Animal Models Of Visceral Pain Research Articles

Correlation between acupoint sensitization of body surface and pituitary adenylate cyclase activating polypeptide expression in myocardial ischemia mice

To explore the relationship between sensitization points of the body surface and the expression of pituitary adenylate cyclase activating polypeptides (PACAP) in myocardial ischemia (MI) mice, so as to reveal the underlying mechanisms of acupoint sensitization from the perspective of molecular biology. Male C57BL/6J mice were randomly divided into control and model groups (28 mice/group). The MI-induced visceral pain model was established by intraperitoneal injection of isoprenaline (ISO, 160 mg/kg). The mice of the control group received intraperitoneal injection of the same dose of normal saline. Six days after modeling, the Evans blue (EB) dye was injected into the tail vein of mice to observe the distribution and quantity of the plasma extravasated EB points at the body surface. Meanwhile, the mechanical pain threshold (MPT) was measured to evaluate the level of pain sensitivity in the activated area on their body surface and left forelimb and hindlimb, respectively. Hematoxylin-eosin (H.E.) staining was used to evaluate the morphologic and pathological changes of the heart tissue in the two groups. Then, the expressions of PACAP in the thoracic (T)1-T5 dorsal root ganglia (DRGs), spinal cord and skin in the dominant area of body surface were detected by Western blot and immunofluorescence staining, respectively. Compared with the control group, the heart tissue of the model group was hypertrophic and the myocardial tissue showed obvious inflammatory cell infiltration and fibrosis. In addition to these pathologic changes, the number of EB exudation points on the body surface was significantly increased (P<0.01), and was mainly distributed in the innervated region of T1-T5 segments of the spinal cord, and the MPT of these EB exudation points was lower than that of non-exudation points (P<0.01). At the same time, the MPTs of left forelimb and hindlimb were significantly decreased in the model group (P<0.001). More importantly, the level of protein and positive expression of PACAP were significantly higher in the model group than those in the control group, which was observed in the innervated body surface, spinal cord and its DRG neurons of T1-T5 segments (P<0.01, P<0.001, P<0.05). ISO injection resulted in histological lesions and cardiogenic referred pain on the body surface after the formation of MI in mice. The expression of PACAP in the body surface of the sensitization points, the corresponding T1-T5 segments of spinal cord and DRG neurons were significantly increased, which may partly explain the reason for acupoint sensitization in the animal model of visceral pain.

Modulation of visceral pain by brain nuclei and brain circuits and the role of acupuncture: a narrative review.

Visceral pain is a complex and heterogeneous pain condition that is often associated with pain-related negative emotional states, including anxiety and depression, and can exert serious effects on a patient's physical and mental health. According to modeling stimulation protocols, the current animal models of visceral pain mainly include the mechanical dilatation model, the ischemic model, and the inflammatory model. Acupuncture can exert analgesic effects by integrating and interacting input signals from acupuncture points and the sites of pain in the central nervous system. The brain nuclei involved in regulating visceral pain mainly include the nucleus of the solitary tract, parabrachial nucleus (PBN), locus coeruleus (LC), rostral ventromedial medulla (RVM), anterior cingulate cortex (ACC), paraventricular nucleus (PVN), and the amygdala. The neural circuits involved are PBN-amygdala, LC-RVM, amygdala-insula, ACC-amygdala, claustrum-ACC, bed nucleus of the stria terminalis-PVN and the PVN-ventral lateral septum circuit. Signals generated by acupuncture can modulate the central structures and interconnected neural circuits of multiple brain regions, including the medulla oblongata, cerebral cortex, thalamus, and hypothalamus. This analgesic process also involves the participation of various neurotransmitters and/or receptors, such as 5-hydroxytryptamine, glutamate, and enkephalin. In addition, acupuncture can regulate visceral pain by influencing functional connections between different brain regions and regulating glucose metabolism. However, there are still some limitations in the research efforts focusing on the specific brain mechanisms associated with the effects of acupuncture on the alleviation of visceral pain. Further animal experiments and clinical studies are now needed to improve our understanding of this area.

Critical evaluation of animal models of visceral pain for therapeutics development: A focus on irritable bowel syndrome.

