Colorectal Distension In Rats Research Articles

Intracerebroventricular administration of TRH Agonist, RX-77368 alleviates visceral pain induced by colorectal distension in rats

Thyrotropin-releasing hormone (TRH) acts centrally to exert pleiotropic actions independently from its endocrine function, including antinociceptive effects against somatic pain in rodents. Whether exogenous or endogenous activation of TRH signaling in the brain modulates visceral pain is unknown. Adult male Sprague-Dawley rats received an intracerebroventricular (ICV) injection of the stable TRH analog, RX-77368 (10, 30 and 100 ng/rat) or saline (5 µl) or were semi-restrained and exposed to cold (4°C) for 45 min. The visceromotor response (VMR) to graded phasic colorectal distensions (CRD) was monitored using non-invasive intracolonic pressure manometry. Naloxone (1 mg/kg) was injected subcutaneously 10 min before ICV RX-77368 or saline. Fecal pellet output was monitored for 1 h after ICV injection. RX-77368 ICV (10, 30 and 100 ng/rat) reduced significantly the VMR by 56.7%, 67.1% and 81.1% at 40 mmHg and by 30.3%, 58.9% and 87.4% at 60 mmHg respectively vs ICV saline. Naloxone reduced RX-77368 (30 and 100 ng, ICV) analgesic response by 51% and 28% at 40 mmHg and by 30% and 33% at 60 mmHg respectively, but had no effect per se. The visceral analgesia was mimicked by the acute exposure to cold. At the doses of 30 and 100 ng, ICV RX-77368 induced defecation within 30 min. These data established the antinociceptive action of RX-77368 injected ICV in a model of visceral pain induced by colonic distension through recruitment of both opioid and non-opioid dependent mechanisms.

Microglia-derived TNF-α inhibiting GABAergic neurons in the anterior lateral bed nucleus of the stria terminalis precipitates visceral hypersensitivity induced by colorectal distension in rats

Irritable bowel syndrome (IBS) is a common and debilitating gastrointestinal disorder that is exacerbated by stress and characterized by abdominal pain. Although microglia in the CNS have been implicated as an important mediator of the stress response, the role of microglia and microglia-GABAergic neuron interactions in the limbic area, most notably BNST, in the development of colorectal hypersensitivity has not been determined. We established a neonatal colorectal distension-induced chronic visceral hyperalgesia model in rats. The results showed that the frequency of spontaneous discharges of alBNST GABAergic neurons and the expression of GAD65/67 were significantly decreased in rats with chronic visceral pain. Moreover, ablation of BNST GABAergic neurons significantly reduced the visceral pain threshold in normal rats. Meanwhile, the number of M1 proinflammatory microglia and the expression of the M1 proinflammatory microglia-derived cytokines IL-6 and TNF-α were increased in the alBNST of rats with chronic visceral pain. Furthermore, alBNST infusion of the microglial inhibitor minocycline or IL-6 and TNF-α neutralizing antibodies significantly increased the visceral pain threshold. The decreased frequency of spontaneous discharges of alBNST GABAergic neurons in rats with chronic visceral pain was mimicked by a bath perfusion of TNF-α, but not IL-6, and was abolished by a perfusion of the microglial inhibitor minocycline. In addition, the alBNST infusion of the microglial inhibitor minocycline upregulated the expression of GAD65/67. Moreover, ablation of BNST GABAergic neurons significantly decreased the visceral pain threshold in normal rats, which was not reversed by a subsequent infusion of the microglial inhibitor minocycline. Our findings revealed this microglia-GABAergic neuron circuit in the alBNST, and this microglia-driven disinhibitory mechanism is essential for brain and gut dysfunction in stressful condition, providing a novel potential target for treating patients with IBS presenting visceral pain that is worsened during episodes of stress.

A Monoclonal Anti-Calcitonin Gene-Related Peptide Antibody Decreases Stress-Induced Colonic Hypersensitivity.

