OCP002, a Mixed Agonist of Opioid and Cannabinoid Receptors, Produces Potent Antinociception With Minimized Side Effects.

BACKGROUND: Increasing attention has been attracted to the development of bifunctional compounds to minimize the side effects of opioid analgesics. Pharmacological studies have verified the functional interaction between opioid and cannabinoid systems in pain management, suggesting that coactivation of the opioid and cannabinoid receptors may provide synergistic analgesia with fewer adverse reactions. Herein, we developed and characterized a novel bifunctional compound containing the pharmacophores of the mu-opioid receptor agonist DALDA and the cannabinoid peptide VD-Hpα-NH2, named OCP002. METHODS: The opioid and cannabinoid agonistic activities of OCP002 were investigated in calcium mobilization and western blotting assays, respectively. Moreover, the central and peripheral antinociceptive effects of OCP002 were evaluated in mouse preclinical models of tail-flick test, carrageenan-induced inflammatory pain, and acetic acid-induced visceral pain, respectively. Furthermore, the potential opioid and cannabinoid side effects of OCP002 were systematically investigated in mice after intracerebroventricular (ICV) and subcutaneous (SC) administrations. RESULTS: OCP002 functioned as a mixed agonist toward mu-opioid, kappa-opioid, and cannabinoid CB1 receptors in vitro. ICV and SC injections of OCP002 produced dose-dependent antinociception in mouse models of nociceptive (the median effective dose [ED50] values with 95% confidence interval [CI] are 0.14 [0.12-0.15] nmol and 0.32 [0.29-0.35] µmol/kg for ICV and SC injections, respectively), inflammatory (mechanical stimulation: ED50 values [95% CI] are 0.76 [0.64-0.90] nmol and 1.23 [1.10-1.38] µmol/kg for ICV and SC injections, respectively; thermal stimulation: ED50 values [95% CI] are 0.13 [0.10-0.17] nmol and 0.23 [0.08-0.40] µmol/kg for ICV and SC injections, respectively), and visceral pain (ED50 values [95% CI] are 0.0069 [0.0050-0.0092] nmol and 1.47 [1.13-1.86] µmol/kg for ICV and SC injections, respectively) via opioid and cannabinoid receptors. Encouragingly, OCP002 cannot cross the blood-brain barrier and exerted nontolerance-forming analgesia over 6-day treatment at both supraspinal and peripheral levels. Consistent with these behavioral results, repeated OCP002 administration did not elicit microglial hypertrophy and proliferation, the typical features of opioid-induced tolerance, in the spinal cord. Furthermore, at the effective analgesic doses, SC OCP002 exhibited minimized opioid and cannabinoid side effects on motor performance, body temperature, gastric motility, physical and psychological dependence, as well as sedation in mice. CONCLUSIONS: This study demonstrates that OCP002 produces potent and nontolerance-forming antinociception in mice with reduced opioid- and cannabinoid-related side effects, which strengthen the candidacy of bifunctional drugs targeting opioid/cannabinoid receptors for translational-medical development to replace or assist the traditional opioid analgesics.

Effect of butorphanol on visceral pain in patients undergoing gastrointestinal endoscopy: a randomized controlled trial.

BACKGROUND: Butorphanol slightly influences the respiratory and circulatory systems, has a better effect on relieving the discomfort caused by mechanical traction, and has a low incidence of postoperative nausea and vomiting (PONV). Combined butorphanol and propofol may suppress postoperative visceral pain, which is avoidable in gastrointestinal endoscopy. Thus, we hypothesized that butorphanol could decrease the incidence of postoperative visceral pain in patients undergoing gastroscopy and colonoscopy. METHODS: This was a randomized, placebo-controlled, and double-blinded trial. Patients undergoing gastrointestinal endoscopy were randomized to intravenously receive either butorphanol (Group I) or normal saline (Group II). The primary outcome was visceral pain after the procedure 10 min after recovery. The secondary outcomes included the rate of safety outcomes and adverse events. Postoperative visceral pain was defined as a visual analog scale (VAS) score ≥ 1. RESULTS: A total of 206 patients were enrolled in the trial. Ultimately, 203 patients were randomly assigned to Group I (n = 102) or Group II (n = 101). In total, 194 patients were included in the analysis: 95 in Group I and 99 in Group II. The incidence of visceral pain at 10 min after recovery was found to be statistically lower with butorphanol than with the placebo (31.5% vs. 68.5%, respectively; RR: 2.738, 95% CI [1.409-5.319], P = 0.002), and the notable difference was in pain level or distribution of visceral pain (P = 0.006). CONCLUSIONS: The trial indicated that adding butorphanol to propofol results in a lower incidence of visceral pain after surgery without noticeable fluctuations in circulatory and respiratory functions for gastrointestinal endoscopy patients. TRIAL REGISTRATION: Clinicaltrials.gov NCT04477733 (PI: Ruquan Han; date of registration: 20/07/2020).

