalities, as manifested by increased colonic goblet cell counts and mucin2 protein expression, decreased aquaporin3 protein expression, improved abdominal withdrawal reflex scores, reduced visceral pain threshold, upregulated serum 5-hydroxytryptamine, and downregulated vasoactive intestinal peptide levels. Furthermore, PF activated the colonic ASIC3/ERK pathway in STC rats, and ASIC3 inhibition partially counteracted PF's modulatory effects on intestinal fluid and visceral sensation. In conclusion, PF alleviated impaired intestinal fluid metabolism and abnormal visceral sensitization in STC rats and thus relieved their symptoms through activation of the ASIC3/ERK pathway., (© 2024 The Authors. The Kaohsiung Journal of Medical Sciences published by John Wiley & Sons Australia, Ltd on behalf of Kaohsiung Medical University.)

Visceral hypersensitivity resulting from compromised gut barrier with activated immune system is a key feature of irritable bowel syndrome (IBS). Corticotropin-releasing factor (CRF) and Toll-like receptor 4 (TLR4) activate proinflammatory cytokine signaling to induce these changes, which is one of the mechanisms of IBS. As activation of the NLRP3 inflammasome by lipopolysaccharide (LPS) or TLR4 leads to release interleukin (IL)-1β, the NLRP3 inflammasome may be involved in the pathophysiology of IBS. Tranilast, an anti-allergic drug has been demonstrated to inhibit the NLRP3 inflammasome, and we evaluated the impact of tranilast on visceral hypersensitivity and colonic hyperpermeability induced by LPS or CRF (IBS rat model). Visceral pain threshold caused by colonic balloon distention was measured by monitoring abdominal muscle contractions electrophysiologically. Colonic permeability was determined by quantifying the absorbed Evans blue within the colonic tissue. Colonic protein levels of NLRP3 and IL-1β were assessed by immunoblot or ELISA. Intragastric administration of tranilast (20-200 mg/kg) for 3 days inhibited LPS (1 mg/kg)-induced visceral hypersensitivity and colonic hyperpermeability in a dose-dependent manner. Simultaneously, tranilast also abolished these alterations induced by CRF (50 µg/kg). LPS increased colonic protein levels of NLRP3 and IL-1β, and tranilast inhibited these changes. β-hydroxy butyrate, an NLRP3 inhibitor, also abolished visceral hypersensitivity and colonic hyperpermeability caused by LPS. In contrast, IL-1β induced similar GI alterations to LPS, which were not modified by tranilast. In conclusion, tranilast improved visceral pain and colonic barrier by suppression of the NLRP3 inflammasome in IBS rat models. Tranilast may be useful for IBS treating., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Background: Irritable bowel syndrome (IBS) is one of the most frequent and debilitating conditions leading to gastroenterological referrals. However, recommended treatments remain limited, yielding only limited therapeutic gains. Chitin-glucan (CG) is a novel dietary prebiotic classically used in humans at a dosage of 1.5-3.0 g/d and is considered a safe food ingredient by the European Food Safety Authority. To provide an alternative approach to managing patients with IBS, we performed preclinical molecular, cellular, and animal studies to evaluate the role of chitin-glucan in the main pathophysiological mechanisms involved in IBS., Aim: To evaluate the roles of CG in visceral analgesia, intestinal inflammation, barrier function, and to develop computational molecular models., Methods: Visceral pain was recorded through colorectal distension (CRD) in a model of long-lasting colon hypersensitivity induced by an intra-rectal administration of TNBS [15 milligrams (mg)/kilogram (kg)] in 33 Sprague-Dawley rats. Intracolonic pressure was regularly assessed during the 9 wk-experiment (weeks 0, 3, 5, and 7) in animals receiving CG ( n = 14) at a human equivalent dose (HED) of 1.5 g/d or 3.0 g/d and compared to negative control (tap water, n = 11) and positive control (phloroglucinol at 1.5 g/d HED, n = 8) groups. The anti-inflammatory effect of CG was evaluated using clinical and histological scores in 30 C57bl6 male mice with colitis induced by dextran sodium sulfate (DSS) administered in their drinking water during 14 d. HT-29 cells under basal conditions and after stimulation with lipopolysaccharide (LPS) were treated with CG to evaluate changes in pathways related to analgesia (µ-opioid receptor (MOR), cannabinoid receptor 2 (CB2), peroxisome proliferator-activated receptor alpha, inflammation [interleukin (IL)-10, IL-1b, and IL-8] and barrier function [mucin 2-5AC, claudin-2, zonula occludens (ZO)-1, ZO-2] using the real-time PCR method. Molecular modelling of CG, LPS, lipoteichoic acid (LTA), and phospholipomannan (PLM) was developed, and the ability of CG to chelate microbial pathogenic lipids was evaluated by docking and molecular dynamics simulations. Data were expressed as the mean ± SEM., Results: Daily CG orally-administered to rats or mice was well tolerated without including diarrhea, visceral hypersensitivity, or inflammation, as evaluated at histological and molecular levels. In a model of CRD, CG at a dosage of 3 g/d HED significantly decreased visceral pain perception by 14% after 2 wk of administration ( P < 0.01) and reduced inflammation intensity by 50%, resulting in complete regeneration of the colonic mucosa in mice with DSS-induced colitis. To better reproduce the characteristics of visceral pain in patients with IBS, we then measured the therapeutic impact of CG in rats with TNBS-induced inflammation to long-lasting visceral hypersensitivity. CG at a dosage of 1.5 g/d HED decreased visceral pain perception by 20% five weeks after colitis induction ( P < 0.01). When the CG dosage was increased to 3.0 g/d HED, this analgesic effect surpassed that of the spasmolytic agent phloroglucinol, manifesting more rapidly within 3 wk and leading to a 50% inhibition of pain perception ( P < 0.0001). The underlying molecular mechanisms contributing to these analgesic and anti-inflammatory effects of CG involved, at least in part, a significant induction of MOR, CB2 receptor, and IL-10, as well as a significant decrease in pro-inflammatory cytokines IL-1b and IL-8. CG also significantly upregulated barrier-related genes including muc5AC, claudin-2, and ZO-2. Molecular modelling of CG revealed a new property of the molecule as a chelator of microbial pathogenic lipids, sequestering gram-negative LPS and gram-positive LTA bacterial toxins, as well as PLM in fungi at the lowesr energy conformations., Conclusion: CG decreased visceral perception and intestinal inflammation through master gene regulation and direct binding of microbial products, suggesting that CG may constitute a new therapeutic strategy for patients with IBS or IBS-like symptoms., Competing Interests: Conflict-of-interest statement: Desreumaux reports personal fees from Abbvie, personal fees from Abbott, personal fees from Amgen, personal fees from Biocodex, personal fees from Biofortis, personal fees from Biogen, personal fees from Biokuris, personal fees from Dr Falk, personal fees from Ferring, personal fees from Galapagos, personal fees from Fresenius, personal fees from Janssen, personal fees from Intestinal Biotech Development, personal fees from Kitozyme, personal fees from Lesaffre, personal fees from MSD, personal fees from Norgine, personal fees from Pfizer, personal fees from Sandoz, personal fees from Shire, personal fees from Takeda, personal fees from Tillotts, and personal fees from UCB outside the submitted work; Dr. Desreumaux has issued a patent (WO2009103884) issued; Christel Rousseaux is Chief Executive Officer at Intestinal Biotech Development; Veronique Maquet is a Product Development Manager at Kitozyme; Salvatore Modica is Chief Operating Officer at Biokuris, a spin-off company of Kitozyme; The other authors have nothing to disclose., (©The Author(s) 2024. Published by Baishideng Publishing Group Inc. All rights reserved.)

Aims: Irritable bowel syndrome (IBS) is a common functional gastrointestinal disorder, but its pathogenesis remains incompletely understood, particularly the involvements of central nervous system sensitization in colorectal visceral pain. Our study was to investigate whether the paraventricular thalamus (PVT) projected to the insular cortex (IC) to regulate colorectal visceral pain in neonatal colonic inflammation (NCI) mice and underlying mechanisms., Methods: We applied optogenetic, chemogenetic, or pharmacological approaches to manipulate the glutamatergic PVT-IC pathway. Fiber photometry was used to assess neuronal activity. Electromyography activities in response to colorectal distension (CRD) were measured to evaluate the colorectal visceral pain., Results: NCI enhanced c-Fos expression and calcium activity upon CRD in the IC Glu , and optogenetic manipulation of them altered colorectal visceral pain responses accordingly. Viral tracing indicated that the PVT Glu projected to the IC Glu . Optogenetic manipulation of PVT Glu changed colorectal visceral pain responses. Furthermore, selective optogenetic modulation of PVT projections in the IC influenced colorectal visceral pain, which was reversed by chemogenetic manipulation of downstream IC Glu ., Conclusions: This study identified a novel PVT-IC neural circuit playing a critical role in colorectal visceral pain in a mouse model of IBS., (© 2023 The Authors. CNS Neuroscience & Therapeutics published by John Wiley & Sons Ltd.)