Irritable bowel syndrome (IBS) is a brain-gut disorder characterized by abdominal pain and altered bowel habits. Although the etiology of IBS remains unclear, stress in adulthood or in early life has been shown to be a significant factor in the development of IBS symptomatology. Evidence suggests that aberrant calcitonin gene-related peptide (CGRP) signaling may be involved in afferent sensitization and visceral organ hypersensitivity. Here, we used a monoclonal anti-CGRP divalent antigen-binding fragment [F(ab')2] antibody to test the hypothesis that inhibition of peripheral CGRP signaling reverses colonic hypersensitivity induced by either chronic adult stress or early life stress. A cohort of adult male rats was exposed to repeated water avoidance stress. Additionally, a second cohort consisting of female rats was exposed to a female-specific neonatal odor-attachment learning paradigm of unpredictable early life stress. Colonic sensitivity was then assessed in adult animals via behavioral responses to colorectal distension (CRD). To analyze spinal nociceptive signaling in response to CRD, dorsal horn extracellular signal-regulated kinase (ERK) 1/2 phosphorylation was measured via immunohistochemistry. Repeated psychologic stress in adulthood or unpredictable stress in early life induced colonic hypersensitivity and enhanced evoked ERK1/2 phosphorylation in the spinal cord after CRD in rats. These phenotypes were reversed by administration of a monoclonal anti-CGRP F(ab')2 fragment antibody. Stress-induced changes in visceral sensitivity and spinal nociceptive signaling were reversed by inhibition of peripheral CGRP signaling, which suggests a prominent role for CGRP in central sensitization and the development of stress-induced visceral hypersensitivity. SIGNIFICANCE STATEMENT: Targeting peripheral calcitonin gene-related peptide (CGRP) with a monoclonal anti-CGRP divalent antigen-binding fragment antibody reduced central sensitization and attenuated colonic hypersensitivity induced by either chronic adult stress or early life stress. CGRP-targeting antibodies are approved for migraine prevention, and the results of this study suggest that targeting CGRP may provide a novel treatment strategy for irritable bowel syndrome-related, stress-induced visceral pain.

Small-Conductance Ca2+-Activated K+ Channels 2 in the Hypothalamic Paraventricular Nucleus Precipitates Visceral Hypersensitivity Induced by Neonatal Colorectal Distension in Rats.

Visceral hypersensitivity is one of the pivotal pathophysiological features of visceral pain in irritable bowel syndrome (IBS). Small-conductance Ca2+-activated K+ channel (SK) is critical for a variety of functions in the central nervous system (CNS), nonetheless, whether it is involved in the pathogenesis of visceral hypersensitivity remain elusive. In this study, we examined mechanism of SK2 in hypothalamic paraventricular nucleus (PVN) in the pathogenesis of visceral hypersensitivity induced by neonatal colorectal distension (CRD). Rats undergoing neonatal CRD presented with visceral hypersensitivity as well as downregulated membrane SK2 channel and p-PKA. Intra-PVN administration of either the membrane protein transport inhibitor dynasore or the SK2 activator 1-EBIO upregulated the expression of membrane SK2 in PVN and mitigated visceral hypersensitivity. In addition, 1-EBIO administration reversed the increase in neuronal firing rates in PVN in rats undergoing neonatal CRD. On the contrary, intra-PVN administration of either the SK2 inhibitor apamin or PKA activator 8-Br-cAMP exacerbated the visceral hypersensitivity. Taken together, these findings demonstrated that visceral hypersensitivity is related to the downregulation of membrane SK2 in PVN, which may be attributed to the activation of PKA; pharmacologic activation of SK2 alleviated visceral hypersensitivity, which brings prospect of SK2 activators as a new intervention for visceral pain.

Aluminum Ingestion Promotes Colorectal Hypersensitivity in Rodents.