Bifunctional Peptidomimetic G Protein-Biased Mu-Opioid Receptor Agonist and Neuropeptide FF Receptor Antagonist KGFF09 Shows Efficacy in Visceral Pain without Rewarding Effects after Subcutaneous Administration in Mice.

There is still an unmet clinical need to develop new pharmaceuticals for effective and safe pain management. Current pharmacotherapy offers unsatisfactory solutions due to serious side effects related to the chronic use of opioid drugs. Prescription opioids produce analgesia through activation of the mu-opioid receptor (MOR) and are major contributors to the current opioid crisis. Multifunctional ligands possessing activity at more than one receptor represent a prominent therapeutic approach for the treatment of pain with fewer adverse effects. We recently reported on the design of a bifunctional MOR agonist/neuropeptide FF receptor (NPFFR) antagonist peptididomimetic, KGFF09 (H-Dmt-DArg-Aba-ßAla-Bpa-Phe-NH2), and its antinociceptive effects after subcutaneous (s.c.) administration in acute and persistent pain in mice with reduced propensity for unwanted side effects. In this study, we further investigated the antinociceptive properties of KGFF09 in a mouse model of visceral pain after s.c. administration and the potential for opioid-related liabilities of rewarding and sedation/locomotor dysfunction following chronic treatment. KGFF09 produced a significant dose-dependent inhibition of the writhing behavior in the acetic acid-induced writhing assay with increased potency when compared to morphine. We also demonstrated the absence of harmful effects caused by typical MOR agonists, i.e., rewarding effects (conditioned-place preference test) and sedation/locomotor impairment (open-field test), at a dose shown to be highly effective in inhibiting pain behavior. Consequently, KGFF09 displayed a favorable benefit/side effect ratio regarding these opioid-related side effects compared to conventional opioid analgesics, such as morphine, underlining the development of dual MOR agonists/NPFFR antagonists as improved treatments for various pain conditions.

Acute Visceral Pain in Rats: Vagal Nerve Block Compared to Bupivacaine Administered Intramuscularly.

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.

Effects of Bepridil and Pimozide, Existing Medicines Capable of Blocking T-Type Ca2+ Channels, on Visceral Pain in Mice.

T-Type Ca2+ channels (T-channels), particularly Cav3.2, are now considered as therapeutic targets for treatment of intractable pain including visceral pain. Among existing medicines, bepridil, a multi-channel blocker, used for treatment of arrhythmia and angina, and pimozide, a dopamine D2 receptor antagonist, known as a typical antipsychotic, have potent T-channel blocking activity. We thus tested whether bepridil and pimozide could suppress visceral pain in mice. Colonic and bladder pain were induced by intracolonic administration of 2,4,6-trinitrobenzene sulfonic acid (TNBS) and systemic administration of cyclophosphamide (CPA), respectively. Referred hyperalgesia was assessed by von Frey test, and colonic hypersensitivity to distension by a volume load with intracolonic water injection and spontaneous bladder pain were evaluated by observing nociceptive behaviors in conscious mice. The mice exhibited referred hyperalgesia and colonic hypersensitivity to distension on day 6 after TNBS treatment. Systemic administration of bepridil at 10-20 mg/kg or pimozide at 0.1-0.5 mg/kg strongly reduced the referred hyperalgesia on the TNBS-induced referred hyperalgesia and colonic hypersensitivity to distension. CPA treatment caused bladder pain-like nociceptive behavior and referred hyperalgesia, which were reversed by bepridil at 10-20 mg/kg or pimozide at 0.5-1 mg/kg. Our data thus suggest that bepridil and pimozide, existing medicines capable of blocking T-channels, are useful for treatment of colonic and bladder pain, and serve as seeds for the development of new medicines for visceral pain treatment.

Extrinsic Primary Afferent Neurons Link Visceral Pain to Colon Motility Through a Spinal Reflex in Mice.

