Colorectal Distension In Mice Research Articles

An anesthesia protocol for robust and repeatable measurement of behavioral visceromotor responses to colorectal distension in mice.

Visceral motor responses (VMR) to graded colorectal distension (CRD) have been extensively implemented to assess the level of visceral pain in awake rodents, which are inevitably confounded by movement artifacts and cannot be conveniently implemented to assess invasive neuromodulation protocols for treating visceral pain. In this report, we present an optimized protocol with prolonged urethane infusion that enables robust and repeatable recordings of VMR to CRD in mice under deep anesthesia, providing a two-hour window to objectively assess the efficacy of visceral pain management strategies. During all surgical procedures, C57BL/6 mice of both sexes (8-12 weeks, 25-35 g) were anesthetized with 2% isoflurane inhalation. An abdominal incision was made to allow Teflon-coated stainless steel wire electrodes to be sutured to the oblique abdominal musculature. A thin polyethylene catheter (Φ 0.2 mm) was placed intraperitoneally and externalized from the abdominal incision for delivering the prolonged urethane infusion. A cylindric plastic-film balloon (Φ 8 mm x 15 mm when distended) was inserted intra-anally, and its depth into the colorectum was precisely controlled by measuring the distance between the end of the balloon and the anus. Subsequently, the mouse was switched from isoflurane anesthesia to the new urethane anesthesia protocol, which consisted of a bout of infusion (0.6 g urethane per kg weight, g/kg) administered intraperitoneally via the catheter and continuous low-dose infusion throughout the experiment at 0.15-0.23 g per kg weight per hour (g/kg/h). Using this new anesthesia protocol, we systematically investigated the significant impact of balloon depth into the colorectum on evoked VMR, which showed a progressive reduction with increased balloon insertion depth from the rectal region into the distal colonic region. Intracolonic TNBS treatment induced enhanced VMR to CRD of the colonic region (>10 mm from the anus) only in male mice, whereas colonic VMR was not significantly altered by TNBS in female mice. Conducting VMR to CRD in anesthetized mice using the current protocol will enable future objective assessments of various invasive neuromodulatory strategies for alleviating visceral pain.

Identification of bacterial lipopeptides as key players in IBS

ObjectivesClinical studies revealed that early-life adverse events contribute to the development of IBS in adulthood. The aim of our study was to investigate the relationship between prenatal stress (PS), gut...

Agonist that activates the µ-opioid receptor in acidified microenvironments inhibits colitis pain without side effects

ObjectiveThe effectiveness of µ-opioid receptor (MOPr) agonists for treatment of visceral pain is compromised by constipation, respiratory depression, sedation and addiction. We investigated whether a fentanyl analogue, (±)-N-(3-fluoro-1-phenethylpiperidine-4-yl)-N-phenyl propionamide (NFEPP),...

Firing patterns and functional roles of different classes of spinal afferents in rectal nerves during colonic migrating motor complexes in mouse colon

The functional role of the different classes of visceral afferents that innervate the large intestine is poorly understood. Recent evidence suggests that low-threshold, wide-dynamic-range rectal afferents play an important role in the detection and transmission of visceral pain induced by noxious colorectal distension in mice. However, it is not clear which classes of spinal afferents are activated during naturally occurring colonic motor patterns or during intense contractions of the gut smooth muscle. We developed an in vitro colorectum preparation to test how the major classes of rectal afferents are activated during spontaneous colonic migrating motor complex (CMMC) or pharmacologically induced contraction. During CMMCs, circular muscle contractions increased firing in low-threshold, wide-dynamic-range muscular afferents and muscular-mucosal afferents, which generated a mean firing rate of 1.53 ± 0.23 Hz (n = 8) under isotonic conditions and 2.52 ± 0.36 Hz (n = 17) under isometric conditions. These low-threshold rectal afferents were reliably activated by low levels of circumferential stretch induced by increases in length (1-2 mm) or load (1-3 g). In a small proportion of cases (5 of 34 units), some low-threshold muscular and muscular-mucosal afferents decreased their firing rate during the peak of the CMMC contractions. High-threshold afferents were never activated during spontaneous CMMC contractions or tonic contractions induced by bethanechol (100 μM). High-threshold rectal afferents were only activated by intense levels of circumferential stretch (10-20 g). These results show that, in the rectal nerves of mice, low-threshold, wide-dynamic-range muscular and muscular-mucosal afferents are excited during contraction of the circular muscle that occurs during spontaneous CMMCs. No activation of high-threshold rectal afferents was detected during CMMCs or intense contractile activity in naïve mouse colorectum.

Mo1658 Acute Psychological Stress Induces an Immediate Naloxone-Independent Visceral Analgesia to Colorectal Distension in Mice

Mo1658 Acute Psychological Stress Induces an Immediate Naloxone-Independent Visceral Analgesia to Colorectal Distension in Mice

Antihyperalgesic Effect of Stable Pansomatostatin Agonist, ODT-8 and Selective Somatostatin Type 1 Receptor (SST 1) Agonist Injected Intraperitoneally During Colorectal Distension in Mice

Antihyperalgesic Effect of Stable Pansomatostatin Agonist, ODT-8 and Selective Somatostatin Type 1 Receptor (SST 1) Agonist Injected Intraperitoneally During Colorectal Distension in Mice

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.