Background & AimsIrritable bowel syndrome (IBS) is a multifactorial disease arising from a complex interplay between genetic predisposition and environmental influences. To date, environmental triggers are not well known. Aluminum is commonly present in food, notably by its use as food additive. We investigated the effects of aluminum ingestion in rodent models of visceral hypersensitivity, and the mechanisms involved.MethodsVisceral hypersensitivity was recorded by colorectal distension in rats administered with oral low doses of aluminum. Inflammation was analyzed in the colon of aluminum-treated rats by quantitative PCR for cytokine expression and by immunohistochemistry for immune cells quantification. Involvement of mast cells in the aluminum-induced hypersensitivity was determined by cromoglycate administration of rats and in mast cell-deficient mice (KitW-sh/W-sh). Proteinase-activated receptor-2 (PAR2) activation in response to aluminum was evaluated and its implication in aluminum-induced hypersensitivity was assessed in PAR2 knockout mice.ResultsOrally administered low-dose aluminum induced visceral hypersensitivity in rats and mice. Visceral pain induced by aluminum persisted over time even after cessation of treatment, reappeared and was amplified when treatment resumed. As observed in humans, female animals were more sensitive than males. Major mediators of nociception were up-regulated in the colon by aluminum. Activation of mast cells and PAR2 were required for aluminum-induced hypersensitivity.ConclusionsThese findings indicate that oral exposure to aluminum at human dietary level reproduces clinical and molecular features of IBS, highlighting a new pathway of prevention and treatment of visceral pain in some susceptible patients.

Transient Hypertension after Spinal Cord Injury Leads to Cerebrovascular Endothelial Dysfunction and Fibrosis.

We aimed to create a clinically relevant pre-clinical model of transient hypertension, and then evaluate the pathophysiological cerebrovascular processes resulting from this novel stimulus, which has recently been epidemiologically linked to cerebrovascular disease. We first developed a clinically relevant model of transient hypertension, secondary to induced autonomic dysreflexia after spinal cord injury and demonstrated that in both patients and rats, this stimulus leads to drastic acute cerebral hyperperfusion. For this, iatrogenic urodynamic filling/penile vibrostimulation was completed while measuring beat-by-beat blood pressure and cerebral blood flow (CBF) in patients. We then developed a rodent model mimicking the clinical reality by performing colorectal distention (to induce autonomic dysreflexia) using pre-clinical beat-by-beat blood pressure and CBF assessments. We then performed colorectal distension in rats for four weeks (6x/day) to evaluate the long-term cerebrovascular consequences of transient hypertension. Outcome measures included middle cerebral artery endothelial function, remodeling, profibrosis and perivascular innervation; measured via pressure myography, immunohistochemistry, molecular biology, and magnetic resonance imaging. Our model demonstrates that chronic repetitive cerebral hyperperfusion secondary to transient hypertension because of autonomic dysreflexia: (1) impairs cerebrovascular endothelial function; (2) leads to profibrotic cerebrovascular stiffening characterized by reduced distensibility and increased collagen deposition; and (3) reduces perivascular sympathetic cerebrovascular innervation. These changes did not occur concurrent to hallmark cerebrovascular changes from chronic steady-state hypertension, such as hypertrophic inward remodeling, or reduced CBF. Chronic exposure to repetitive transient hypertension after spinal cord injury leads to diverse cerebrovascular impairment that appears to be unique pathophysiology compared with steady-state hypertension in non-spinal cord injured models.

α-lipoic acid suppresses neuronal excitability and attenuates colonic hypersensitivity to colorectal distention in diabetic rats.

AimPatients with long-standing diabetes often demonstrate intestinal dysfunction, characterized as constipation or colonic hypersensitivity. Our previous studies have demonstrated the roles of voltage-gated sodium channels NaV1.7 and NaV1.8 in dorsal root ganglion (DRG) in colonic hypersensitivity of rats with diabetes. This study was designed to determine roles of antioxidant α-lipoic acid (ALA) on sodium channel activities and colonic hypersensitivity of rats with diabetes.MethodsStreptozotocin was used to induce diabetes in adult female rats. Colonic sensitivity was measured by behavioral responses to colorectal distention in rats. The excitability and sodium channel currents of colon projection DRG neurons labeled with DiI were measured by whole-cell patch-clamp recordings. The expressions of NaV1.7 and NaV1.8 of colon DRGs were measured by western blot analysis.ResultsALA treatment significantly increased distention threshold in responding to colorectal distension in diabetic rats compared with normal saline treatment. ALA treatment also hyper-polarized the resting membrane potentials, depolarized action potential threshold, increased rheobase, and decreased frequency of action potentials evoked by ramp current stimulation. Furthermore, ALA treatment also reduced neuronal sodium current densities of DRG neurons innervating the colon from rats with diabetes. In addition, ALA treatment significantly downregulated NaV1.7 and NaV1.8 expression in colon DRGs from rats with diabetes.ConclusionOur results suggest that ALA plays an analgesic role, which was likely mediated by downregulation of NaV1.7 and NaV1.8 expressions and functions, thus providing experimental evidence for using ALA to treat colonic hypersensitivity in patients with diabetic visceral pain.