BACKGROUND & AIMS: Proper colon function requires signals from extrinsic primary afferent neurons (ExPANs) located in spinal ganglia. Most ExPANs express the vanilloid receptor TRPV1, and a dense plexus of TRPV1-positive fibers is found around myenteric neurons. Capsaicin, a TRPV1 agonist, can initiate activity in myenteric neurons and produce muscle contraction. ExPANs might therefore form motility-regulating synapses onto myenteric neurons. ExPANs mediate visceral pain, and myenteric neurons mediate colon motility, so we investigated communication between ExPANs and myenteric neurons and the circuits by which ExPANs modulate colon function. METHODS: In live mice and colon tissues that express a transgene encoding the calcium indicator GCaMP, we visualized levels of activity in myenteric neurons during smooth muscle contractions induced by application of capsaicin, direct colon stimulation, stimulation of ExPANs, or stimulation of preganglionic parasympathetic neuron (PPN) axons. To localize central targets of ExPANs, we optogenetically activated TRPV1-expressing ExPANs in live mice and then quantified Fos immunoreactivity to identify activated spinal neurons. RESULTS: Focal electrical stimulation of mouse colon produced phased-locked calcium signals in myenteric neurons and produced colon contractions. Stimulation of the L6 ventral root, which contains PPN axons, also produced myenteric activation and contractions that were comparable to those of direct colon stimulation. Surprisingly, capsaicin application to the isolated L6 dorsal root ganglia, which produced robust calcium signals in neurons throughout the ganglion, did not activate myenteric neurons. Electrical activation of the ganglia, which activated even more neurons than capsaicin, did not produce myenteric activation or contractions unless the spinal cord was intact, indicating that a complete afferent-to-efferent (PPN) circuit was necessary for ExPANs to regulate myenteric neurons. In TRPV1-channel rhodopsin-2 mice, light activation of ExPANs induced a pain-like visceromotor response and expression of Fos in spinal PPN neurons. CONCLUSIONS: In mice, ExPANs regulate myenteric neuron activity and smooth muscle contraction via a parasympathetic spinal circuit, linking sensation and pain to motility.

Bile acids induce visceral hypersensitivity via mucosal mast cell-to-nociceptor signaling that involves the farnesoid X receptor/nerve growth factor/transient receptor potential vanilloid 1 axis.

Increased colonic bile acid (BA) exposure, frequent in diarrhea-predominant irritable bowel syndrome (IBS-D), can affect gut function. Nerve growth factor (NGF) is implicated in the development of visceral hypersensitivity (VH). In this study, we tested the hypothesis that BAs cause VH via mucosal mast cell (MMC)-to-nociceptor signaling, which involves the farnesoid X receptor (FXR)/NGF/transient receptor potential vanilloid (TRPV)1 axis. BAs were intracolonically administered to rats for 15 d. Visceral sensitivity to colorectal distention and colonic NGF expression were examined. BAs caused VH, an effect that involved MMC-derived NGF and was accompanied by enhanced TRPV1 expression in the dorsal root ganglia. Anti-NGF treatment and TRPV1 antagonism inhibited BA-induced VH. BAs induced NGF mRNA and protein expression and release in cultured mast cells. Colonic supernatants from patients with IBS-D with elevated colonic BA content transcriptionally induced NGF expression. In FXR-/- mice, visceral sensitivity and colonic NGF expression were unaltered after BA treatment. Pharmacological antagonism and FXR silencing suppressed BA-induced NGF expression and release in mast cells. Mitogen-activated protein kinase kinase (MKK) 3/6/p38 MAPK/NF-κB signaling was mechanistically responsible for FXR-mediated NGF expression and secretion. The findings show an MMC-dependent and FXR-mediated pronociceptive effect of BAs and identify the BA/FXR/NGF/TRPV1 axis as a key player in MMC-to-neuron communication during pain processing in IBS.-Li, W.-T., Luo, Q.-Q., Wang, B., Chen, X., Yan, X.-J., Qiu, H.-Y., Chen, S.-L. Bile acids induce visceral hypersensitivity via mucosal mast cell-to-nociceptor signaling that involves the farnesoid X receptor/nerve growth factor/transient receptor potential vanilloid 1 axis.

MiR-34a affects dexmedetomidine-inhibited chronic inflammatory visceral pain by targeting to HDAC2.