Aims: Basolateral amygdala (BLA), as a center for stress responses and emotional regulation, is involved in visceral hypersensitivity of irritable bowel syndrome (IBS) induced by stress. In the present study, we aimed to investigate the role of EphB2 receptor (EphB2) in BLA and explore the underlying mechanisms in this process., Methods: Visceral hypersensitivity was induced by water avoidance stress (WAS). Elevated plus maze test, forced swimming test, and sucrose preference test were applied to assess anxiety- and depression-like behaviors. Ibotenic acid or lentivirus was used to inactivate BLA in either the induction or maintenance stage of visceral hypersensitivity. The expression of protein was determined by quantitative PCR, immunofluorescence, and western blot., Results: EphB2 expression was increased in BLA in WAS rats. Inactivation of BLA or downregulation of EphB2 in BLA failed to induce visceral hypersensitivity as well as anxiety-like behaviors. However, during the maintenance stage of visceral pain, visceral hypersensitivity was only partially relieved but anxiety-like behaviors were abolished by inactivation of BLA or downregulation of EphB2 in BLA. Chronic WAS increased the expression of EphB2, N-methyl-D-aspartate receptors (NMDARs), and postsynaptic density protein (PSD95) in BLA. Downregulation of EphB2 in BLA reduced NMDARs and PSD95 expression in WAS rats. However, activation of NMDARs after the knockdown of EphB2 expression still triggered visceral hypersensitivity and anxiety-like behaviors., Conclusions: Taken together, the results suggest that EphB2 in BLA plays an essential role in inducing visceral hypersensitivity. In the maintenance stage, the involvement of EphB2 is crucial but not sufficient. The increase in EphB2 induced by WAS may enhance synaptic plasticity in BLA through upregulating NMDARs, which results in IBS-like symptoms. These findings may give insight into the treatment of IBS and related psychological distress., (© 2024 The Authors. CNS Neuroscience & Therapeutics published by John Wiley & Sons Ltd.)

Sex differences in visceral nociception have been reported in clinical and preclinical studies, but the potential differences in sensory neural encoding of the colorectum between males and females are not well understood. In this study, we systematically assessed sex differences in colorectal neural encoding by conducting high-throughput optical recordings in intact dorsal root ganglia (DRGs) from control and visceral hypersensitive mice. We found an apparent sex difference in zymosan-induced behavioral visceral hypersensitivity: enhanced visceromotor responses to colorectal distension were observed only in male mice, not in female mice. In addition, a higher number of mechanosensitive colorectal afferents were identified per mouse in the zymosan-treated male group than in the saline-treated male group, whereas the mechanosensitive afferents identified per mouse were comparable between the zymosan- and saline-treated female groups. The increased number of identified afferents in zymosan-treated male mice was predominantly from thoracolumbar (TL) innervation, which agrees with the significant increase in the TL afferent proportion in the zymosan group as compared with the control group in male mice. In contrast, female mice showed no difference in the proportion of colorectal neurons between saline- and zymosan-treated groups. Our results revealed a significant sex difference in colorectal afferent innervation and sensitization in the context of behavioral visceral hypersensitivity, which could drive differential clinical symptoms in male and female patients. NEW & NOTEWORTHY We used high-throughput GCaMP6f recordings to study 2,275 mechanosensitive colorectal afferents in mice. Our results revealed significant sex differences in the zymosan-induced behavioral visceral hypersensitivity, which were present in male but not female mice. Male mice also showed sensitization of colorectal afferents in the thoracolumbar pathway, whereas female mice did not. These findings highlight sex differences in sensory neural anatomy and function of the colorectum, with implications for sex-specific therapies for treating visceral pain.

Background: Despite achieving endoscopic remission, over 20% of inflammatory bowel disease (IBD) patients experience chronic abdominal pain. Visceral pain and the microbiome exhibit sex-dependent interactions, while visceral pain in IBD shows a sex bias. Our aim was to evaluate whether post-inflammatory microbial perturbations contribute to visceral hypersensitivity in a sex-dependent manner., Methods: Males, cycling females, ovariectomized, and sham-operated females were given dextran sodium sulfate to induce colitis and allowed to recover. Germ-free recipients received sex-appropriate and cross-sex fecal microbial transplants (FMT) from post-inflammatory donor mice. Visceral sensitivity was assessed by recording visceromotor responses to colorectal distention. The composition of the microbiota was evaluated via 16S rRNA gene V4 amplicon sequencing, while the metabolome was assessed using targeted (short chain fatty acids - SCFA) and semi-targeted mass spectrometry., Results: Post-inflammatory cycling females developed visceral hyperalgesia when compared to males. This effect was reversed by ovariectomy. Both post-inflammatory males and females exhibited increased SCFA-producing species, but only males had elevated fecal SCFA content. FMT from post-inflammatory females transferred visceral hyperalgesia to both males and females, while FMT from post-inflammatory males could only transfer visceral hyperalgesia to males., Conclusions: Female sex, hormonal status as well as the gut microbiota play a role in pain modulation. Our data highlight the importance of considering biological sex in the evaluation of visceral pain.

Early-life stress (ELS) is thought to cause the development of visceral pain disorders. While some individuals are vulnerable to visceral pain, others are resilient, but the intrinsic circuit and molecular mechanisms involved remain largely unclear. Herein, we demonstrate that inbred mice subjected to maternal separation (MS) could be separated into susceptible and resilient subpopulations by visceral hypersensitivity evaluation. Through a combination of chemogenetics, optogenetics, fiber photometry, molecular and electrophysiological approaches, we discovered that susceptible mice presented activation of glutamatergic projections or inhibition of GABAergic projections from the anteroventral bed nucleus of the stria terminalis (avBNST) to paraventricular nucleus (PVN) corticotropin-releasing hormone (CRH) neurons. However, resilience develops as a behavioral adaptation partially due to restoration of PVN SK2 channel expression and function. Our findings suggest that PVN CRH neurons are dually regulated by functionally opposing avBNST neurons and that this circuit may be the basis for neurobiological vulnerability to visceral pain., (© 2023. The Author(s), under exclusive licence to American College of Neuropsychopharmacology.)

Chronic visceral pain is a common symptom of irritable bowel syndrome (IBS). Exosomes are involved in the development of pain. Rab27a can mediate the release of exosomes. The purpose of this study is to investigate how Rab27a-mediated exosome secretion in the anterior cingulate cortex (ACC) regulates visceral hyperalgesia induced with neonatal maternal deprivation (NMD) in adult mice. The colorectal distension method was adopted to measure visceral pain. The BCA protein assay kit was applied to detect the exosome protein concentration. Western blotting, quantitative PCR, and immunofluorescence technique were adopted to detect the expression of Rab27a and the markers of exosomes. Exosomes extracted from ACC were more in NMD mice than in control (CON) mice. Injection of the exosome-specific inhibitor GW4869 in ACC attenuated colorectal visceral pain of NMD mice. Injection of NMD-derived exosomes produced colorectal visceral pain in CON mice. Rab27a was upregulated in ACC of NMD mice. Rab27a was highly expressed in ACC neurons of NMD mice, rather than astrocytes and microglia. Injection of Rab27a-siRNA reduced the release of exosomes and attenuated the colorectal visceral pain in NMD mice. This study suggested that overexpression of Rab27a increased exosome secretion in ACC neurons, thus contributing to visceral hyperalgesia in NMD mice. NEW & NOTEWORTHY This work demonstrated that the expression of Rab27a in the anterior cingulate cortex was upregulated, which mediated multivesicular bodies trafficking to the plasma membrane and led to the increased release of neuronal exosomes, thus contributing to colorectal visceral pain in neonatal maternal deprivation (NMD) mice. Blocking the release of exosomes or downregulation of Rab27a could alleviate colorectal visceral pain in NMD mice. These data may provide a promising strategy for the treatment of visceral pain in irritable bowel syndrome patients.