Medulla Oblongata Mechanism of Inhibitory Effect of Thermal Stimulation to Nociceptive Colorectal Distention in Rats

Objective: To discuss mechanism of moxibustion (thermal stimulation) effect and best moxibustion stimulus parameter. Methods: Experiments were performed on 48 male Sprague-Dawley rats. Unit discharges from individual single neuron were recorded extracellularly with glass-microelectrode in Subnucleus Reticularis Dorsalis (SRD). Visceral-intrusive stimulation is done by colorectal distension. Thermal stimulation with different temperature (40°C, 42°C, 44°C, 46°C, 48°C, 50°C, 52°C) and different stimulus area (diameter of circle : 1.0 cm, 1.5 cm, 2.0 cm, 2.5 cm, 3.0 cm, 3.5 cm, 4.0cm) was applied around RN12 during nociceptive colorectal distension. Results: SRD neurons could be activated by visceral stimulation within noxious range. Under low temperature of stimulus, especially under 45°C of pain threshold to ordinary people, visceral nociceptive afferent facilitated thermal stimulus from the body surface. While after thermal stimulation reached a harmful degree, the thermal stimulus will inhibit visceral nociceptive afferent. Moreover, statistics show that the higher the temperature is, the smaller the size of stimulation area is needed, and they correlate with each other negatively. Conclusion: Visceral nociception could be inhibited by somatic thermal stimulation with specific parameter at medulla level. According to our finding, best thermal stimulation temperature is around 48°C and the best size of stimulation area is around 3.14-7.07cm2 (with 2.0-3.0cm diameter).

Activation of corticotropin-releasing factor neurons and microglia in paraventricular nucleus precipitates visceral hypersensitivity induced by colorectal distension in rats

Visceral hypersensitivity is a major contributor to irritable bowel syndrome and other disorders with visceral pain. Substantial evidence has established that glial activation and neuro-glial interaction play a key role in the establishment and maintenance of visceral hypersensitivity. We recently demonstrated that activation of spinal microglial toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear factor κB (NF-κB) signaling facilitated the development of visceral hypersensitivity in a rat model developed by neonatal and adult colorectal distensions (CRDs). Hypothalamic paraventricular nucleus (PVN) plays a pivotal role in the pathogenesis of chronic pain. In this study, we examined the mechanism by which microglia and neurons in PVN establish and maintain visceral hypersensitivity and the involvement of TLR4 signaling. Visceral hypersensitivity was precipitated by adult colorectal distension (CRD) only in rats that experienced neonatal CRDs. Visceral hypersensitivity was associated with an increase in the expression of c-fos, corticotropin-releasing factor (CRF) protein and mRNA in PVN, which could be prevented by intra-PVN infusion of lidocaine or small interfering RNA targeting the CRF gene. These results suggest PVN CRF neurons modulate visceral hypersensitivity. Adult CRD induced an increase in the expression of Iba-1 (a microglial marker), TLR4 protein, and its downstream effectors MyD88, NF-κB, as well as proinflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) only in rats that experienced neonatal CRDs. Intra-PVN infusion of minocycline, a nonselective microglial inhibitor, attenuated the hyperactivity of TLR4 signaling cascade, microglial activation, and visceral hypersensitivity. Taken together, these data suggest that neonatal CRDs induce a glial activation in PVN. Adult CRD potentiates the glial and CRF neuronal activity, and precipitates visceral hypersensitivity and pain. TLR4 signaling and proinflammatory cytokines TNF-α and IL-1β may participate in neuro-glial interaction during the pathogenesis of visceral hypersensitivity.