BACKGROUND: Dexmedetomidine (DEX) has been used as an anesthetic for decades. The present investigation aimed to elucidate the analgesic impact of DEX on 2,4,6-Trinitrobenzenesulfonic acid (TNBS)-induced chronic inflammatory visceral pain (CIVP) in rats. METHODS: TNBS with or without DEX to Male Sprague-Dawley SD rats were randomly divided into four groups: normal, CIVP, DEX, and vehicle. Pain behaviors were assessed and the abdominal withdrawal reflex, mechanical withdrawal threshold, and thermal withdrawal latency were recorded. Quantitative polymerase chain reaction data showed increased expressions of pro-inflammatory cytokines (IL-6, IL-1ß and TNF-α) in the spinal cord tissues of rats. RESULTS: RNA microarray and quantitative polymerase chain reaction results indicated that miR-34a was downregulated by TNBS induction, but it was upregulated by DEX administration. Further studies showed that transfection of adenovirus-miR-34a inhibitor reversed the effect of DEX on the pain behaviors and spinal-cord pro-inflammatory-cytokine generation in CIVP rats. Additionally, we found that miR-34a targeted the 3'-UTR of the HDAC2 gene, as evinced by the increased HDAC2 expression in the CIVP and DEX + miR-34a inhibitor groups, and decreased HDAC2 signaling in the DEX group. Moreover, knock-down of HDAC2 restored DEX-attenuated pain behaviors and reduced pro-inflammatory cytokine production. CONCLUSIONS: DEX thus exhibited an analgesic effect on CIVP rats through the miR-34a-mediated HDAC2 pathway and suppressed visceral hypersensitivity.

CGRPα within the Trpv1-Cre population contributes to visceral nociception.

The role of calcitonin gene-related peptide (CGRP) in visceral and somatic nociception is incompletely understood. CGRPα is highly expressed in sensory neurons of dorsal root ganglia and particularly in neurons that also express the transient receptor potential cation channel subfamily V member 1 (Trpv1). Therefore, we investigated changes in visceral and somatic nociception following deletion of CGRPα from the Trpv1-Cre population using the Cre/lox system. In control mice, acetic acid injection (0.6%, ip) caused significant immobility (time stationary), an established indicator of visceral pain. In CGRPα-mCherrylx/lx;Trpv1-Cre mice, the duration of immobility was significantly less than controls, and the distance CGRPα-mCherrylx/lx;Trpv1-Cre mice traveled over 20 min following acetic acid was significantly greater than controls. However, following acetic acid injection, there was no difference between genotypes in the writhing reflex, number of abdominal licks, or forepaw wipes of the cheek. CGRPα-mCherrylx/lx;Trpv1-Cre mice developed more pronounced inflammation-induced heat hypersensitivity above baseline values compared with controls. However, analyses of noxious acute heat or cold transmission revealed no difference between genotypes. Also, odor avoidance test, odor preference test, and buried food test for olfaction revealed no differences between genotypes. Our findings suggest that CGRPα-mediated transmission within the Trpv1-Cre population plays a significant role in visceral nociceptive pathways underlying voluntary movement. Monitoring changes in movement over time is a sensitive parameter to identify differences in visceral nociception, compared with writhing reflexes, abdominal licks, or forepaw wipes of the cheek that were unaffected by deletion of CGRPα- from Trpv1-Cre population and likely utilize different mechanisms. NEW & NOTEWORTHY The neuropeptide calcitonin gene-related peptide (CGRP) is highly colocalized with transient receptor potential cation channel subfamily V member 1 (TRPV1)-expressing primary afferent neurons, but the functional role of CGRPα specifically in these neurons is unknown in pain processing from visceral and somatic afferents. We used cre-lox recombination to conditionally delete CGRPα from TRPV1-expressing neurons in mice. We show that CGRPα from within TRPV1-cre population plays an important role in visceral nociception but less so in somatic nociception.

Assessment of the antinociceptive and ulcerogenic activity of the tapentadol-diclofenac combination in rodents.

Preclinical Research & Development The objective of the present study was to evaluate the tapentadol-diclofenac combination in three dose-ratios in the mouse acetic acid-induced visceral pain and their ulcerogenic activity on the stomachal mucous. Dose-response curves were generated for tapentadol, diclofenac, and their combination in the acetic acid-induced writhing test in mice. Moreover, the stomachs of animals were surgically removal and gastrointestinal ulcerogenic action of the combination was assessed. The isobolographic analysis, interaction index, and ANOVA were used to analyze the data. The isobolographic analysis and interaction index showed a similar antinociceptive activity for the three combinations of the analgesic mixture. Moreover, tapentadol and the proportions 1:1 or 3:1 of the analgesic combination caused a mild gastrointestinal damage. These data indicate that the systemic co-administration of tapentadol and diclofenac produced a synergistic interaction in the acetic acid-induced visceral pain test with an acceptable gastric damage profile in mice.

