Identification of the Visceral Pain Pathway Activated by Noxious Colorectal Distension in Mice

In patients with irritable bowel syndrome, visceral pain is evoked more readily following distension of the colorectum. However, the identity of extrinsic afferent nerve pathway that detects and transmits visceral pain from the colorectum to the spinal cord is unclear. In this study, we identified which extrinsic nerve pathway(s) underlies nociception from the colorectum to the spinal cord of rodents. Electromyogram recordings were made from the transverse oblique abdominal muscles in anesthetized wild type (C57BL/6) mice and acute noxious intraluminal distension stimuli (100–120 mmHg) were applied to the terminal 15 mm of colorectum to activate visceromotor responses (VMRs). Lesioning the lumbar colonic nerves in vivo had no detectable effect on the VMRs evoked by colorectal distension. Also, lesions applied to the right or left hypogastric nerves failed to reduce VMRs. However, lesions applied to both left and right branches of the rectal nerves abolished VMRs, regardless of whether the lumbar colonic or hypogastric nerves were severed. Electrical stimulation applied to either the lumbar colonic or hypogastric nerves in vivo, failed to elicit a VMR. In contrast, electrical stimulation (2–5 Hz, 0.4 ms, 60 V) applied to the rectum reliably elicited VMRs, which were abolished by selective lesioning of the rectal nerves. DiI retrograde labeling from the colorectum (injection sites 9–15 mm from the anus, measured in unstretched preparations) labeled sensory neurons primarily in dorsal root ganglia (DRG) of the lumbosacral region of the spinal cord (L6-S1). In contrast, injection of DiI into the mid to proximal colon (injection sites 30–75 mm from the anus, measured in unstretched preparations) labeled sensory neurons in DRG primarily of the lower thoracic level (T6-L2) of the spinal cord. The visceral pain pathway activated by acute noxious distension of the terminal 15 mm of mouse colorectum is transmitted predominantly, if not solely, through rectal/pelvic afferent nerve fibers to the spinal cord. The sensory neurons of this spinal afferent pathway lie primarily in the lumbosacral region of the spinal cord, between L6 and S1

Stress-related alterations of visceral sensation: animal models for irritable bowel syndrome study.

Stressors of different psychological, physical or immune origin play a critical role in the pathophysiology of irritable bowel syndrome participating in symptoms onset, clinical presentation as well as treatment outcome. Experimental stress models applying a variety of acute and chronic exteroceptive or interoceptive stressors have been developed to target different periods throughout the lifespan of animals to assess the vulnerability, the trigger and perpetuating factors determining stress influence on visceral sensitivity and interactions within the brain-gut axis. Recent evidence points towards adequate construct and face validity of experimental models developed with respect to animals' age, sex, strain differences and specific methodological aspects such as non-invasive monitoring of visceromotor response to colorectal distension as being essential in successful identification and evaluation of novel therapeutic targets aimed at reducing stress-related alterations in visceral sensitivity. Underlying mechanisms of stress-induced modulation of visceral pain involve a combination of peripheral, spinal and supraspinal sensitization based on the nature of the stressors and dysregulation of descending pathways that modulate nociceptive transmission or stress-related analgesic response