Hippocampal microglial activation and glucocorticoid receptor down-regulation precipitate visceral hypersensitivity induced by colorectal distension in rats

Visceral hypersensitivity is a common characteristic in patients suffering from irritable bowel syndrome (IBS) and other disorders with visceral pain. Although the pathogenesis of visceral hypersensitivity remains speculative due to the absence of pathological changes, the long-lasting sensitization in neuronal circuitry induced by early life stress may play a critical role beyond the digestive system even after complete resolution of the initiating event. The hippocampus integrates multiple sources of afferent inputs and sculpts integrated autonomic outputs for pain and analgesia regulation. Here, we examined the hippocampal mechanism in the pathogenesis of visceral hypersensitivity with a rat model induced by neonatal and adult colorectal distensions (CRDs). Neither neonatal nor adult CRD evoked behavioral abnormalities in adulthood; however, adult re-exposure to CRD induced persistent visceral hypersensitivity, depression-like behaviors, and spatial learning impairment in rats that experienced neonatal CRD. Rats that experienced neonatal and adult CRDs presented a decrease in hippocampal glucocorticoid receptor (GR) immunofluorescence staining and protein expression, and increases in hippocampal microglial activation and cytokine (IL-1β and TNF-α) accumulation. The decrease in hippocampal GR expression and increase in hippocampal IL-1β and TNF-α accumulation could be prevented by hippocampal local infusion of minocycline, a microglial inhibitor. These results suggest that neonatal CRD can increase the vulnerability of hippocampal microglia, and adult CRD challenge facilitates the hippocampal cytokine release from the sensitized microglia, which down-regulates hippocampal GR protein expression and, subsequently, precipitates visceral hypersensitivity.

5‐HT 3 Receptor Signaling in a Rat Model of Sex Specific Visceral Hypersensitivity

Background Irritable bowel syndrome (IBS) is a functional gastrointestinal motor and visceral sensation disorder that is more prominent in women compared to men. 5-Hydroxytryptamine (5-HT, serotonin) signaling is disrupted in some IBS patients possibly due to polymorphic variations in the gene encoding the serotonin transporter (SERT) which result in increased extracellular 5-HT availability. Female SERT knockout (KO) rats exhibit visceral hypersensitivity to colonic distention that mimics colonic hypersensitivity known to occur in female IBS patients. Objective. We tested the hypothesis that 5-HT action at 5-HT3 receptors contributes to visceral hypersensitivity. Methods. We examined the effects of acute administration of morphine and the 5-HT3 receptor antagonists alosetron, granisetron, and ondansetron on visceral sensitivity in SERT KO and wild type (WT) rats. We measured the visceromotor response (VMR) to colorectal distension (CRD). Results. Ondansetron (0.1 mg/kg, s.c) did not affect visceral sensitivity, while alosetron (0.1 mg/kg, s.c.) increased the VMR to CRD in SERT KO female rats and WT male rats. Granisetron (0.1 mg/kg, s.c.) increased VMR to CRD in SERT KO female rats only. Morphine (3 mg/kg s.c.) suppressed the VMR to CRD in all rats. Conclusions The results suggest that acute peripheral administration of some 5-HT3 receptor antagonists causes a paradoxical increase in the VMR to CRD. 5-HT3 receptor antagonists may interact with ascending pain pathways and descending pain inhibitory pathways. Supported by: 5R21NS075841

Colorectal distention induces acute and delayed visceral hypersensitivity: role of peripheral corticotropin-releasing factor and interleukin-1 in rats.

Most studies evaluating visceral sensation measure visceromotor response (VMR) to colorectal distention (CRD). However, CRD itself induces visceral sensitization, and little is known about the detailed characteristics of this response. The present study tried to clarify this question. VMR was determined by measuring abdominal muscle contractions as a response to CRD in rats. The CRD set consisted of two isobaric distentions (60 mmHg for 10 min twice, with a 30-min rest), and the CRD set was performed on two separate days, i.e., days 1 and 3, 8. On day 1, VMR to the second CRD was increased as compared with that to the first CRD, which is the acute sensitization. VMR to the first CRD on day 3 returned to the same level as that to the first CRD on day 1, and total VMR, i.e., the whole response to the CRD set, was not different between day 1 and day 3. However, total VMR was significantly increased on day 8 as compared with that on day 1, suggesting CRD induced the delayed sensitization. Intraperitoneally administered astressin (200 µg/kg), a corticotropin-releasing factor receptor antagonist, at the end of the first CRD blocked the acute sensitization, but anakinra (20 mg/kg, intraperitoneally), an interleukin-1 receptor antagonist, did not modify it. Astressin (200 µg/kg, twice before CRD on day 8) did not alter the delayed sensitization, but anakinra (20 mg/kg, twice) abolished it. CRD induced both acute sensitization and delayed sensitization, which were mediated through peripheral corticotropin-releasing factor and interleukin-1 pathways, respectively.