Visceral and somatic pain modalities reveal NaV 1.7-independent visceral nociceptive pathways.

KEY POINTS: Voltage-gated sodium channels play a fundamental role in determining neuronal excitability. Specifically, voltage-gated sodium channel subtype NaV 1.7 is required for sensing acute and inflammatory somatic pain in mice and humans but its significance in pain originating from the viscera is unknown. Using comparative behavioural models evoking somatic and visceral pain pathways, we identify the requirement for NaV 1.7 in regulating somatic (noxious heat pain threshold) but not in visceral pain signalling. These results enable us to better understand the mechanisms underlying the transduction of noxious stimuli from the viscera, suggest that the investigation of pain pathways should be undertaken in a modality-specific manner and help to direct drug discovery efforts towards novel visceral analgesics. ABSTRACT: Voltage-gated sodium channel NaV 1.7 is required for acute and inflammatory pain in mice and humans but its significance for visceral pain is unknown. Here we examine the role of NaV 1.7 in visceral pain processing and the development of referred hyperalgesia using a conditional nociceptor-specific NaV 1.7 knockout mouse (NaV 1.7Nav1.8 ) and selective small-molecule NaV 1.7 antagonist PF-5198007. NaV 1.7Nav1.8 mice showed normal nociceptive behaviours in response to intracolonic application of either capsaicin or mustard oil, stimuli known to evoke sustained nociceptor activity and sensitization following tissue damage, respectively. Normal responses following induction of cystitis by cyclophosphamide were also observed in both NaV 1.7Nav1.8 and littermate controls. Loss, or blockade, of NaV 1.7 did not affect afferent responses to noxious mechanical and chemical stimuli in nerve-gut preparations in mouse, or following antagonism of NaV 1.7 in resected human appendix stimulated by noxious distending pressures. However, expression analysis of voltage-gated sodium channel α subunits revealed NaV 1.7 mRNA transcripts in nearly all retrogradely labelled colonic neurons, suggesting redundancy in function. By contrast, using comparative somatic behavioural models we identify that genetic deletion of NaV 1.7 (in NaV 1.8-expressing neurons) regulates noxious heat pain threshold and that this can be recapitulated by the selective NaV 1.7 antagonist PF-5198007. Our data demonstrate that NaV 1.7 (in NaV 1.8-expressing neurons) contributes to defined pain pathways in a modality-dependent manner, modulating somatic noxious heat pain, but is not required for visceral pain processing, and advocate that pharmacological block of NaV 1.7 alone in the viscera may be insufficient in targeting chronic visceral pain.

Modulation of P2X3 and P2X2/3 Receptors by Monoclonal Antibodies.

Purinergic homomeric P2X3 and heteromeric P2X2/3 receptors are ligand-gated cation channels activated by ATP. Both receptors are predominantly expressed in nociceptive sensory neurons, and an increase in extracellular ATP concentration under pathological conditions, such as tissue damage or visceral distension, induces channel opening, membrane depolarization, and initiation of pain signaling. Hence, these receptors are considered important therapeutic targets for pain management, and development of selective antagonists is currently progressing. To advance the search for novel analgesics, we have generated a panel of monoclonal antibodies directed against human P2X3 (hP2X3). We have found that these antibodies produce distinct functional effects, depending on the homomeric or heteromeric composition of the target, its kinetic state, and the duration of antibody exposure. The most potent antibody, 12D4, showed an estimated IC50 of 16 nm on hP2X3 after short term exposure (up to 18 min), binding to the inactivated state of the channel to inhibit activity. By contrast, with the same short term application, 12D4 potentiated the slow inactivating current mediated by the heteromeric hP2X2/3 channel. Extending the duration of exposure to ∼20 h resulted in a profound inhibition of both homomeric hP2X3 and heteromeric hP2X2/3 receptors, an effect mediated by efficient antibody-induced internalization of the channel from the plasma membrane. The therapeutic potential of mAb12D4 was assessed in the formalin, complete Freund's adjuvant, and visceral pain models. The efficacy of 12D4 in the visceral hypersensitivity model indicates that antibodies against P2X3 may have therapeutic potential in visceral pain indications.