Background: Electroacupuncture (EA) can effectively relieve visceral hypersensitivity (VH). However, its mechanisms are still unclear., Objective: To investigate the impact of EA on VH caused by ileitis, and whether EA relieves VH by modulating the endogenous cannabinoid system (ECS)., Methods: Thirty male native goats were randomly divided into a saline-treated control group (Saline, n = 9) and three 2,4,6-trinitro-benzenesulfonic acid (TNBS)-treated VH model groups that underwent injection of TNBS into the ileal wall to induce VH and remained untreated (TNBS, n = 9) or received six sessions of EA (for 30 min every 3 days) (TNBS + EA, n = 6) or sham acupuncture (TNBS + Sham, n = 6). The visceromotor response (VMR) to colorectal distention (CRD) was measured after each EA treatment. Three goats in the Saline/TNBS groups were euthanized after 7 days for histopathological examination; the remaining 24 (n = 6/group) underwent sampling of the ileal wall, T11 spinal cord and brain nuclei/areas related to visceral regulation and ascending pain modulation system on day 22. Expression of cannabinoid receptor 1 (CB1R), fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) was detected by immunohistochemistry., Results: VMR to CRD was greater in TNBS-treated goats than in saline-treated goats (p < 0.01) from day 7 to 22. After day 7, EA-treated goats showed a decreased (p < 0.05) VMR compared with untreated TNBS-exposed goats. TNBS treatment decreased CB1R and increased FAAH and MAGL expression in the ileum and related nuclei/areas; this was reversed by EA., Conclusion: EA ameliorates VH, probably by regulating the ECS in the intestine and nuclei/areas related to visceral regulation and descending pain modulation systems.

Recent studies have demonstrated the vital role of P2X4 receptors (a family of ATP-gated non-selective cation channels) in the transmission of neuropathic and inflammatory pain. In this study, we investigated the role of spinal P2X4 receptors in chronic functional visceral hypersensitivity of neonatal maternal separation (NMS) rats. A rat model of irritable bowel syndrome was established by neonatal maternal separation. Visceral sensitivity was assessed by recording the response of the external oblique abdominal muscle to colorectal distension. P2X4 receptor antagonist and agonist were administrated intrathecally. The expression of P2X4 receptor was examined by Western Blot and immunofluorescence. The effect of P2X4 receptor antagonist on expression of brain-derived neurotrophic factor (BDNF) was assessed by Western Blot. We found neonatal maternal separation enhanced visceral hypersensitivity and increased the expression of P2X4 receptor in spinal thoracolumbar and lumbosacral segments of rats. Pharmacological results showed that visceral sensitivity was attenuated after intrathecal injection of P2X4 receptor antagonist, 5-BDBD, at doses of 10 nM or 100 nM, while visceral sensitivity was enhanced after intrathecal injection of P2X4 receptor agonist C5-TDS at doses of 10 μM or 15 μM. In addition, the spinal expression of BDNF significantly increased in NMS rats and intrathecal injection of 5-BDBD significantly decreased the expression of BDNF especially in NMS rats. C5-TDS failed to increase EMG amplitude in the presence of ANA-12 in control rats. Our results suggested the spinal P2X4 receptors played an important role in visceral hypersensitivity of NMS rats through BDNF., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

This study explored the role of P2X7 receptors in spinal cord astrocytes in the electroacupuncture-induced inhibition of visceral hypersensitivity (VH) in rats with irritable bowel syndrome (IBS). Visceral hypersensitivity of IBS was intracolonically induced by 2,4,6-trinitrobenzene sulfonic acid (TNBS). Visceromotor responses to colorectal distension (CRD-20,40,60,80 mmHg) and abdominal withdrawal reflex scoring (AWRs) were recorded after electroacupuncture at bilateral Zusanli (ST36) and Sanyinjiao (SP6) acupoints to evaluate the analgesic effect of electroacupuncture on visceral pain in rats with IBS. Fluorocitric acid (FCA), an astrocyte activity inhibitor, was injected intrathecally before electroacupuncture intervention and AWRs were recorded. Western blot and real-time qPCR were used to detect the expression of NMDA and P2X7 receptor to observe the regulation effect of electroacupuncture on NMDA receptor in the spinal cord of rats with visceral hypersensitivity. Intrathecal injection of P2X7 agonist or antagonist was administered before electroacupuncture treatment. To observe the effect of P2X7 receptor in spinal astrocytes on the inhibition of visceral hyperalgesia by electroacupuncture, the changes of AWR score, NMDA receptor in the spinal cord, and GFAP expression in astrocytes were detected. Inflammation of the colon had basically subsided at day 21 post-TNBS; persistent visceral hypersensitivity could be suppressed by electroacupuncture. This analgesic effect could be inhibited by FCA. The analgesic effect, downregulation of NMDA receptor NR1 subunit, and P2X7 protein of electroacupuncture were all reversed by FCA. P2X7 receptor antagonist A740003 can cooperate with EA to carry out analgesic effect in rats with visceral pain and downregulate the expression of NR1, NR2B, and GFAP in spinal dorsal horn. However, the P2X7 receptor agonist BzATP could partially reverse the analgesic effect of EA, inhibiting the downregulatory effect of EA on the expression of NR1, NR2B, and GFAP. These results indicate that EA may downregulate the expression of the NMDA receptor by inhibiting the P2X7 receptor in the spinal cord, thereby inhibiting spinal cord sensitization in IBS rats with visceral pain, in which astrocytes are an important medium., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Inflammatory and functional gastrointestinal disorders such as irritable bowel syndrome (IBS) and obstructive bowel disorder (OBD) underlie the most prevalent forms of visceral pain. Although visceral pain can be generally provoked by mechanical distension/stretch, the mechanisms that underlie visceral mechanosensitivity in colon-innervating visceral afferents remain elusive. Here, we show that virally mediated ablation of colon-innervating TRPV1-expressing nociceptors markedly reduces colorectal distention (CRD)-evoked visceromotor response (VMR) in mice. Selective ablation of the stretch-activated Piezo2 channels from TRPV1 lineage neurons substantially reduces mechanically evoked visceral afferent action potential firing and CRD-induced VMR under physiological conditions, as well as in mouse models of zymosan-induced IBS and partial colon obstruction (PCO). Collectively, our results demonstrate that mechanosensitive Piezo2 channels expressed by TRPV1-lineage nociceptors powerfully contribute to visceral mechanosensitivity and nociception under physiological conditions and visceral hypersensitivity under pathological conditions in mice, uncovering potential therapeutic targets for the treatment of visceral pain., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Visceral pain (VP) is a global problem with complex etiologies and limited therapeutic options. Guanylyl cyclase C (GUCY2C), an intestinal receptor producing cyclic GMP(cGMP), which regulates luminal fluid secretion, has emerged as a therapeutic target for VP. Indeed, FDA-approved GUCY2C agonists ameliorate VP in patients with chronic constipation syndromes, although analgesic mechanisms remain obscure. Here, we revealed that intestinal GUCY2C was selectively enriched in neuropod cells, a type of enteroendocrine cell that synapses with submucosal neurons in mice and humans. GUCY2Chi neuropod cells associated with cocultured dorsal root ganglia neurons and induced hyperexcitability, reducing the rheobase and increasing the resulting number of evoked action potentials. Conversely, the GUCY2C agonist linaclotide eliminated neuronal hyperexcitability produced by GUCY2C-sufficient - but not GUCY2C-deficient - neuropod cells, an effect independent of bulk epithelial cells or extracellular cGMP. Genetic elimination of intestinal GUCY2C amplified nociceptive signaling in VP that was comparable with chemically induced VP but refractory to linaclotide. Importantly, eliminating GUCY2C selectively in neuropod cells also increased nociceptive signaling and VP that was refractory to linaclotide. In the context of loss of GUCY2C hormones in patients with VP, these observations suggest a specific role for neuropod GUCY2C signaling in the pathophysiology and treatment of these pain syndromes.

Visceral pain caused by inflammatory bowel disease (IBD) greatly diminishes the quality of life in affected patients. Yet, the mechanism of how IBD causes visceral pain is currently not fully understood. Previous studies have suggested that the central nervous system (CNS) and gut-brain axis (GBA) play an important role in IBD-inducing visceral pain. As one of the treatments for IBD, electroacupuncture (EA) has been used to treat various types of pain and gastrointestinal diseases in clinical practice. However, whether EA relieves the visceral pain of IBD through the gut-brain axis has not been confirmed. To verify the relationship between visceral pain and CNS, the following experiments were conducted. 1 H-NMR analysis was performed on the prefrontal cortex (PFC) tissue obtained from IBD rat models to determine the link between the metabolites and their role in EA treatment against visceral pain. Western blot assay was employed for detecting the contents of glutamate transporter excitatory amino acid transporters 2 (EAAT2) and the glutamate receptor N-methyl-D-aspartate (NMDA) to verify whether EA treatment can alleviate neurotoxic symptoms induced by abnormal increases of glutamate. Study results showed that the glutamate content was significantly increased in the PFC of TNBS-induced IBD rats. This change was reversed after EA treatment. This process was associated with increased EAAT2 expression and decreased expression of NMDA receptors in the PFC. In addition, an increase in intestinal glutamic-metabolizing bacteria was observed. In conclusion, this study suggests that EA treatment can relieve visceral pain by reducing glutamine toxicity in the PFC, and serves an alternative clinical utility., Competing Interests: The authors declare no conflict of interest., (Copyright © 2023 Hao Jiang et al.)