The classification of chronic visceral pain is complex, resulting from persistent inflammation, vascular (ischemic) mechanisms, cancer, obstruction or distension, traction or compression, and combined mechanisms, as well as unexplained functional mechanisms. Despite the prevalence, treatment options for chronic visceral pain are limited. Given this unmet clinical need, the development of novel analgesic agents, with defined targets derived from preclinical studies, is urgently needed. While various animal models have played an important role in our understanding of visceral pain, our knowledge is far from complete. Due to the complexity of visceral pain, this document will focus on chronic abdominal pain, which is the major complaint in patients with disorders of the gut-brain interaction, also referred to as functional gastrointestinal disorders, such as irritable bowel syndrome (IBS). Models for IBS are faced with challenges including a complex clinical phenotype, which is comorbid with other conditions including anxiety, depression, painful bladder syndrome, and chronic pelvic pain. Based upon the multifactorial nature of IBS with complicated interactions between biological, psychological, and sociological variables, no single experimental model recapitulates all the symptoms of IBS. This position paper will contextualize chronic visceral pain using the example of IBS and focus on its pathophysiology while providing a critical review of current animal models that are most relevant, robust, and reliable in which to screen promising therapeutics to alleviate visceral pain and delineate the gaps and challenges with these models. We will also highlight, prioritize, and come to a consensus on the models with the highest face/construct validity.

Human visceral nociception: findings from translational studies in human tissue.

Peripheral sensitization of nociceptors during disease has long been recognized as a leading cause of inflammatory pain. However, a growing body of data generated over the last decade has led to the increased understanding that peripheral sensitization is also an important mechanism driving abdominal pain in highly prevalent functional bowel disorders, in particular, irritable bowel syndrome (IBS). As such, the development of drugs that target pain-sensing nerves innervating the bowel has the potential to be a successful analgesic strategy for the treatment of abdominal pain in both organic and functional gastrointestinal diseases. Despite the success of recent peripherally restricted approaches for the treatment of IBS, not all drugs that have shown efficacy in animal models of visceral pain have reduced pain end points in clinical trials of IBS patients, suggesting innate differences in the mechanisms of pain processing between rodents and humans and, in particular, how we model disease states. To address this gap in our understanding of peripheral nociception from the viscera and the body in general, several groups have developed experimental systems to study nociception in isolated human tissue and neurons, the findings of which we discuss in this review. Studies of human tissue identify a repertoire of human primary afferent subtypes comparable to rodent models including a nociceptor population, the targeting of which will shape future analgesic development efforts. Detailed mechanistic studies in human sensory neurons combined with unbiased RNA-sequencing approaches have revealed fundamental differences in not only receptor/channel expression but also peripheral pain pathways.

Toward an effective peripheral visceral analgesic: responding to the national opioid crisis.

This minireiew summarizes recent new developments in visceral analgesics. This promising field is important, as a new approach to address abdominal pain with peripheral visceral analgesics is considered a key approach to addressing the current opioid crisis. Some of the novel compounds address peripheral pain mechanisms through modulation of opioid receptors via biased ligands, nociceptin/orphanin FQ opioid peptide (NOP) receptor, or dual action on NOP and μ-opioid receptor, buprenorphine and morphiceptin analogs. Other compounds target nonopioid mechanisms, including cannabinoid (CB2), N-methyl-d-aspartate, calcitonin gene-related peptide, estrogen, and adenosine A2B receptors and transient receptor potential (TRP) channels (TRPV1, TRPV4, and TRPM8). Although current evidence is based predominantly on animal models of visceral pain, early human studies also support the evidence from the basic and animal research. This augurs well for the development of nonaddictive, visceral analgesics for treatment of chronic abdominal pain, an unmet clinical need.

Animal models of gastrointestinal and liver diseases. Animal models of visceral pain: pathophysiology, translational relevance, and challenges.

Visceral pain describes pain emanating from the thoracic, pelvic, or abdominal organs. In contrast to somatic pain, visceral pain is generally vague, poorly localized, and characterized by hypersensitivity to a stimulus such as organ distension. Animal models have played a pivotal role in our understanding of the mechanisms underlying the pathophysiology of visceral pain. This review focuses on animal models of visceral pain and their translational relevance. In addition, the challenges of using animal models to develop novel therapeutic approaches to treat visceral pain will be discussed.