Peripheral Mediators of Colorectal Nociception and Sensitization

Several ion channels are thought to facilitate colorectal afferent neuron sensitization, which contributes to abdominal pain in irritable bowel syndrome (IBS). In the present work, I hypothesized that two such channels – TRPV1 and P2X3 – cooperate to mediate colorectal pain and hypersensitivity. To test this, I employed TRPV1-P2X3 double knockout (TPDKO) mice and pharmacological antagonists and evaluated combined channel contributions to whole- organism responses to colorectal distension (CRD) and afferent fiber responses to colorectal stretch. Baseline responses to CRD were unexpectedly greater in TPDKO compared with control mice, but zymosan-produced CRD hypersensitivity was absent in TPDKO mice. Relative to control mice, proportions of afferent mechano-sensitive and -insensitive classes were not different in TPDKO mice. Whereas responses of mucosal and serosal class afferents to mechanical probing were unaffected, responses of muscular (but not muscular/mucosal) afferents to stretch were significantly attenuated in TPDKO mice as was sensitization by inflammatory soup of both muscular and muscular/mucosal afferents. In pharmacological studies, the TRPV1 antagonist A889425 and P2X3 antagonist TNP-ATP, alone and in combination, applied onto stretch-sensitive afferent endings attenuated afferent responses to stretch; combined antagonism produced greater attenuation. In the aggregate, these observations suggest that: (1) genetic manipulation of TRPV1 and P2X3 leads to reduction in colorectal mechanosensation peripherally and compensatory changes and/or disinhibition of other channels centrally and (2) combined pharmacological antagonism produces more robust attenuation of mechanosensation peripherally than single antagonism. The relative importance of these channels appears to be enhanced in hypersensitivity, highlighting the potential utility of multi-target pharmacotherapy in IBS

Mechanisms contributing to visceral hypersensitivity : focus on splanchnic afferent nerve signalling

This is the peer reviewed version of the following article: Deiteren, A., De Man, J. G., Keating, C., Jiang, W., De Schepper, H. U., Pelckmans, P. A., Francque, S. M. and De Winter, B. Y. (2015), Mechanisms contributing to visceral hypersensitivity: focus on splanchnic afferent nerve signaling. Neurogastroenterology & Motility, 27: 1709–1720, which has been published in final form at doi:10.1111/nmo.12667. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.Visceral hypersensitivity is a main characteristic of functional bowel disorders and is mediated by both peripheral and central factors. We investigated whether enhanced splanchnic afferent signaling in vitro is associated with visceral hypersensitivity in vivo in an acute and postinflammatory rat model of colitis.Peer reviewedFinal Accepted Versio

How should we define a nociceptor in the gut-brain axis?

In the past few years, there has been extraordinary interest in how the gut communicates with the brain. This is because substantial and gathering data has emerged to suggest that sensory nerve pathways between the gut and brain may contribute much more widely in heath and disease, than was originally presumed. In the skin, the different types of sensory nerve endings have been thoroughly characterized, including the morphology of different nerve endings and the sensory modalities they encode. This knowledge is lacking for most types of visceral afferents, particularly spinal afferents that innervate abdominal organs, like the gut. In fact, only recently have the nerve endings of spinal afferents in any visceral organ been identified. What is clear is that spinal afferents play the major role in pain perception from the gut to the brain. Perhaps surprisingly, the majority of spinal afferent nerve endings in the gut express the ion channel TRPV1, which is often considered to be a marker of nociceptive neurons. And, a majority of gut-projecting spinal afferent neurons expressing TRPV1 are activated at low thresholds, in the normal physiological range, well below the normal threshold for detection of painful sensations. This introduces a major conundrum regarding visceral nociception. How should we define a nociceptor in the gut? We discuss the notion that nociception from the gut wall maybe a process encrypted into multiple different morphological types of spinal afferent nerve ending, rather than a single class of sensory ending, like free-endings, suggested to underlie nociception in skin

Extrinsic primary afferent signalling in the gut

Visceral sensory neurons activate reflex pathways that control gut function and also give rise to important sensations, such as fullness, bloating, nausea, discomfort, urgency and pain. Sensory neurons are organised into three distinct anatomical pathways to the central nervous system (vagal, thoracolumbar and lumbosacral). Although remarkable progress has been made in characterizing the roles of many ion channels, receptors and second messengers in visceral sensory neurons, the basic aim of understanding how many classes there are, and how they differ, has proven difficult to achieve. We suggest that just five structurally distinct types of sensory endings are present in the gut wall that account for essentially all of the primary afferent neurons in the three pathways. Each of these five major structural types of endings seems to show distinctive combinations of physiological responses. These types are: 'intraganglionic laminar' endings in myenteric ganglia; 'mucosal' endings located in the subepithelial layer; 'muscular–mucosal' afferents, with mechanosensitive endings close to the muscularis mucosae; 'intramuscular' endings, with endings within the smooth muscle layers; and 'vascular' afferents, with sensitive endings primarily on blood vessels. 'Silent' afferents might be a subset of inexcitable 'vascular' afferents, which can be switched on by inflammatory mediators. Extrinsic sensory neurons comprise an attractive focus for targeted therapeutic intervention in a range of gastrointestinal disorders.Australian National Health and Medical Research Counci