Background: Irritable bowel syndrome (IBS) is a common functional gastrointestinal disorder, and its specific pathogenesis is still unclear. We have previously reported that TTX-resistant (TTX-R) sodium channels in colon-specific dorsal root ganglion (DRG) neurons were sensitized in a rat model of visceral hypersensitivity induced by neonatal colonic inflammation (NCI). However, the detailed molecular mechanism for activation of sodium channels remains unknown. This study was designed to examine roles for melatonin (MT) in sensitization of sodium channels in NCI rats., Methods: Colorectal distention (CRD) in adult male rats as a measure of visceral hypersensitivity. Colon-specific dorsal root ganglion (DRG) neurons were labeled with DiI and acutely dissociated for measuring excitability and sodium channel current under whole-cell patch clamp configurations. Western blot and Immunofluorescence were employed to detect changes in expression of Na v 1.8 and MT2., Results: The results showed that rats exhibited visceral hypersensitivity after NCI treatment. Intrathecal application of melatonin significantly increased the threshold of CRD in NCI rats with a dose-dependent manner, but has no role in the control group. Whole-cell patch clamp recording showed that melatonin remarkably decreased the excitability and the density of TTX-R sodium channel in DRG neurons from NCI rats. The expression of MT2 receptor at the protein level was markedly lower in NCI rats. 8MP, an agonist of MT2 receptor, enhanced the distention threshold in NCI rats. Application of 8MP reversed the enhanced hypersensitivity of DRG neurons from NCI rats. 8MP also reduced TTX-R sodium current density and modulated dynamics of TTX-R sodium current activation., Conclusions: These data suggest that sensitization of sodium channels of colon DRG neurons in NCI rats is most likely mediated by MT2 receptor, thus identifying a potential target for treatment for chronic visceral pain in patients with IBS.

Pain involves both sensory and affective elements. An aspect of the affective dimension of pain is its sustained unpleasantness, characterized by emotional feelings. Pain results from interactions between memory, attentional, and affective brain circuitry, and it has attracted enormous interest in pain research. However, the brain targets and signaling mechanism involved in pain remain elusive. Using a conditioned place avoidance (CPA) paradigm, we show that colorectal distention (CRD magnitude ≤ 35 mmHg, a subthreshold for pain) paired with a distinct environment can cause significant aversion to a location associated with pain-related insults in rats. We show a substantial increase in the L-lactate concentration in the anterior cingulate cortex (ACC) following CPA training. Local exogenous infusion of lactate into the ACC enhances aversive memory and induces the expression of the memory-related plasticity genes pCREB, CREB, and Erk1/2. The pharmacological experiments revealed that the glycogen phosphorylation inhibitor 1,4-dideoxy-1,4-imino-D-arabinitol (DAB) impairs memory consolidation. Furthermore, short-term Gi pathway activation of ACC astrocytes before CPA training significantly decreases the lactate level and suppresses pain-related aversive learning. The effects were reversed by the local infusion of lactate into the ACC. Our study demonstrates that lactate is released from astrocytes in vivo following visceral pain-related aversive learning and memory retrieval and induces the expression of the plasticity-related immediate early genes CREB, pCREB, and Erk1/2 in the ACC. Chronic visceral pain is an important factor in the pathophysiology of irritable bowel syndrome (IBS). The current study provides evidence that astrocytic activity in the ACC is required for visceral pain-related aversive learning and memory., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Visceral pain is a prominent feature of various gastrointestinal diseases. The P2X7 receptor is expressed by multiple cell types including dorsal root ganglion satellite glial cells, macrophages, and spinal microglia, all of which have been implicated in nociceptive sensitization. We have used the selective and CNS penetrant P2X7 receptor antagonist Lu AF27139 to explore this receptor's role in distinct rat models of inflammatory and visceral hypersensitivity. Rats injected with CFA in the hindpaw displayed a marked reduction in hindpaw mechanical threshold, which was dose-dependently reversed by Lu AF27139 (3-30 mg/kg, p.o.). In rats injected with TNBS in the proximal colon, the colorectal distension threshold measured distally was significantly lower than sham treated rats at 7 days post-injection (P < 0.001), indicative of a marked central sensitization. Colonic hypersensitivity was also reversed by Lu AF27139 (10-100 mg/kg) and by the κ-opioid receptor agonist U-50,488H (3 mg/kg, s.c.). Moreover, both Lu AF27139 and U-50,488H prevented a TNBS-induced increase in spinal and brain levels of PGE2 and LTB4, as well as an increase in brain levels of PGF2α and TXB2. Lu AF27139 was well tolerated as revealed by a lack of significant effect on rotarod motor function and coordination at all doses tested up to 300 mg/kg. Thus, P2X7 receptor antagonism is efficacious in a rat model of visceral pain, via a mechanism which potentially involves attenuation of microglial function within spinal and/or supraspinal pain circuits, albeit a peripheral site of action cannot be excluded., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

Chronic visceral pain is a major challenge for both patients and health providers. Although the central sensitization of the brain is thought to play an important role in the development of visceral pain, the detailed neural circuits remain largely unknown. Using a well-established chronic visceral hypersensitivity model induced by neonatal maternal deprivation (NMD) in male mice, we identified a distinct pathway whereby the claustrum (CL) glutamatergic neuron projecting to the anterior cingulate cortex (ACC) is critical for visceral pain but not for CFA-evoked inflammatory pain. By a combination of in vivo circuit-dissecting extracellular electrophysiological approaches and visceral pain related electromyographic (EMG) recordings, we demonstrated that optogenetic inhibition of CL glutamatergic activity suppressed the ACC neural activity and visceral hypersensitivity of NMD mice whereas selective activation of CL glutamatergic activity enhanced the ACC neural activity and evoked visceral pain of control mice. Further, optogenetic studies demonstrate a causal link between such neuronal activity and visceral pain behaviors. Chemogenetic activation or inhibition of ACC neural activities reversed the effects of optogenetic manipulation of CL neural activities on visceral pain responses. Importantly, molecular detection showed that NMD significantly enhances the expression of NMDA receptors and activated CaMKIIα in the ACC postsynaptic density (PSD) region. Together, our data establish a functional role for CL→ACC glutamatergic neurons in gating visceral pain, thus providing a potential treatment strategy for visceral pain. SIGNIFICANCE STATEMENT Studies have shown that sensitization of anterior cingulate cortex (ACC) plays an important role in chronic pain. However, it is as yet unknown whether there is a specific brain region and a distinct neural circuit that helps the ACC to distinguish visceral and somatic pain. The present study demonstrates that claustrum (CL) glutamatergic neurons maybe responding to colorectal distention (CRD) rather than somatic stimulation and that a CL glutamatergic projection to ACC glutamatergic neuron regulates visceral pain in mice. Furthermore, excessive NMDA receptors and overactive CaMKIIα in the ACC postsynaptic density (PSD) region were observed in mice with chronic visceral pain. Together, these findings reveal a novel neural circuity underlying the central sensitization of chronic visceral pain., (Copyright © 2022 the authors.)

Visceral pain is unbearable, and natural methods are needed to relieve it. Electroacupuncture is a relatively new technique that helps relieve visceral pain by improving blood circulation and providing energy to clogged parts of the body. However, its analgesic effect and mechanism in colorectal pain are still unknown. In this study, the visceral pain models of electroacupuncture in rats were compared and discussed, using nanocomponents to stimulate the expression and mechanism of the nerve growth factor in colorectal pain and electroacupuncture and to observe the expression and mechanism of nerve growth factor in visceral pain relief rats induced by nanocomponents and electroacupuncture. The results show that nanocomponents can effectively relieve visceral pain under the action of electroacupuncture. NGF can activate endogenous proliferation, migration, differentiation, and integration. NSC can promote nerve regeneration and recovery after injury., Competing Interests: There are no conflicts of interest related to this research work and article., (Copyright © 2022 Dongdong Liang et al.)