Guanylate cyclase-C/cGMP: an emerging pathway in the regulation of visceral pain

Activation of guanylate cyclase-C (GC-C) expressed predominantly on intestinal epithelial cells by guanylin, uroguanylin or the closely related GC-C agonist peptide, linaclotide, stimulates generation, and release of cyclic guanosine-3′,5′-monophosphate (cGMP). Evidence that the visceral analgesic effects of linaclotide are mediated by a novel, GC-C-dependent peripheral sensory mechanism was first demonstrated in animal models of visceral pain. Subsequent studies with uroguanylin or linaclotide have confirmed the activation of a GC-C/cGMP pathway leading to increased submucosal cGMP mediated by cGMP efflux pumps, which modulates intestinal nociceptor function resulting in peripheral analgesia. These effects can be reproduced by the addition of exogenous cGMP and support a role for GC-C/cGMP signaling in the regulation of visceral sensation, a physiological function that has not previously been linked to the GC-C/cGMP pathway. Notably, targeting the GC-C/cGMP pathway for treatment of gastrointestinal pain and abdominal sensory symptoms has now been validated in the clinic. In 2012, linaclotide was approved in the United States and European Union for the treatment of adult patients with irritable bowel syndrome with constipation.

σ1Receptors Are Involved in the Visceral Pain Induced by Intracolonic Administration of Capsaicin in Mice

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. 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). 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. 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.

Relative potency of pregabalin, gabapentin, and morphine in a mouse model of visceral pain

Pregabalin is probably more effective than prototype gabapentin in different kinds of pain treatments. This study was performed to compare the potency of gabapentin, pregabalin, and morphine in a well-established model of visceral pain. The number of abdominal contractions was counted for 30 min in adult male mice that received different doses of pregabalin, gabapentin, morphine, or placebo intraperitoneally 30 min before receiving 0.6% acetic acid 10 mL·kg(-1).The antinociceptive effect of each drug dose was determined as a percentage of the reduction in the number of acetic acid-induced writhes. The effective doses, for 20%, 50%, and 80% response (ED(20), ED(50), and ED(80), respectively), of each drug were calculated using least squares linear regression analysis, and then dose-response curves were compared. Pregabalin, gabapentin, and morphine produced a linear dose-dependent antinociceptive effect (coefficient of determination [r(2)] > 0.9). No difference was observed between slopes of dose-response curves. The ED(50) estimates (95% confidence interval) for pregabalin, gabapentin, and morphine were 17.1 (12.9 to 22.1) mg·kg(-1), 87.1 (45.8 to 129.8) mg·kg(-1), and 0.2 (0.1 to 0.3) mg·kg(-1), respectively. In this animal model of visceral pain, all three drugs exhibited parallel dose-response curves. Pregabalin had five times the potency of gabapentin and 1/85(th) the potency of morphine. Similar potency ratios may apply in clinical practice. Despite some limitations of animal studies, this model could be useful for comparing new analgesics in visceral pain treatment.

Achieving translation in models of visceral pain

The failure of drugs to modify pain end points in clinical trials for irritable bowel syndrome (IBS) highlights the knowledge gap that exists in the translation of efficacy in animal models of visceral pain into the clinic. Recent progress has been made towards improving the translation of visceral pain, particularly with regard to the activation of the sensory nerves which relay pain from the gut to the brain. This review will focus on studies which have identified the presence of an altered gastrointestinal and immune environment in IBS patients. The development of human gastrointestinal visceral afferent recordings has allowed direct comparison between sensory nerve studies in animals and human, as well as important advances in our understanding of the ion channels that underpin the changes in sensory nerve excitability.

The Safety, Tolerability, Systemic Exposure, and Effect on Bowel Habits of Single and Multiple Doses of the Intestinal Sodium Re-Uptake Inhibitor RDX5791 in Normal Healthy Volunteers