Enhanced responses of the anterior cingulate cortex neurones to colonic distension in viscerally hypersensitive rats

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/65998/1/jphysiol.2005.096073.pd

Expression and Function of Galanin in Colonic Sensory Neurones

The gastrointestinal (GI) tract is innervated by both the enteric nervous system and the sensory nervous system, and it is the latter that is responsible for giving rise to conscious sensations arising from the GI tract, such as pain. In particular, the distal colon is a key source of visceral pain in both inflammatory bowel disease (e.g. ulcerative colitis and Crohn’s disease) and irritable bowel syndrome. In both conditions, pain relief is complicated by the GI side effects of commonly prescribed analgesics, such as non-steroidal anti-inflammatory drugs and opioids. Therefore, furthering knowledge about how sensory innervation of the GI tract is modulated in health and disease has the potential to identify new therapeutic avenues. Galanin is a neuropeptide that has various functions within the central and peripheral nervous systems, e.g. regulation of feeding and modulation of nociceptive pathways respectively. Galanin has been previously demonstrated to modulate the mechanosensitivity of vagal sensory afferents innervating the upper GI tract, but nothing is known about its role in the distal colon, i.e. the lower GI tract. Using mice, I therefore aimed to determine if galanin also modulates the lumbar splanchnic nerve (LSN), which innervates the distal colon, in both healthy and inflamed (hypersensitive) conditions. Using ex vivo LSN electrophysiological recordings I found that galanin dose-dependently inhibits LSN responses to mechanical stimuli and the mechanical hypersensitivity induced by acutely applied inflammatory mediators. Using galanin receptor agonists (GalR1: M671, GalR2: spexin), I identified that GalR1 mediates the inhibitory effects on LSN mechanosensitivity. Using a mouse model of colitis, I found that the LSN was hypersensitive to mechanical stimuli, but that galanin no longer produced any inhibitory effect. Immunohistochemistry experiments using thoracolumbar (T13-L1) and lumbosacral (L6-S1) dorsal root ganglia (DRG) retrograde labelled from the colon demonstrated that galanin is primarily expressed by putative nociceptors, but that no major expression changes occur during colitis. In summary, I have demonstrated that galanin inhibits LSN mechanosensitivity and inflammatory mediators induced mechanohypersensitivity, but that this inhibition is lost in an in vivo model. This work highlights the potential for targeting the galaninergic system to treat GI pain.Cambridge Trust - Vice Chancellor's Award Rosetrees Trust - Postdoctoral Grant (A1296) BBSRC - BB/R006210/