The inhibitory effect of granisetron on ventrolateral medulla neuron responses to colorectal distension in rats

Irritable bowel syndrome (IBS) is one of the most widespread functional gastrointestinal disorders characterized by abdominal pain. A key pathophysiological mechanism of abdominal pain is associated with disturbances of serotonergic transmission in feedback control loops of endogenous pain modulation in which the ventrolateral medulla (VLM) plays an important role. The receptors to serotonin (5-HT), and particularly the serotonin 3 (5-HT3) receptors have been extensively used as a potential target for abdominal pain treatment of IBS patients due to antinociceptive features of the 5-HT3 receptor antagonists. The precise mechanisms underlying the antinociceptive action of these antagonists remain unclear. The main objective of our study was to evaluate the involvement of the 5-HT3 receptors in abdominal pain transmission within the VLM. Experiments were carried out on urethane-anaesthetized rats using the animal model of abdominal pain. Noxious colorectal distension (CRD) with a pressure of 80mmHg induced a significant increase in VLM neuron-evoked activity and depressor reactions (171.1±12.7% and 64±1.8% to baseline, accordingly). Selective blockade of the 5-HT3 receptors with granisetron at doses of 1.0 or 2.0mg/kg (i.v) resulted in long-lasting (90min) dose-dependent inhibition of VLM neuron-evoked activity and depressor reactions. When brainstem dorsal surface applications of granisetron (10 or 20µM) were used, the changes were more pronounced. These results suggest involvement of the 5-HT3 receptors in abdominal pain transmission within the VLM, which will be discussed in relation to the central antinociceptive effect of granisetron.

Effects of DA-9701, a Novel Prokinetic Agent, on Phosphorylated Extracellular Signal-Regulated Kinase Expression in the Dorsal Root Ganglion and Spinal Cord Induced by Colorectal Distension in Rat

Background/AimsDA-9701, a standardized extract of Pharbitis Semen and Corydalis Tuber, is a new prokinetic agent that exhibits an analgesic effect on the abdomen. We investigated whether DA-9701 affects visceral pain induced by colorectal distension (CRD) in rats.MethodsA total of 21 rats were divided into three groups: group A (no CRD+no drug), group B (CRD+no drug), and group C (CRD+DA-9701). Expression of pain-related factors, substance P (SP), c-fos, and phosphorylated extracellular signal-regulated kinase (p-ERK) in the dorsal root ganglion (DRG) and spinal cord was determined by immunohistochemical staining and Western blotting.ResultsThe proportions of neurons in the DRG and spinal cord expressing SP, c-fos, and p-ERK were higher in group B than in group A. In the group C, the proportion of neurons in the DRG and spinal cord expressing p-ERK was lower than that in group B. Western blot results for p-ERK in the spinal cord indicated a higher level of expression in group B than in group A and a lower level of expression in group C than in group B.ConclusionsDA-9701 may decrease visceral pain via the downregulation of p-ERK in the DRG and spinal cord.

Gastrointestinal pain: Unraveling a novel endogenous pathway through uroguanylin/guanylate cyclase-C/cGMP activation

The natural hormone uroguanylin regulates intestinal fluid homeostasis and bowel function through activation of guanylate cyclase-C (GC-C), resulting in increased intracellular cyclic guanosine-3′,5′-monophosphate (cGMP). We report the effects of uroguanylin-mediated activation of the GC-C/cGMP pathway in vitro on extracellular cGMP transport and in vivo in rat models of inflammation- and stress-induced visceral hypersensitivity. In vitro exposure of intestinal Caco-2 cells to uroguanylin stimulated bidirectional, active extracellular transport of cGMP into luminal and basolateral spaces. cGMP transport was significantly and concentration dependently decreased by probenecid, an inhibitor of cGMP efflux pumps. In ex vivo Ussing chamber assays, uroguanylin stimulated cGMP secretion from the basolateral side of rat colonic epithelium into the submucosal space. In a rat model of trinitrobenzene sulfonic acid (TNBS)-induced visceral hypersensitivity, orally administered uroguanylin increased colonic thresholds required to elicit abdominal contractions in response to colorectal distension (CRD). Oral administration of cGMP mimicked the antihyperalgesic effects of uroguanylin, significantly decreasing TNBS- and restraint stress–induced visceromotor response to graded CRD in rats. The antihyperalgesic effects of cGMP were not associated with increased colonic spasmolytic activity, but were linked to significantly decreased firing rates of TNBS-sensitized colonic afferents in rats in response to mechanical stimuli. In conclusion, these data suggest that the continuous activation of the GC-C/cGMP pathway along the intestinal tract by the endogenous hormones guanylin and uroguanylin results in significant reduction of gastrointestinal pain. Extracellular cGMP produced on activation of GC-C is the primary mediator in this process via modulation of sensory afferent activity.