Lack of interaction between psychological stress and DSS-induced colitis affecting colonic sensitivity during colorectal distension in mice

Inflammation and stress have been associated to colorectal hypersensitivity in functional gastrointestinal disorders. We explored if colonic inflammation and stress, alone or combined, participate in the development of visceral hypersensitivity in a mouse model. First, female mice were exposed to 3% dextran sulphate sodium (DSS, 5 days) to induce chronic colitis, followed by repeated psychological stress (water avoidance stress, WAS; 1 h daily/10 days). In a separate experiment, female mice were subjected to WAS and thereafter received 3% DSS. Colitis was evaluated at necropsy. Faecal pellet output served as a marker of stress effect on colonic motility. Visceral pain was assessed at fixed time-points monitoring abdominal contractions during colorectal distension (CRD). DSS provoked a mild chronic colitis that was not affected by previous WAS or aggravated by subsequent stress. WAS induced a significant increase in pellet output, although the response was attenuated in animals with colitis. Responses to CRD were similar in all experimental groups, with transient hyperalgesia observed only during acute colitis. Under the present conditions, neither colitis nor stress, alone or in combination, significantly affected the responses to CRD in mice, suggesting that there is not a direct relationship between inflammation and stress and the development of visceral hypersensitivity.

Involvement of κ‐opioid receptors in visceral nociception in mice

It has been shown that the behavioural responses to chemically evoked visceral nociception are increased in transgenic mice lacking the kappa-opioid receptor (KOR). The aim of the present study was to evaluate the contribution of KOR in mechanically evoked visceral pain by performing colorectal distension (CRD) and monitoring the subsequent visceromotor response (VMR) in control mice (KOR(+/+)) and in mice lacking KOR (KOR(-/-)). Pseudo-affective visceral pain responses were evoked in conscious mice using increasing (10-80 mmHg) and repeated (12 x 55 mmHg) phasic CRD paradigms. The resulting VMR was determined by monitoring the electromyographic activity of the abdominal muscle. The increasing and repeated CRD paradigms, respectively, evoked similar responses in both KOR(+/+) and KOR(-/-) mice. The selective KOR-agonists U-69593 (5 and 25 mg kg(-1), s.c.) and asimadoline (25 mg kg(-1), s.c.) significantly decreased the VMR in KOR(+/+) mice, while having no effect in KOR(-/-) mice. In contrast, the selective mu-opioid receptor agonist fentanyl significantly reduced the VMR in both types of mice and appeared more efficacious in KOR(-/-) mice. The opioid receptor antagonist naloxone (0.3-30 mg kg(-1) s.c.) did not affect the response to CRD in C57BL/6 mice at any dose tested. In conclusion, the data confirm that the KOR agonists used in this study inhibit the VMR to CRD in mice by acting via KOR receptors. In addition, the data suggest that the endogenous opioid system is not likely to modulate the VMR to mechanically evoked visceral pain in mice.

Assessment of Visceral Pain-Related Pseudo-Affective Responses to Colorectal Distension in Mice by Intracolonic Manometric Recordings

Recently, a new manometric method has been proposed to quantify visceromotor responses (VMR) to colorectal distension (CRD) in rats. This method is based on monitoring pressure changes within the distending balloon during CRD. This study assesses the applicability of such a technique to the quantification of VMRs to CRD in mice. Electrical activity of the abdominal muscles and pressure changes within the distending balloon (mechanical response) were simultaneously recorded in conscious mice during CRD (phasic ascending, 10-80 mm Hg, or repetitive, 55 mm Hg). There was a clear stimulus-response relationship with a strong correlation between electrical and mechanical responses during the ascending (r 2 = 0.899, n = 7) or repetitive phasic CRD (r 2 = 0.926, n = 8). Repetitive phasic distensions (55 mm Hg) increased the mechanical and electrical responses by 71 ± 20% and 42 ± 16%, respectively (pulses 10-12 vs. 1-3; n = 8, both P < .01). Atropine (0.5 or 1 mg/kg, subcutaneously) did not affect the mechanical response to CRD. The μ-opioid agonist, fentanyl (0.05 mg/kg, subcutaneously), completely prevented the sensitizing response associated to repetitive distensions. These results show that noninvasive, surgery-free manometry of intracolonic pressure is a reliable method to assess VMRs to CRD in mice. The analgesic effect of compounds could be determined, indicating that the method can be used in pharmacologic studies. Perspective The model presented to assess visceral pain in mice allows a broad use of this species in pharmacological studies and will be of use in the characterization of potential targets and new drugs for the treatment of human pathologies with visceral pain arising from the gut as a significant component.

Effect of DSS‐induced colitis on visceral sensitivity to colorectal distension in mice

The present study aimed at evaluating the effect of dextran sodium sulphate (DSS)-induced colitis on visceral sensitivity, measured as the visceromotor response (VMR) to colorectal distension (CRD) in BALB/c and C57Bl/6 male mice. Inflammation was induced by the addition of 4% DSS to the drinking water for 5 (C57Bl/6) or 6-7 days (BALB/c). Parallel groups were used to monitor histopathological changes and visceral sensitivity. Pseudo-affective visceral pain responses were evoked using an increasing phasic CRD paradigm (10-60 mmHg) in conscious mice on predetermined days (pretreatment controls, 12, 16, 20, 30, 40 and 51). In both mouse strains, significant histopathological changes developed between days 2 and 5 of DSS treatment, and persisted until day 12 (P < 0.05). On day 15, inflammatory scores were reduced by about 50%. Despite evidence of inflammation in DSS-treated mice, no differences could be shown in the VMR to CRD between DSS-treated mice and controls at any time point tested. In addition, no differences were seen before and after DSS treatment in the same group of mice. In conclusion, these data suggest that DSS-induced colonic inflammation does not affect the visceral sensitivity to CRD, neither at short or long term, in BALB/c or C57Bl/6 male mice.