Urine Trouble: A Molecular and Anatomical Examination of Bladder Pain

Interstitial cystitis or painful bladder syndrome (IC/PBS) is a chronic pain syndrome affecting 3-6% of women in the US. Some evidence suggests urothelial injury may cause some cases of IC/PBS. To examine how urothelial injury modulates bladder pain, three injury models were used: Infection with uropathogenic Escherichia coli (UPEC, inflammatory), Protamine sulfate (PS, non-inflammatory chemical), and lipopolysaccharide (LPS, non-infectious and inflammatory) treatment. Surprisingly, injury with PS decreased, while UPEC alone increased the pain response to bladder distention. These data suggest that changes in the urothelium may modulate some forms of IC/PBS. However, in most cases, patients with IC/PBS have no histological abnormalities. Therefore, other molecular mediators may modulate IC/PBS. Glutamate, the major excitatory neurotransmitter in the CNS, modulates a host of physiological responses through the actions of ionotropic (iGluRs) and metabotropic receptors (mGluRs). Metabotropic glutamate receptor 5 (mGluR5) has previously been shown to have an important role in somatic inflammatory and neuropathic pain models. However, there is limited evidence that mGluR5 in the CNS has a role in bladder pain. To determine the function of mGluR5 in non-inflammatory bladder pain, mice with a targeted genetic deletion of mGluR5 (mGluR5 KO) were used. Both mGluR5 KO mice and mice given a specific mGluR5 antagonist (fenobam) had a reduced bladder pain response. Systemic treatment with fenobam in mice infected with UPEC also resulted in a reduced response to bladder distention. Thus suggesting that mGluR5 is necessary for the full expression of inflammatory and non-inflammatory bladder pain. However, these techniques do not indicate the site of action because mGluR5 is expressed throughout the pain neuroaxis. Antagonism of mGuR5 in the right central nucleus of the amygdala (CeA) is analgesic in a somatic inflammatory injury model. To examine the role of mGluR5 in the CeA, pharmacological activation of Group I mGluRs (mGluR5 and 1) was examined. Activation of mGluR5/1 in the CeA was sufficient to increase the pain response to noxious bladder distention. Additionally, pharmacological inhibition and virally mediated conditional deletion of mGluR5 in the CeA reduced the evoked response to bladder distention. Finally, optogenetic activation of the CeA increases the pain response to bladder distention suggesting that mGluR5 activation increases neuronal excitability in the CeA, increasing sensitivity to bladder distention. Overall, these data suggest a role of urothelial injury and mGluR5 in bladder pain

NMDA receptor subunit expression and PAR2 receptor activation in colospinal afferent neurons (CANs) during inflammation induced visceral hypersensitivity

<p>Abstract</p> <p>Background</p> <p>Visceral hypersensitivity is a clinical observation made when diagnosing patients with functional bowel disorders. The cause of visceral hypersensitivity is unknown but is thought to be attributed to inflammation. Previously we demonstrated that a unique set of enteric neurons, colospinal afferent neurons (CANs), co-localize with the NR1 and NR2D subunits of the NMDA receptor as well as with the PAR2 receptor. The aim of this study was to determine if NMDA and PAR2 receptors expressed on CANs contribute to visceral hypersensitivity following inflammation. Recently, work has suggested that dorsal root ganglion (DRG) neurons expressing the transient receptor potential vanilloid-1 (TRPV1) receptor mediate inflammation induced visceral hypersensitivity. Therefore, in order to study CAN involvement in visceral hypersensitivity, DRG neurons expressing the TRPV1 receptor were lesioned with resiniferatoxin (RTX) prior to inflammation and behavioural testing.</p> <p>Results</p> <p>CANs do not express the TRPV1 receptor; therefore, they survive following RTX injection. RTX treatment resulted in a significant decrease in TRPV1 expressing neurons in the colon and immunohistochemical analysis revealed no change in peptide or receptor expression in CANs following RTX lesioning as compared to control data. Behavioral studies determined that both inflamed non-RTX and RTX animals showed a decrease in balloon pressure threshold as compared to controls. Immunohistochemical analysis demonstrated that the NR1 cassettes, N1 and C1, of the NMDA receptor on CANs were up-regulated following inflammation. Furthermore, inflammation resulted in the activation of the PAR2 receptors expressed on CANs.</p> <p>Conclusion</p> <p>Our data show that inflammation causes an up-regulation of the NMDA receptor and the activation of the PAR2 receptor expressed on CANs. These changes are associated with a decrease in balloon pressure in response to colorectal distension in non-RTX and RTX lesioned animals. Therefore, these data suggest that CANs contribute to visceral hypersensitivity during inflammation.</p