Mechanisms of Pelvic Organ Crosstalk: 1. Peripheral Modulation of Bladder Inhibition by Colorectal Distention in Rats

Bladder activity can be inhibited by afferent input from the colorectum (inhibitory rectovesical reflex). We evaluated the functional response of the rat bladder to nonnoxious and noxious colorectal distention, and investigated the mechanical and pharmacological peripheral modulation of this response. In 70 female Sprague-Dawley® rats we evaluated the effect of nonnoxious (20 mm Hg) and noxious (40 and 60 mm Hg) colorectal distention on the micturition volume threshold and on bladder activity in a filled bladder. We also studied the effect of rectal balloon size (1.5 vs 3.5 cm long), and rectal administration of 2% lidocaine jelly or 1 mM allyl isothiocyanate solution on the inhibitory rectovesical reflex. Colorectal distention at 60 mm Hg increased the micturition volume threshold (mean ± SE 0.640 ± 0.056 vs 0.448 ± 0.035 ml in controls, p <0.001). Bladder contraction frequency was significantly decreased by 40 and 60 mm Hg colorectal distention vs controls (mean 0.62 ± 0.06 and 0.33 ± 0.05 per minute, respectively, vs 0.77 ± 0.03, each p <0.001). These effects were reversible and pressure dependent (p <0.001), and more pronounced using a large rectal balloon (mean 40 vs 60 mm Hg colorectal distention 0.35 ± 0.12 vs 0.07 ± 0.04 per minute, p = 0.004). We noted no significant graded inhibition of bladder contraction amplitude or duration. The inhibitory rectovesical reflex was reversibly abolished by intrarectal lidocaine administration. Intrarectal allyl isothiocyanate administration significantly increased the effect of noxious colorectal distention on bladder contraction frequency. Only noxious levels of colorectal distention initiated the inhibitory rectovesical reflex. The effect increased with rectal balloon size and with intrarectal administration of allyl isothiocyanate. It was reversibly abolished by lidocaine. Results suggest that spinal interneurons are the mechanism behind the inhibitory rectovesical reflex.

Visceral pain readouts in experimental medicine

Visceral pain is studied at the level of the primary afferent fiber, spinal cord, subcortical, and cortical levels electrophysiologically and using brain imaging, which provides an objective measure of excitation at each level. However, correlation of these with actual perception of pain in conscious animal models has been problematic, and we rely on indirect measures in most preclinical research. The main method is electromyographic recording of abdominal muscle contractions in response to colorectal distension (CRD), which may reflect reflexes set up at several levels of the above pathway. Several experimental treatments for visceral pain have failed in clinical trials, possibly because of failure to translate from preclinical observations on CRD responses in animals to perception of spontaneous events in patients. Therefore, we need more objective outcomes. In this NGM issue, Hultin et al. show feasibility of routine recordings of cortical evoked electrical potentials (CEP) using implanted cranial electrodes in response to graded CRD in rats. CEP comprised three temporal components with latencies of approximately 20-50 ms, 90-180 ms, and 300 ms, which were reproducible and graded in intensity and latency with distension pressure. From this basic study it is clear that colorectal evoked potentials can be recorded reliably in awake rats and may serve as an objective marker for centrally projecting visceral sensory signals in rodents. It remains to be seen how these responses are affected by drugs under development for clinical management of visceral pain, and if there is improved translation.