Synergism between Naproxen and Rutin in a Mouse Model of Visceral Pain.

Preclinical Research The aim of the present study was to evaluate the antinociceptive interaction between naproxen and the glycoside flavonoid, rutin in the acetic acid-induced writhing test in mice. Naproxen (5, 20, 50, and 100 mg/kg p.o.) or rutin (10, 25, 50, and 100mg/kg p.o.) were administered 60 min before the intraperitoneal administration with acetic acid. The dose-response curve of each individual compound and the experimental effective dose 50 (ED50 ) value were obtained to determinate different proportions of the combinations between the two compounds (naproxen-rutin 1:1, 3:1, and 3:1) in the writhing test. The results indicated a synergistic antinociceptive interaction between two drugs with different mechanism of action, naproxen and rutin in all the combinations. Drug Dev Res 78 : 184-188, 2017. © 2017 Wiley Periodicals, Inc.

Elevated H2 O2 levels in trinitrobenzene sulfate-induced colitis rats contributes to visceral hyperalgesia through interaction with the transient receptor potential ankyrin 1 cation channel.

BACKGROUND AND AIM: Inflammatory bowel disease is associated with chronic abdominal pain. Transient receptor potential ankyrin 1 (TRPA1) is a well-known pain sensor expressed in primary sensory neurons. Recent studies indicate that reactive oxygen species such as hydrogen peroxide (H2 O2 ) may activate TRPA1. METHODS: Colonic inflammation was induced by intra-colonic administration of trinitrobenzene sulfate (TNBS) in adult male Sprague-Dawley rats. Visceromotor response (VMR) to colorectal distention (CRD) was recorded to evaluate the visceral hyperalgesia. Rats were sacrificed 1 day after treatment with saline or TNBS; colonic tissues from the inflamed region were removed and then processed to assess the H2 O2 content. H2 O2 scavenger N-acetyl-l-cysteine or a TRPA1 antagonist, HC-030031, was intravenously administrated to the TNBS-treated rats or saline-treated rats. In a parallel experiment, intra-colonic H2 O2 -induced visceral hyperalgesia in naïve rats and the effect of intravenous HC-030031 were measured based on the VMR to CRD. RESULTS: Trinitrobenzene sulfate treatment resulted in significant increase in VMR to CRD at day 1. The H2 O2 content in the inflamed region of the colon in TNBS-treated rats was significantly higher than that of saline-treated rats. N-acetyl-l-cysteine or HC-030031 significantly suppressed the enhanced VMR in TNBS-treated rats while saline-treated rats remained unaffected. Moreover, blockade of TRPA1 activation by HC-030031 significantly reversed the exogenous H2 O2 -induced visceral hyperalgesia. CONCLUSION: These results suggest that H2 O2 content of the colonic tissue is increased in the early stage of TNBS-induced colitis. The increased H2 O2 content may contribute to the visceral hyperalgesia by activating TRPA1.

Ellagic acid enhances the antinociceptive action of carbamazepine in the acetic acid writhing test with mice.

CONTEXT: Ellagic acid (EA) produced antinociceptive and anti-inflammatory effects through the central and peripheral sites of action. OBJECTIVE: The objective of the current study was to examine the functional interaction between ellagic acid and carbamazepine (CBZ) on pain. MATERIALS AND METHODS: Fourteen groups of mice (8-10 each) were used in this study. Pain was induced by intraperitoneal acetic acid in mice (writhing test) and the functional interaction was analyzed using the isobolographic method. EA at doses 0.3, 1, 3, and 10 mg/kg and carbamazepine at doses 3, 10, 20, and 30 mg/kg, alone and also in combination (1/2, 1/4, and 1/8 of the drug's ED50) were intraperitoneally administered 30 min before acetic acid (0.6% v/v). Then, the abdominal writhes were counted during a 25-min period. RESULTS: EA (0.3-10 mg/kg, i.p.) and CBZ (3-30 mg/kg, i.p.) inhibited the writhing response evoked by acetic acid. Fifty percent effective dose (ED50) values against this tonic pain were 1.02 mg/kg and 6.40 mg/kg for EA and CBZ, respectively. The antinociception induced by EA showed higher potency than that of carbamazepine. Co-administration of increasing fractional increments of ED50 values of EA and CBZ produced additive interaction against writhing responses, as revealed by isobolographic analysis. DISCUSSION AND CONCLUSION: These results suggest that a combination of carbamazepine and ellagic acid may be a new strategy for the management of neuropathic pain such as what occurs in trigeminal neuralgia, since the use of carbamazepine is often limited by its adverse effects and by reduction of its analgesic effect through microsomal enzyme induction.