Aims: Visceral hypersensitivity in irritable bowel syndrome (IBS) is widespread, but effective therapies for it remain elusive. As a canonical anti-inflammatory protein, suppressor of cytokine signaling 3 (SOCS3) reportedly relays exchange protein 1 directly activated by cAMP (Epac1) signaling and inhibits the intracellular response to inflammatory cytokines. Despite the inhibitory effect of SOCS3 on the pro-inflammatory response and neuroinflammation in PVN, the systematic investigation of Epac1-SOCS3 signaling involved in visceral hypersensitivity remains unknown. This study aimed to explore Epac1-SOCS3 signaling in the activity of hypothalamic paraventricular nucleus (PVN) corticotropin-releasing factor (CRF) neurons and visceral hypersensitivity in adult rats experiencing neonatal colorectal distension (CRD)., Methods: Rats were subjected to neonatal CRD to simulate visceral hypersensitivity to investigate the effect of Epac1-SOCS3 signaling on PVN CRF neurons. The expression and activity of Epac1 and SOCS3 in nociceptive hypersensitivity were determined by western blot, RT-PCR, immunofluorescence, radioimmunoassay, electrophysiology, and pharmacology., Results: In neonatal-CRD-induced visceral hypersensitivity model, Epac1 and SOCS3 expressions were downregulated and IL-6 levels elevated in PVN. However, infusion of Epac agonist 8-pCPT in PVN reduced CRF neuronal firing rates, and overexpression of SOCS3 in PVN by AAV-SOCS3 inhibited the activation of PVN neurons, reduced visceral hypersensitivity, and precluded pain precipitation. Intervention with IL-6 neutralizing antibody also alleviated the visceral hypersensitivity. In naïve rats, Epac antagonist ESI-09 in PVN increased CRF neuronal firing. Consistently, genetic knockdown of Epac1 or SOCS3 in PVN potentiated the firing rate of CRF neurons, functionality of HPA axis, and sensitivity of visceral nociception. Moreover, pharmacological intervention with exogenous IL-6 into PVN simulated the visceral hypersensitivity., Conclusions: Inactivation of Epac1-SOCS3 pathway contributed to the neuroinflammation accompanied by the sensitization of CRF neurons in PVN, precipitating visceral hypersensitivity and pain in rats experiencing neonatal CRD., (© 2022 The Authors. CNS Neuroscience & Therapeutics published by John Wiley & Sons Ltd.)

Visceral pain is a leading cause of morbidity in gastrointestinal diseases, which is exacerbated by the gut-related side-effects of many analgesics. New treatments are needed and further understanding of the mediators and mechanisms underpinning visceral nociception in disease states is required to facilitate this. The pro-inflammatory cytokine TNFα is linked to pain in both patients with inflammatory bowel disease and irritable bowel syndrome, and has been shown to sensitize colonic sensory neurons. Somatic, TNFα-triggered thermal and mechanical hypersensitivity is mediated by TRPV1 signalling and p38 MAPK activity respectively, downstream of TNFR1 receptor activation. We therefore hypothesized that TNFR1-evoked p38 MAPK activity may also be responsible for TNFα sensitization of colonic afferent responses to the TRPV1 agonist capsaicin, and noxious distension of the bowel. Using Ca 2+ imaging of dorsal root ganglion sensory neurons, we observed TNFα-mediated increases in intracellular [Ca 2+ ] and sensitization of capsaicin responses. The sensitizing effects of TNFα were dependent on TNFR1 expression and attenuated by p38 MAPK inhibition. Consistent with these findings, ex vivo colonic afferent fibre recordings demonstrated an enhanced response to noxious ramp distention of the bowel and bath application of capsaicin following TNFα pre-treatment. Responses were reversed by p38 MAPK inhibition and absent in tissue from TNFR1 knockout mice. Our findings demonstrate a contribution of TNFR1, p38 MAPK and TRPV1 to TNFα-induced sensitization of colonic afferents, highlighting the potential utility of these drug targets for the treatment of visceral pain in gastrointestinal disease. KEY POINTS: The pro-inflammatory cytokine TNFα is elevated in gastrointestinal disease and sensitizes colonic afferents via modulation of TRPA1 and Na V 1.8 activity. We further develop this understanding by demonstrating a role for p38 MAPK and TRPV1 in TNFα-mediated colonic afferent sensitization. Specifically, we show that: TNFα sensitizes sensory neurons and colonic afferents to the TRPV1 agonist capsaicin. TNFα-mediated sensitization of sensory neurons and colonic nociceptors is dependent on TNFR1 expression. TNFα sensitization of sensory neurons and colonic afferents to capsaicin and noxious ramp distension is abolished by inhibition of p38 MAPK. Collectively these data support the utility of targeting TNFα, TNFR1 and their downstream signalling via p38 MAPK for the treatment of visceral pain in gastrointestinal disease., (© 2022 The Authors. The Journal of Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Purpose: To analyze the effect and mechanism of dexmedetomidine (DEX) analgesia pretreatment on functional chronic visceral pain in rats., Methods: Rats were divided into six groups: W1, W2, W3, W4, W5, and W6. The behavioral changes and electrophysiological indexes of rats in each group before and after DEX treatment were detected., Results: The levels of abdominal withdrawal reflex (AWR) in W5 and W6 groups were significantly lower than those in group W3, while the levels of thermal withdrawal latency (TWL) and mechanical withdrawal threshold (MWT) were significantly higher than those in group W3 (p < 0.05). The electromyographic signals of W1, W5, and W6 groups showed little fluctuation, while those of groups W2, W3, and W4 showed obvious fluctuation. TLR4 mRNA expression, IRF3, P65, and phosphorylation levels in W4, W5, and W6 groups were significantly lower than those in group W2 (p < 0.05)., Conclusions: Dexmedetomidine epidural anesthesia pretreatment could significantly inhibit visceral pain response in rats with functional chronic visceral pain, and its mechanism was related to the activation of TLR4 in spinal dorsal horn tissue of rats and the activation inhibition of IRF3 and P65 in the downstream key signals.

Aims: Sustained visceral hypersensitivity is a hallmark of irritable bowel syndrome (IBS) could be partially explained by enteric neural remodeling. Particularly, synaptic plasticity in the enteric nervous system, a form of enteric "memory", has been speculated as a participant in the pain maintenance in IBS. This study aimed to elucidate the role of ephrinB2/ephB2 in enteric synaptic plasticity and visceral pain in IBS., Materials and Methods: EphrinB2/ephB2 expression and synaptic plasticity were assessed in colonic tissues from IBS patients, and rats induced by Trichinella spiralis infection and those treated with ephB2-Fc (an ephB2 receptor blocker) or ifenprodil (a selective NR2B antagonist). Furthermore, abdominal withdrawal reflex scores to colorectal distention and mesenteric afferent firing were assessed. EphrinB2-Fc(an ephB2 receptor activator) induced enteric synaptic plasticity was further evaluated in longitudinal muscle-myenteric plexus(LMMP) cultures and primary cultured myenteric neurons., Key Findings: EphrinB2/ephB2 was specifically expressed in colonic nerves and upregulated in IBS patients and rats, which was correlated with pain severity. The functional synaptic plasticity, visceral sensitivity to colorectal distention and colonic mesenteric afferent activity to mechanical and chemical stimulus were enhanced in IBS rats, and were blocked by ephB2-Fc or ifenprodil treatment. EphrinB2-Fc promoted the phosphorylation of NR2B in IBS rats and LMMP cultures, and could mediate sustained neural activation via increased [Ca 2+ ] i and raised expression of synaptic plasticity-related early immediate genes, including c-fos and arc., Significance: EphrinB2/ephB2 facilitated NR2B-mediated synaptic potentiation in the enteric nervous system that may be a novel explanation and potential therapeutic target for sustained pain hypersensitivity in IBS., (Copyright © 2022. Published by Elsevier Inc.)

Irritable bowel syndrome is a gastrointestinal disorder of unknown etiology characterized by widespread, chronic abdominal pain associated with altered bowel movements. Increasing amounts of evidence indicate that injury and inflammation during the neonatal period have long-term effects on tissue structure and function in the adult that may predispose to gastrointestinal diseases. In this study we aimed to investigate how the epigenetic regulation of DNA demethylation of the p2x7r locus guided by the transcription factor GATA binding protein 1 (GATA1) in spinal astrocytes affects chronic visceral pain in adult rats with neonatal colonic inflammation (NCI). The spinal GATA1 targeting to DNA demethylation of p2x7r locus in these rats was assessed by assessing GATA1 function with luciferase assay, chromatin immunoprecipitation, patch clamp, and interference in vitro and in vivo. In addition, a decoy oligodeoxynucleotide was designed and applied to determine the influence of GATA1 on the DNA methylation of a p2x7r CpG island. We showed that NCI caused the induction of GATA1, Ten-eleven translocation 3 (TET3), and purinergic receptors (P2X7Rs) in astrocytes of the spinal dorsal horn, and demonstrated that inhibiting these molecules markedly increased the pain threshold, inhibited the activation of astrocytes, and decreased the spinal sEPSC frequency. NCI also markedly demethylated the p2x7r locus in a manner dependent on the enhancement of both a GATA1-TET3 physical interaction and GATA1 binding at the p2x7r promoter. Importantly, we showed that demethylation of the p2x7r locus (and the attendant increase in P2X7R expression) was reversed upon knockdown of GATA1 or TET3 expression, and demonstrated that a decoy oligodeoxynucleotide that selectively blocked the GATA1 binding site increased the methylation of a CpG island in the p2x7r promoter. These results demonstrate that chronic visceral pain is mediated synergistically by GATA1 and TET3 via a DNA-demethylation mechanism that controls p2x7r transcription in spinal dorsal horn astrocytes, and provide a potential therapeutic strategy by targeting GATA1 and p2x7r locus binding., (© 2021. The Author(s).)