Repeated Psychological Stress Induces an Immediate Naloxone-Independent Visceral Analgesia to Colorectal Distension in Rats

Repeated Psychological Stress Induces an Immediate Naloxone-Independent Visceral Analgesia to Colorectal Distension in Rats

Oral baclofen reduces visceral pain-related pseudo-affective responses to colorectal distension in rats: Relation between plasma exposure and efficacy

Objective. We previously showed that activation of GABAB receptors by intravenous baclofen reduces pseudo-affective responses to colorectal distension in rats. Here we evaluate the potential clinical significance of these observations. Material and Methods. Clinically relevant colorectal distension protocols were used to assess the effects of oral baclofen on visceromotor and autonomic cardiovascular responses in conscious rats. Plasma levels of baclofen were monitored to provide clinical relevance to the doses used. Conscious female Sprague–Dawley rats were subjected to repeated noxious colorectal distension (12 × 80 mmHg), ascending-phasic colorectal distension (10–80 mmHg, 10 mmHg increments) or ramp colorectal distension (10 min ramp at 8 mmHg/min). Visceromotor and cardiovascular responses (mean arterial blood pressure and heart rate) were monitored. Pain-related response thresholds were assessed using ascending-phasic and ramp colorectal distension. Results. Baclofen (1–10 μmol/kg, p.o.) reduced the visceromotor response to colorectal distension, reaching a 40% maximal inhibition (p < 0.05). The highest dose (10 μmol/kg, p.o.) also inhibited pain-related cardiovascular responses in telemetrized rats (50–55% reduction in colorectal distension-evoked hypertensive and tachycardic responses; p < 0.05). Similar thresholds for pain-related visceromotor responses were determined during ramp or ascending-phasic colorectal distension (34.1 ± 1.9 and 31.7 ± 3.2 mmHg, respectively). Baclofen (10 μmol/kg, p.o.) increased thresholds to 71.1 ± 3.7 and 77.5 ± 1.8 mmHg during ramp and ascending-phasic colorectal distension, respectively (p < 0.001). Plasma levels of baclofen were 3.3 ± 0.2 μmol/l at 90 min post-dosing, corresponding to the end of the colorectal distension procedure. Conclusions. Oral baclofen, at plasma levels similar to those reported safe and within a therapeutic range in humans, produced significant visceral anti-nociceptive effects in rats.

Luminal serotonin time-dependently modulates vagal afferent driven antinociception in response to colorectal distention in rats

Compelling evidence shows that vagal afferents mediate antinociception in response to visceral insults. Our recent findings implied that luminal serotonin (5-hydroxytryptamine, 5-HT) might mediate chronic food allergen sensitized visceral hyperalgesia, in which vagal afferents might be implicated. Here, to test this hypothesis, we investigated the effects of luminal infused 5-HT on visceral nociception and the involvement of vagal antinociceptive pathway. The vagus-intact or vagotomized rats were given acute intraluminally or intraperitoneally administered 5-HT, or chronic luminal infusion of 5-HT. The visceromotor response (VMR) to colorectal distension (CRD) was electrophysiologically recorded. Acute intraluminal infusion of 5-HT (10 or 100 nmol) significantly attenuated VMR to CRD, while systemic administered 5-HT at similar doses resulted in markedly augmented nociception. Pretreatment with luminal application of granisetron or lidocaine, or pharmacological depletion of endogenous 5-HT with injection of p-chlorophenylalanine, a 5-HT synthesis inhibitor, and subdiaphragmatic vagotomy or functional deafferentation with capsaicin abolished the effect of luminal (but not systemic) 5-HT. Chronic infusion of 5-HT (10 nmol d(-1) for 5 days) produced gradual augmentation of baseline VMR. And, the VMR to CRD after 5-HT infusion decreased on day 1 and 2, then gradually increased from day 3. Surgical vagotomy or daily preperfusion with granisetron canceled these time-dependent patterns. Luminal 5-HT time-dependently modulates vagal afferent driven antinociception. Acute infusion of 5-HT attenuates visceral nociception via activation of vagal afferent 5-HT type 3 receptors (5-HT(3)Rs)within intestinal mucosa; while chronic luminal 5-HT caused gradually developed visceral hyperalgesia, which may also involve vagal 5-HT(3)Rs.