Purpose: RDX5791 is a first-in-class minimally systemic, small molecule NHE3 inhibitor in clinical development for the treatment of IBS-C and other constipation-related diseases. The intestinal Na+/H+ antiport protein NHE3 plays a key role in the uptake of sodium, and thus water, from the intestinal lumen. RDX5791 increases intestinal sodium leading to enhanced intestinal fluid volume and transit. RDX5791 has also been shown to have an anti-nociceptive effect in an IBS animal model of visceral pain. The purpose of this study was to evaluate the safety, tolerability, systemic exposure and pharmacodynamics of RDX5791, after single and 7 daily oral doses to healthy volunteers. Methods: A randomized, double-blind, placebo-controlled, study was performed in 80 healthy male and female volunteers at single ascending doses of 10, 50, 150, 450, and 900 mg (6 treated, 2 placebo/group) and multiple ascending doses of 3, 10, 30, and 100 mg (8 treated, 2 placebo). Safety assessments were performed routinely and included vital signs, ECGs, hematology, and serum chemistry evaluations as well as adverse event assessments. The pharmacodynamics of RDX5791 on bowel habits was assessed by recording stool frequency (time), weight, and consistency utilizing the Bristol Stool Form Scale (BSFS). Results: RDX5791 was very well tolerated and exhibited minimal systemic absorption. There were no SAEs observed and very few adverse events. In the single-dose phase, there was no RDX5791 detectable in plasma samples at doses of 10 and 50 mg (LOQ 0.5 ng/mL) and transient amounts (0.5 to 1ng/mL) at one time point in 11 of 18 subjects at doses of 150, 450 and 900 mg. In the multiple-dose phase, only 2 of 576 plasma samples had any detectable RDX5791 (< 1 ng/mL). In the single-dose phase, RDX5791 caused a decrease in time to first stool (TTFS) in all dose groups, most notably at 50 mg, (4.3 ± 2.4 hr versus 23.2 ± 7.8 hr for placebo). RDX5791 also caused statistically significant increases from placebo in BSFS (at 50 mg) and stool weight (at 150 mg). In the multiple-dose phase, RDX5791 caused a dose-related increase in BSFS with no increase in stool frequency as compared to placebo. RDX5791 also caused a decrease in TTFS and an increase in the % of days with a spontaneous bowel movement. Conclusion: RDX5791 is a minimally systemic, first-in-class inhibitor of NHE3, that was well tolerated at all doses tested (10-900 mg). RDX5791 exhibited pharmacodynamic activity in healthy volunteers consistent with its mechanism of action. Preclinical and clinical data corroborate the potential of RDX5791 for the treatment of IBS-C and other constipation-related diseases. In addition to the study described here, a phase 2 study to assess the efficacy of RDX5791 in IBS-C is ongoing. Disclosure: All authors are employees of Ardelyx, the sponsor of the clinical trial This research was supported by an industry grant from Ardelyx.

Role of oxidative stress in animal model of visceral pain

Reactive oxygen species play an important role both in physiological and pathophysiological reactions. The aim of this study was to determine the role of free radicals and antioxidants in the development of visceral pain. Visceral pain was produced by colorectal distension (CRD) in adult rats. CRD was caused by insertion of a lubricated latex balloon into the descending colon and rectum followed by inflation to 80 mm Hg for 10 min. During CRD, visceral pain was rated on 0–3.5 point scale. Oxidative stress was determined indirectly by measurement of free radical scavenging enzymes (glutathione peroxidase (GPx) and superoxide dismutase (SOD)) in the blood, liver and brain. Following CRD we observed (1) all rats expressed signs of visceral pain (overall rating was 1.83), (2) SOD and GPx levels were increased in the liver and blood, and decreased in the brain samples and (3) administration of the antioxidant Trolox, a water-soluble derivate of vitamin E, prior to CRD, prevented SOD and GPx changes in the liver, blood and brain, but did not affect pain scores. It was concluded, that CRD as a model of visceral pain, increases oxidative stress in animals, which could be prevented by prior administration of antioxidants; however, antioxidants did not attenuate signs of visceral pain caused by CRD.

Up-regulation of galanin and corticotropin-releasing hormone mRNAs in the key hypothalamic and amygdaloid nuclei in a mouse model of visceral pain

Cyclophosphamide (CP)-induced cystitis is often used as an animal model of visceral pain. Various neuropeptides in the hypothalamic and amygdaloid nuclei are implicated in pain-induced responses. However, little information is available regarding the regulation of the neuropeptides in response to visceral pain. In the present study, we examined the effects of CP-induced cystitis on the levels of mRNAs encoding galanin, corticotropin-releasing hormone (CRH), substance P, and enkephalins in the hypothalamic and limbic nuclei using in situ hybridization histochemistry in mouse. Galanin mRNA levels in CP-treated group increased significantly in the arcuate nucleus and the paraventricular nucleus (PVN) but not in the medial preoptic area after the intraperitoneal administration of CP (200 mg/kg body weight) in comparison to those in saline-treated group. CRH mRNA levels in CP-treated group also increased significantly in the central amygdala as well as the PVN after the CP administration. In contrast, CP-induced cystitis failed to upregulate the preprotachykinin-A and preproenkephalin genes which encode substance P and enkephalins, respectively in the hypothalamic and limbic nuclei at any of the time points examined. These results suggest that visceral nociception may upregulate both galanin and CRH gene expression in the hypothalamic and limbic nuclei.