Chronic visceral pain (CVP) is one of the common symptoms of many diseases triggered by underlying diseases of the internal organs of the human body. Its causes include vascular mechanisms, mechanical factors, persistent inflammation, and unexplained functional mechanisms. Although the pathogenesis is unclear, more and more research has begun to shift from the neuronal aspect to the glial cells in recent years. Some data highlight that the spinal glial cells, particularly the microglia and astrocytes, play an essential role in CVP. Based on this, we highlight the mechanisms of microglia and astrocytes in CVP concerning the release of cytokines, chemokines, and neuroactive substances and alterations in intracellular signaling pathways during the process. Finally, because CVP is widespread in various diseases, we present future perspectives targeting microglia and astrocytes for treatment., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Cognitive behavioral therapy (CBT) improves the quality of life for patients with brain-gut disorders; however, the underlying mechanisms of CBT remain to be explored. Previously, we showed that environmental enrichment (EE), an experimental paradigm that mirrors positive behavioral intervention, ameliorates chronic stress-induced visceral hypersensitivity in a rodent model via mechanisms involving altered activity in the central nucleus of amygdala (CeA). In the present study, we investigated whether microglia-mediated synaptic plasticity in the CeA is a potential mechanism underlying the protective effects of EE against stress-induced visceral hypersensitivity. We stereotaxically implanted corticosterone (CORT) micropellets onto the dorsal margin of the CeA shown previously to induce colonic hypersensitivity. Animals were housed in EE cages or standard cages for 14 days after CORT implantation. Visceral sensitivity was assessed via visceromotor behavioral response to colorectal distension. Microglial morphology, microglia-mediated synaptic engulfment, and the expression of synaptic pruning-related signals complement component 1q (C1q), complement component 3 (C3), and C3 receptor (C3R) were measured using immunofluorescence and RNAscope assay. We found that housing CORT implanted rats in EE cages for 14 days attenuated visceral hypersensitivity in both male and female rats as compared with control rats maintained in standard housing. EE reduced CORT-induced microglial remodeling and microglia-mediated synaptic pruning with reduced C1q and CR3, but not C3, expression. Our data suggest that exposure to EE is sufficient to ameliorate stress-induced visceral pain via reducing amygdala microglia-modulated neuronal plasticity. NEW & NOTEWORTHY Clinical studies show that cognitive behavioral therapy (CBT) is effective in ameliorating visceral pain in patient with irritable bowel syndrome (IBS), yet the underlying mechanisms remain unexplored. By using environmental enrichment (EE), an experimental paradigm that mirrors positive behavioral intervention, we demonstrated that microglia-mediated synaptic plasticity in the CeA explains, plays a role, at least in part, in the positive effects of EE to reduce visceral hypersensitivity.

Background: Inflammatory visceral pain is endogenously controlled by enkephalins locally released by mucosal CD4 + T lymphocytes in mice. The present study aimed at identifying opioid receptor(s) expressed on nociceptive sensory nerves involved in this peripheral opioid-mediated analgesia., Methods: The peripheral analgesia associated with the accumulation of CD4 + T lymphocytes within the inflamed colonic mucosa was assessed in conditional knockout mice specifically deleted for either of the two opioid receptors for enkephalins (i.e., µ (MOR) and δ (DOR) receptors) in Na v 1.8-expressing sensory neurons in the dextran sulfate sodium (DSS)-induced colitis model., Results: Endogenous analgesia is lost in conditional knockout mice for DOR, but not MOR at the later phase of the DSS-induced colitis. The absence of either of the opioid receptors on sensory nerves had no impact on both the colitis severity and the rate of T lymphocytes infiltrating the inflamed colonic mucosa., Conclusion: The key role of DOR on primary afferents in relieving intestinal inflammatory pain opens new therapeutic opportunities for peripherally restricted DOR analgesics to avoid most of the side effects associated with MOR-targeting drugs used in intestinal disorders., (© 2022. The Author(s).)

Patients with post-traumatic stress disorder (PTSD) are usually at an increased risk for chronic disorders, such as irritable bowel syndrome (IBS), characterized by hyperalgesia and allodynia, but its subsequent effect on visceral hyperalgesia and the mechanism remain unclear. The present study employed single prolonged stress (SPS), a model of PTSD-pain comorbidity, behavioral evaluation, intrathecal drug delivery, immunohistochemistry, Western blotting, and RT-PCR techniques. When detecting visceral sensitivity, the score of the abdominal withdrawal reflex (AWR) induced by graded colorectal distention (CRD) was used. The AWR score was reduced in the SPS day 1 group but increased in the SPS day 7 and SPS day 14 groups at 40 mmHg and 60 mmHg, and the score was increased significantly with EphrinB1-Fc administration. The EphB2+ cell density and EphB2 protein and mRNA levels were downregulated in the SPS day 1 group and then upregulated significantly in the SPS day 7 group; these changes were more noticeable with EphrinB1-Fc administration compared with the SPS-only group. The C-Fos-positive reaction induced by SPS was mainly localized in neurons of the spinal dorsal horn, in which the C-Fos-positive cell density and its protein and mRNA levels were upregulated on SPS days 7 and 14; these changes were statistically significant in the SPS + EphrinB1-Fc group compared with the SPS alone group. The present study confirmed the time window for the AWR value, EphB2 and C-Fos changes, and the effect of EphrinB1-Fc on these changes, which suggests that spinal cord EphB2 activation exacerbates visceral pain after SPS.

The current study aimed to investigate the effects of sodium butyrate on the level of colonic protein IRAK1 (interleukin-1 receptor-associated kinase 1) in irritable bowel syndrome (IBS) models as well as revealing the relationship between IRAKI level and visceral sensitivity during the progression of IBS. IBS symptoms were induced using TNBS (2,4,6-trinitrobenzene sulfonic acid) in mice and using IL-33 in HT-29 cells, which were then hanlded with sodium butyrate (100 mM for each mice and 0.05 M for HT-29 cells). The threshold of visceral pain and the expression of IRAKI in mice, and the level of IRAKI in HT-29 cells were detected. The data showed that the level of IRAK1 in IBS mice was higher than that in the control group, while the pre-treatment with sodium butyrate could solidy suppressed the level of IRAK1. Morevoer, it was found that the level of IRAK1 was negatively correlated with the pain threshold. In in vitro assays, the level of IRAK1 was firstly induced by IL-33 stimulation and then suppressed by sodium butyrate pretreatment. Collectively, the level of IRAKI showed an obvioulty positive relation with visceral hypersensitivity in IBS models, and the treatment with sodium butyrate could alleviate visceral hypersensitivity by inhibiting the expression of IRAKI.