The analgesic effect of diclofenac sodium administered via the epidural route in an experimental visceral pain model.

PURPOSE: The aim of this study was to investigate the characteristics of the analgesic effect of diclofenac sodium injected epidurally in single or repeated doses and whether tolerance develops in long-term use. MATERIALS AND METHODS: A total of 30 rats were included in the study. The rats were anesthetized using intraperitoneal ketamine hydrochloride and an epidural catheter (EC) was inserted at the level of 13th dorsal thoraco-lumbar vertebrae (T13). Eleven rats were excluded from the study. The remaining 19 rats were randomly divided into three groups; Group Control (Group C) (n = 6) received 20 µL normal saline solution (NS) via EC for 10 days; Group Single Dose (Group SD) (n = 6) received 20 µL NS for 9 days and 6 µg diclofenac via EC on 10th day; Group Ten Doses (Group TDs) (n = 7) received 6 µg diclofenac via EC in 20 µL NS for 10 days. On the 10th day, 30 min after epidural diclofenac sodium, 300 mg/kg of 3% acetic acid was injected via intraperitoneal route, and the rats were observed for 30 min and number of writhing reflex (WR) was recorded. RESULTS: The values of total number of Writhing Reflex (WRT) and Writhing reflex per minute(WR/min) were found to be significantly higher in Group C compared with Groups SD and TD (P = 0.009). CONCLUSION: Single and repeated doses of diclofenac sodium via epidural route have an analgesic effect in a visceral pain model in rats without developing tolerance.

TRPV1 sensitization mediates postinflammatory visceral pain following acute colitis.

Quiescent phases of inflammatory bowel disease (IBD) are often accompanied by chronic abdominal pain. Although the transient receptor potential vanilloid 1 (TRPV1) ion channel has been postulated as an important mediator of visceral hypersensitivity, its functional role in postinflammatory pain remains elusive. This study aimed at establishing the role of TRPV1 in the peripheral sensitization underlying chronic visceral pain in the context of colitis. Wild-type and TRPV1-deficient mice were separated into three groups (control, acute colitis, and recovery), and experimental colitis was induced by oral administration of dextran sulfate sodium (DSS). Recovery mice showed increased chemically and mechanically evoked visceral hypersensitivity 5 wk post-DSS discontinuation, at which point inflammation had completely resolved. Significant changes in nonevoked pain-related behaviors could also be observed in these animals, indicative of persistent discomfort. These behavioral changes correlated with elevated colonic levels of substance P (SP) and TRPV1 in recovery mice, thus leading to the hypothesis that SP could sensitize TRPV1 function. In vitro experiments revealed that prolonged exposure to SP could indeed sensitize capsaicin-evoked currents in both cultured neurons and TRPV1-transfected human embryonic kidney (HEK) cells, a mechanism that involved TRPV1 ubiquitination and subsequent accumulation at the plasma membrane. Importantly, although TRPV1-deficient animals experienced similar disease severity and pain as wild-type mice in the acute phase of colitis, TRPV1 deletion prevented the development of postinflammatory visceral hypersensitivity and pain-associated behaviors. Collectively, our results suggest that chronic exposure of colon-innervating primary afferents to SP could sensitize TRPV1 and thus participate in the establishment of persistent abdominal pain following acute inflammation.

Butyrate-induced colonic hypersensitivity is mediated by mitogen-activated protein kinase activation in rat dorsal root ganglia.