Pharmacological and Pharmacokinetic Aspects of Functional Gastrointestinal Disorders

Medications are commonly used for the treatment of patients with functional gastrointestinal disorders. The general goal of this report is to review the pharmacokinetics and pharmacology of medications used in functional gastrointestinal disorders. Methods included literature review, consensus evaluation of the evidence for each topic assigned originally to 1 or 2 authors, and broader review at a harmonization session as part of the Rome III process. This report reviews the animal models that have been validated for the study of effects of pharmacologic agents on sensation and motility; the preclinical pharmacology, pharmacokinetics, and toxicology usually required for introduction of novel therapeutic agents; the biomarkers validated for studies of sensation and motility end points with experimental medications in humans; the pharmacogenomics applied to these medications and disorders; and the pharmacology of agents that are applied or have potential for treatment of functional gastrointestinal disorders, including psychopharmacologic agents. Clinician and basic investigators involved in the treatment or investigation of functional gastrointestinal disorders or disease models need to have a comprehensive understanding of a vast range of medications. It is anticipated that the interaction between investigators of basic science, basic and applied pharmacology, and clinical trials will lead to better treatment of these disorders.

Recent advances in understanding molecular mechanisms of primary afferent activation

Thermal, mechanical, and chemical stimuli depolarise specialised damage sensing neurons to initiate electrical signals that may ultimately result in a sensation of pain. Over the past decade many of the...

Characterization of Cyclophosphamide Cystitis, a Model of Visceral and Referred Pain, in the Mouse: Species and Strain Differences

Existing animal models of visceral pain in the mouse are of limited practical usefulness since they are labor intensive or not visceral specific. Recently a rat model of cyclophosphamide (CP) cystitis was developed that requires only intraperitoneal injection and features inflammation confined to the bladder. We adapted this model for use in multiple mouse strains to investigate the genetic basis of variability in visceral nociception. Outbred CD-1 mice and 12 inbred mouse strains were tested for behavioral changes induced by CP (0 to 300 mg/kg intraperitoneally). We noted that despite the absence of postural changes or abdominal crises in CD-1 mice, CP produced dose dependent decreases in voluntary locomotor activity unaccompanied by ataxia measured on the rotarod test; referred hyperalgesia of the tail base region but not of the hind paw, which was inhibited in dose dependent fashion by morphine (0 to 20 mg/kg); and bladder inflammation corresponding to these behavioral indices. Furthermore, the extent of hypolocomotion was genotype dependent across 12 inbred strains. The simple and automatable nature of CP cystitis using hypolocomotion as a dependent measure renders it an attractive model in which to investigate the genetic and physiological bases of visceral pain. Comparison of strain sensitivity to CP induced hypolocomotion with other nociceptive assays suggests that genes specific to visceral nociception may exist.

The capsaicin analogue SDZ249-665 attenuates the hyper-reflexia and referred hyperalgesia associated with inflammation of the rat urinary bladder

This study assessed the effects of the systemically administered capsaicin analogue SDZ249-665 in an animal model of visceral pain and hyper-reflexia. The effects of prophylactic administration of SDZ249-665 (in the dose range 0.05–1 mg/kg) on the viscero-visceral hyper-reflexia (VVH) and the referred viscero-somatic hyperalgesia to mechanical stimuli (VSH) associated with turpentine inflammation of the rat urinary bladder were evaluated. SDZ249-665 attenuated both the VVH and the VSH in a dose related fashion. In the VVH model, following solvent control administration, intra-vesical turpentine administration was associated with a significant reduction in micturition threshold to 43.7% (SEM 6.3) of baseline, indicating the presence of a VVH. This effect was not observed when animals were prophylactically treated with SDZ249-665 alone. At a dose of 0.1 mg/kg the micturition threshold was 90.7% (SEM 10.2) of baseline at 1 h after intra-vesical instillation of turpentine. In the VSH model, curves were plotted of the difference in fore and hind limb withdrawal latencies from a mechanical stimulus and the area under these curves (AUCs) were compared between different treatment protocols. Intra-vesical turpentine was associated with a negative deflection of the curve (AUC −5.2×10 3 SEM 1.7) in comparison with naı̈ve animals (AUC −0.02×10 3 SEM 0.6), indicative of a referred hyperalgesia. This was prevented, in a dose-related manner, by prophylactic administration of SDZ249-665. For example, at a dose of 0.5 mg/kg the AUC was +0.4×10 3 (SEM 0.8). These findings support previous work indicating that capsaicin sensitive neurones participate in patho-physiological events occurring following inflammation of the bladder, and provides evidence that systemically active capsaicin based compounds may be developed for use in the clinical setting.