Background: Visceral and parietal peritoneum layers have different sensory innervations. Most visceral peritoneum sensory information is conveyed via the vagus nerve to the nucleus of the solitary tract (NTS). We already showed in animal models that intramuscular (i.m.) injection of local anesthetics decreases acute somatic and visceral pain and general inflammation induced by aseptic peritonitis. The goal of the study was to compare the effects of parietal block, i.m. bupivacaine, and vagotomy on spinal cord and NTS stimulation induced by a chemical peritonitis., Methods: We induced peritonitis in rats using carrageenan and measured cellular activation in spinal cord and NTS under the following conditions, that is, a parietal nerve block with bupivacaine, a chemical right vagotomy, and i.m. microspheres loaded with bupivacaine. Proto-oncogene c-Fos (c-Fos), cluster of differentiation protein 11b (CD11b), and tumor necrosis factor alpha (TNF-α) expression in cord and NTS were studied., Results: c-Fos activation in the cord was inhibited by nerve block 2 hours after peritoneal insult. Vagotomy and i.m. bupivacaine similarly inhibited c-Fos activation in NTS. Forty-eight hours after peritoneal insult, the number of cells expressing CD11b significantly increased in the cord (P = .010). The median difference in the effect of peritonitis compared to control was 30 cells (CI95, 13.5-55). TNF-α colocalized with CD11b. Vagotomy inhibited this microglial activation in the NTS, but not in the cord. This activation was inhibited by i.m. bupivacaine both in cord and in NTS. The median difference in the effect of i.m. bupivacaine added to peritonitis was 29 cells (80% increase) in the cord and 18 cells (75% increase) in the NTS. Our study underlines the role of the vagus nerve in the transmission of an acute visceral pain message and confirmed that systemic bupivacaine prevents noxious stimuli by inhibiting c-Fos and microglia activation., Conclusions: In rats receiving intraperitoneal carrageenan, i.m. bupivacaine similarly inhibited c-Fos and microglial activation both in cord and in the NTS. Vagal block inhibited activation only in the NTS. Our study underlines the role of the vagus nerve in the transmission of an acute visceral pain message and confirmed that systemic bupivacaine prevents noxious stimuli. This emphasizes the effects of systemic local anesthetics on inflammation and visceral pain., Competing Interests: Conflicts of Interest: See Disclosures at the end of the article., (Copyright © 2021 International Anesthesia Research Society.)

Accumulating evidence shows that agents targeting gut dysbiosis are effective for improving symptoms of irritable bowel syndrome (IBS). However, the potential mechanisms remain unclear. In this study we investigated the effects of berberine on the microbiota-gut-brain axis in two rat models of visceral hypersensitivity, i.e., specific pathogen-free SD rats subjected to chronic water avoidance stress (WAS) and treated with berberine (200 mg· kg -1  ·d -1 , ig, for 10 days) as well as germ-free (GF) rats subjected to fecal microbiota transplantation (FMT) from a patient with IBS (designated IBS-FMT) and treated with berberine (200 mg· kg -1  ·d -1 , ig, for 2 weeks). Before the rats were sacrificed, visceral sensation and depressive behaviors were evaluated. Then colonic tryptase was measured and microglial activation in the dorsal lumbar spinal cord was assessed. The fecal microbiota was profiled using 16S rRNA sequencing, and short chain fatty acids (SCFAs) were measured. We showed that berberine treatment significantly alleviated chronic WAS-induced visceral hypersensitivity and activation of colonic mast cells and microglia in the dorsal lumbar spinal cord. Transfer of fecal samples from berberine-treated stressed donors to GF rats protected against acute WAS. FMT from a patient with IBS induced visceral hypersensitivity and pro-inflammatory phenotype in microglia, while berberine treatment reversed the microglial activation and altered microbial composition and function and SCFA profiles in stools of IBS-FMT rats. We demonstrated that berberine did not directly influence LPS-induced microglial activation in vitro. In both models, several SCFA-producing genera were enriched by berberine treatment, and positively correlated to the morphological parameters of microglia. In conclusion, activation of microglia in the dorsal lumbar spinal cord was involved in the pathogenesis of IBS caused by dysregulation of the microbiota-gut-brain axis, and the berberine-altered gut microbiome mediated the modulatory effects of the agent on microglial activation and visceral hypersensitivity, providing a potential option for the treatment of IBS., (© 2021. The Author(s).)

Introduction: Stress is a known trigger for the symptoms of irritable bowel syndrome (IBS), a gastrointestinal (GI) disorder that presents with abnormal bowel habits and abdominal pain due to visceral hypersensitivity. While behavioral therapies have been used to attenuate IBS symptoms, the underlying mechanisms by which these therapies interact with stress-induced pathology remains to be delineated. Here we use a rat model to test the hypothesis that exposure to environmental enrichment (EE) inhibits stress-induced changes within the brain-gut axis to prevent visceral and somatic hypersensitivity and colonic hyperpermeability., Methods: Female rats (n = 8/group) were housed in EE one week before and one week during exposure to water avoidance stress (WAS) while controls were housed in standard cages (SH). One day after the final WAS exposure, colonic and somatic sensitivity were assessed by the visceromotor response (VMR) to colorectal distension (CRD) and withdrawal threshold elicited by an electronic von Frey on the hind paw of the rats respectively. All rats were returned to SH for 3 weeks before colonic and somatic sensitivity were reassessed on day 28. The rats were then immediately euthanized and the spinal cord was collected to assess changes in neuronal activation (assessed via ERK phosphorylation) in response to noxious CRD. A separate cohort of animals (n = 8/group) that did not undergo behavioral assessments was euthanized the day after the final WAS exposure and the central nucleus of the amygdala (CeA) was collected to investigate WAS and EE induced epigenetic changes at the glucocorticoid receptor (GR) and corticotrophin releasing hormone (CRH) promoter. The colon from these rats was also collected to assess colonic permeability via changes in transepithelial electrical resistance (TEER) in vitro., Results: Exposure to stress persistently increased VMR to CRD (P < 0.01) and decreased the hind paw withdrawal threshold (P < 0.001) in female rats. WAS also decreased TEER in the colon tissue of female rats (p = 0.05). In the CeA, WAS induced a decrease in histone acetylation at the GR promoter but increased histone acetylation at the CRH promoter and reduced GR-CRH interactions in the CeA. Analysis of the spinal cord showed that WAS increased CRD-evoked ERK phosphorylation in the dorsal horn. Exposure to EE prevented WAS-induced changes in the CeA, dorsal horn and colon respectively to prevent visceral and somatic hypersensitivity., Conclusion: Our data reveals that behavioral therapies can produce long lasting molecular and epigenetic changes that can prevent stress-induced pathologies even after completion of the therapy. These results highlight the potential mechanisms by which behavioral therapies may ameliorate visceral pain associated stress-related pathologies such as the irritable bowel syndrome., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Background: We previously reported that female offspring of dams subjected to chronic prenatal stress (CPS) develop enhanced visceral hypersensitivity (VHS) following exposure to chronic stress in adult life that is mediated by up-regulation of spinal cord BDNF. The aims of this study were to examine the roles of estrogen receptor alpha (ERα) and an increase in spinal serotonin signaling in promoting this enhanced VHS in female rats and up-regulation of spinal cord BDNF transcription., Methods: Pregnant dams were exposed to chronic stress from E11 until delivery. At 8 weeks, a chronic adult stress (CAS) protocol was applied for nine days., Key Results: Ovariectomy before CAS or treatment with letrozole before and during CAS significantly prevented the development of enhanced VHS in female CPS+CAS rats. Intrathecal application of ERα siRNA significantly reduced VHS, decreased lumbar-sacral spinal cord expression of both ERα and BDNF, and reversed pro-transcriptional epigenetic modifications at BDNF promoter lX. Cerebrospinal fluid serotonin levels and 5HT3A receptor expression in the LS spinal cord were both significantly increased in female CPS+CAS rats. During CAS, intrathecal infusion of alosetron significantly decreased VHS, reduced BDNF and ERα expression in the LS spinal cord, and attenuated RNA pol II and ERα binding to the BNDF core promoter IX., Conclusions & Inferences: Serotonin-mediated activation of 5HT3A receptors in the spinal cord drives the development of enhanced female-specific VHS in our two hit CPS+CAS through up-regulation of spinal cord ERα., (© 2021 John Wiley & Sons Ltd.)

Persistent pain affecting patients with inflammatory bowel diseases (IBDs) is still very difficult to treat. Carbonic anhydrase (CA) represents an intriguing pharmacological target considering the anti-hyperalgesic efficacy displayed by CA inhibitors in both inflammatory and neuropathic pain models. The aim of this work was to evaluate the effect of inhibiting CA IV, particularly when expressed in the gut, on visceral pain associated with colitis induced by 2,4-di-nitrobenzene sulfonic acid (DNBS) in rats. Visceral sensitivity was assessed by measuring animals' abdominal responses to colorectal distension. Repeated treatment with the selective CA IV inhibitors AB-118 and NIK-67 effectively counteracted the development of visceral pain induced by DNBS. In addition to pain relief, AB-118 showed a protective effect against colon damage. By contrast, the anti-hyperalgesic activity of NIK-67 was independent of colon healing, suggesting a direct protective effect of NIK-67 on visceral sensitivity. The enzymatic activity and the expression of CA IV resulted significantly increased after DNBS injection. NIK-67 normalised CA IV activity in DNBS animals, while AB-118 was partially effective. None of these compounds influenced CA IV expression through the colon. Although further investigations are needed to study the underlying mechanisms, CA IV inhibitors are promising candidates in the search for therapies to relieve visceral pain in IBDs.