OBJECTIVE: Increased faecal butyrate levels have been reported in irritable bowel syndrome. Rectal instillation of sodium butyrate (NaB) increases visceral sensitivity in rats by an unknown mechanism. We seek to examine the signal transduction pathways responsible for the enhanced neuronal excitability in the dorsal root ganglion (DRG) following NaB enemas and demonstrate that this is responsible for the colonic hypersensitivity reported in this animal model. DESIGN: Colorectal distention (CRD) studies were performed in rats treated with NaB rectal instillation with/without intrathecal or intravenous administration of mitogen-activated protein (MAP) kinase kinase inhibitor U0126. Western blot analysis and immunocytochemistry studies elucidated intracellular signalling pathways that modulate IA. Patch-clamp recordings were performed on isolated DRG neurons treated with NaB, with/without U0126. RESULTS: Visceromotor responses (VMR) were markedly enhanced in NaB-treated rats. Western blot analysis of DRG neurons from NaB-treated rats showed a 2.2-fold increase in phosphorylated ERK1/2 (pEKR1/2) and 1.9-fold increase in phosphorylated voltage-gated potassium channel subunit 4.2 (pKv4.2). Intrathecal or intravenous administration of U0126 reduced VMR to CRD in NaB-treated rats and prevented increases in pERK1/2 and pKv4.2. Patch-clamp recordings of isolated DRG neurons showed that NaB caused a reduction in IA to 48.9%±1.4% of control and an increase in neuronal excitability, accompanied by a twofold increase in pERK1/2 and pKv4.2. Concurrent U0126 administration prevented these changes. CONCLUSIONS: Visceral hypersensitivity induced by colonic NaB treatment is mediated by activation of the MAP kinase-ERK1/2 pathway, which phosphorylates Kv4.2. This results in a reduction in IA and an enhancement of DRG neuronal excitability.

Effectiveness of Butorphanol in alleviating intra- and post-operative visceral pain following microwave ablation for hepatic tumor: a dual-central, randomized, controlled trial.

Many patients who underwent hepatic percutaneous microwave ablation (MWA) reported experiencing pain during the procedure. This study utilized a well-designed multicentral, randomized, and placebo-controlled format to investigate the effects of Butorphanol. Patients who underwent MWA were randomly assigned to either Butorphanol or normal saline group. The primary outcomes of the study were assessed by measuring the patients' intraoperative pain levels using a 10-point visual analog scale (VAS). Secondary outcomes included measuring postoperative pain levels at the 6-h mark (VAS) and evaluating comprehensive pain assessment outcomes. A total of 300 patients were divided between the control group (n = 100) and the experimental group (n = 200). Butorphanol showed statistically significant reductions in intraoperative pain levels compared to the placebo during surgery (5.00 ± 1.46 vs. 3.54 ± 1.67, P < 0.001). Significant differences were observed in postoperative pain levels at the 6-h mark and in the overall assessment of pain (1.39 + 1.21 vs. 0.65 + 0.81, P < 0.001). Butorphanol had a significant impact on reducing the heart rate of patients. The empirical evidence supports the effectiveness of Butorphanol in reducing the occurrence of visceral postoperative pain in patients undergoing microwave ablation for hepatic tumor. Furthermore, the study found no noticeable impact on circulatory and respiratory dynamics.

σ1 receptors are involved in the visceral pain induced by intracolonic administration of capsaicin in mice.

BACKGROUND: Visceral pain is an important and prevalent clinical condition whose treatment is challenging. Sigma-1 (σ1) receptors modulate somatic pain, but their involvement in pure visceral pain is unexplored. METHODS: The authors evaluated the role of σ1 receptors in intracolonic capsaicin-induced visceral pain (pain-related behaviors and referred mechanical hyperalgesia to the abdominal wall) using wild-type (WT) (n = 12 per group) and σ1 receptor knockout (σ1-KO) (n = 10 per group) mice, selective σ1 receptor antagonists (BD-1063, S1RA, and NE-100), and control drugs (morphine and ketoprofen). RESULTS: The intracolonic administration of capsaicin (0.01-1%) induced concentration-dependent visceral pain-related behaviors and referred hyperalgesia in both WT and σ1-KO mice. However, the maximum number of pain-related behaviors induced by 1% capsaicin in σ1-KO mice (mean ± SEM, 22 ± 2.9) was 48% of that observed in WT animals (46 ± 4.2). Subcutaneous administration of the σ1 receptor antagonists BD-1063 (16-64 mg/kg), S1RA (32-128 mg/kg), and NE-100 (8-64 mg/kg) dose-dependently reduced the number of behavioral responses (by 53, 62, and 58%, respectively) and reversed the referred hyperalgesia to mechanical control threshold (0.53 ± 0.05 g) in WT mice. In contrast, these drugs produced no change in σ1-KO mice. Thus, the effects of these drugs are specifically mediated by σ1 receptors. Morphine produced an inhibition of capsaicin-induced visceral pain in WT and σ1-KO mice, whereas ketoprofen had no effect in either mouse type. CONCLUSION: These results suggest that σ1 receptors play a role in the mechanisms underlying capsaicin-induced visceral pain and raise novel perspectives for their potential therapeutic value.