Abnormal brain-gut interactions contribute to the development of chronic visceral hypersensitivity (CVH), which is the pivotal feature of irritable bowel syndrome (IBS). Despite the consensus with respect to the vital role of hyperpolarization-activated cyclic nucleotide-gated 2 (HCN2) channels in promoting painful symptoms in the peripheral nervous system, we identified that the upregulation of HCN2 in supraoptic nucleus (SON) was involved in the modulation of CVH in rat model of neonatal colorectal distention (n-CRD). Specifically, colorectal distention (CRD) upregulated the expression of c-Fos in SON in adult CVH rats, indicating the involvement of SON sensitazation in visceral sensation. Moreover, the administration of ZD7288 (the pan-HCN channel inhibitor) rather than 8-Br-cAMP (the non-specific HCN channel agonist) aggravated the CVH symptoms and reduced the phosphorylation level of CaMKII-CREB cascade. Together, the findings indicated that the upregulation of supraoptic HCN2 contributed to the sensitization of SON, which had protective effects on the modulation of CVH with the involvement of CaMKII-CREB cascade in n-CRD rat model., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Interactions between the intestinal microbiota, immune system and nervous system are essential for homeostasis in the gut. Inflammasomes contribute to innate immunity and brain-gut interactions, but their role in microbiota-neuro-immune interactions is not clear. Therefore, we investigated the effect of the inflammasome on visceral pain and local and systemic neuroimmune responses after antibiotic-induced changes to the microbiota. Wild-type (WT) and caspase-1/11 deficient (Casp1 KO) mice were orally treated for 2 weeks with an antibiotic cocktail (Abx, Bacitracin A and Neomycin), followed by quantification of representative fecal commensals (by qPCR), cecal short chain fatty acids (by HPLC), pathways implicated in the gut-neuro-immune axis (by RT-qPCR, immunofluorescence staining, and flow cytometry) in addition to capsaicin-induced visceral pain responses. Abx-treatment in WT-mice resulted in an increase in colonic macrophages, central neuro-immune interactions, colonic inflammasome and nociceptive receptor gene expression and a reduction in capsaicin-induced visceral pain. In contrast, these responses were attenuated in Abx-treated Casp1 KO mice. Collectively, the data indicate an important role for the inflammasome pathway in functional and inflammatory gastrointestinal conditions where pain and alterations in microbiota composition are prominent.

Chronic abdominal pain is a common clinical condition experienced by patients with irritable bowel syndrome (IBS). A general lack of suitable treatment options for the management of visceral pain is the major contributing factor to the debilitating nature of the disease. Understanding the underlying causes of chronic visceral pain is pivotal to identifying new effective therapies for IBS. This review provides the current evidence, demonstrating that mediators and receptors that induce itch in the skin also act as "gut irritants" in the gastrointestinal tract. Activation of these receptors triggers specific changes in the neuronal excitability of sensory pathways responsible for the transmission of nociceptive information from the periphery to the central nervous system leading to visceral hypersensitivity and visceral pain. Accumulating evidence points to significant roles of irritant mediators and their receptors in visceral hypersensitivity and thus constitutes potential targets for the development of more effective therapeutic options for IBS.

Stress can trigger symptoms in patients with irritable bowel syndrome (IBS). Previously we demonstrated that chronic psychological stress induced microglial remodeling in the central nucleus of amygdala (CeA) and contributed to the development of visceral hypersensitivity via synaptic engulfment. However, the specific signaling mechanisms that microglia depend upon to recognize target neurons to facilitate visceral pain remain unknown. Here, we test the hypothesis that the microglia in the CeA contribute to chronic stress-induced visceral hypersensitivity via complement C1q/C3-CR3 signaling-mediated synaptic remodeling. In male and female Fischer-344 rats, micropellets of corticosterone (CORT) or cholesterol (control) were stereotaxically implanted bilaterally onto the CeA. After 7 days, microglial C1q, complement receptor 3 (CR3) expression, and microglia-mediated synaptic engulfment were assessed via RNAscope, quantitative PCR, and immunofluorescence. The microglial inhibitor minocycline, CR3 antagonist neutrophil inhibitory factor (NIF), or vehicle were daily infused into the CeA following CORT implantations. Visceral sensitivity was assessed via a visceromotor response (VMR) to graded pressures of isobaric colorectal distension (CRD). Our results suggest that chronic exposure to elevated CORT in the CeA induced visceral hypersensitivity and amygdala microglial morphological remodeling. CORT increased microglial C1q and CR3 expression and increased microglia-mediated synaptic engulfment. Both groups of animals with minocycline or NIF infusions reversed microglia-mediated synaptic remodeling and attenuated CORT-induced visceral hypersensitivity. Our findings demonstrate that C1q/C3-CR3 signaling is critical for microglia-mediated synaptic remodeling in the CeA and contributes to CORT-induced visceral hypersensitivity. NEW & NOTEWORTHY Patients with irritable bowel syndrome (IBS) show altered amygdala activity. We showed previously that stress induces visceral hypersensitivity partially through microglia-modulated synaptic plasticity in the central nucleus of the amygdala (CeA). Our current data suggest that the C1q/C3-CR3 cascade initiates microglia-mediated synaptic remodeling in the CeA. Blocking C3-CR3 interaction attenuates stress-induced visceral hypersensitivity. These findings uncover a role of microglia-synapse signaling in the brain-gut regulation and support a future therapeutic target to treat visceral pain.

Irritable bowel syndrome (IBS) is one of the most common challenging diseases for clinical treatment. The aim of this study is to investigate whether transcranial direct current stimulation (tDCS) has analgesic effect on visceral hypersensitivity (VH) in an animal model of IBS as well as the underlying mechanism. As the activation of GluN2B in anterior cingulate cortex (ACC) takes part in VH, we examined whether and how GluN2B in ACC takes part in the effect of tDCS. Neonatal maternal deprivation (NMD), a valuable experimental model to study the IBS pathophysiology, was used to induce visceral hypersensitivity of rats. We quantified VH as colorectal distention threshold and performed patch-clamp recordings of ACC neurons. The expression of GluN2B were determined by RT-qPCR and Western blotting. The GluN2B antagonist Ro 25-6981 was microinjected into the rostral and caudal ACC. tDCS was performed for 7 consecutive days. It was found that NMD decreased expression of GluN2B, which could be obviously reversed by tDCS. Injection of Ro 25-6981 into rostral and caudal ACC of normal rats induced VH and also reversed the analgesic effect of tDCS. Our data sheds light on the nonpharmacological therapy for chronic VH in pathological states such as IBS. NEW & NOTEWORTHY Irritable bowel syndrome (IBS) is a gastrointestinal disease characterized by visceral hypersensitivity. This study showed a decrease of GluN2B expression and neural activity in ACC of IBS-model rats, which could be obviously reversed by tDCS. In addition, blockade of GluN2B in rostral and caudal ACC induced VH of normal rats. Furthermore, analgesic effect of tDCS on NMD rats was reversed by GluN2B antagonist.

Early-life stress (ELS) is a high-risk factor for the development of chronic visceral pain in adulthood. Emerging evidence suggests that mast cells play a key role in the development of visceral hypersensitivity through interaction with neurons. The sensitization of corticotropin-releasing factor (CRF) neurons in the hypothalamic paraventricular nucleus (PVN) plays a pivotal role in the pathogenesis of visceral pain. However, the precise mechanism by which mast cells and CRF neurons interact in the PVN in the pathogenesis of visceral hypersensitivity remains elusive. In the present study, we used neonatal maternal separation (MS), an ELS model, and observed that neonatal MS induced visceral hypersensitivity and triggered PVN mast cell activation in adult rats, which was repressed by intra-PVN infusion of the mast cell stabilizer disodium cromoglycate (cromolyn). Wild-type (WT) mice but not mast cell-deficient Kit W-sh/W-sh mice that had experienced neonatal MS exhibited chronic visceral hypersensitivity. MS was associated with an increase in the expression of proinflammatory mediators, the number of CRF + cells and CRF protein in the PVN, which was prevented by intra-PVN infusion of cromolyn. Furthermore, we demonstrated that intra-PVN infusion of the mast degranulator compound 48/80 significantly induced mast cell activation, resulting in proinflammatory mediator release, CRF neuronal sensitization, and visceral hypersensitivity, which was suppressed by cromolyn. Overall, our findings demonstrated that neonatal MS induces the activation of PVN mast cells, which secrete numerous proinflammatory mediators that may participate in neighboring CRF neuronal activity, ultimately directly inducing visceral hypersensitivity in adulthood., (Copyright © 2021 Elsevier B.V. All rights reserved.)