Repeated psychological stress-induced alterations of visceral sensitivity and colonic motor functions in mice: influence of surgery and postoperative single housing on visceromotor responses

Abstract

Visceral pain modulation by chronic stress in mice has been little studied. Electromyography (EMG) recording of abdominal muscle contractions, as a proxy to the visceromotor response (VMR), requires electrode implantation and post-surgical single housing (SH) which could affect the VMR to stress. To test this hypothesis, male mice had electrode implantation surgery (S) plus SH, or no surgery and were group housed (NS-GH) or single housed (NS-SH) and exposed to either water avoidance stress (WAS, 1 h/day) or left undisturbed in their home cages for 10 days. The VMR to phasic ascending colorectal distension (CRD) was assessed before (basal) and 24 h after 10 days of WAS or no stress using a surgery-free method of intraluminal colonic pressure (ICP) recording (solid-state manometry). WAS heightened significantly the VMR to CRD at 30, 45, and 60 mmHg in S-SH vs. NS-GH, but not compared to NS-SH conscious mice. Compared to basal CRD, WAS increased VMR at 60 mmHg in the S-SH group and decreased it at 30-60 mmHg in NS-GH mice, while having no effect in NS-SH mice. The average defecation during the hour of repeated WAS over 10 days was 1.9 and 2.4 fold greater in S-SH vs. NS-GH and NS-SH mice, respectively. These data indicate that the combination of S-SH required for VMR monitoring with EMG is an important component of repeated WAS-induced post-stress visceral hypersensitivity and defecation in mice.

Figure 1

Example traces of simultaneous recording of visceral motor response (VMR) to CRD by electromyography (EMG) and intraluminal colonic pressure (ICP) in conscious mice. (A) Schema of the modified miniaturized pressure catheter equipped with the custom-made polyethylene plastic balloon to measure ICP. The 2 cm-long polyethylene balloon is ligated 1 cm below the pressure probe. (B) Original and rectified representative EMG and ICP traces recorded simultaneously from the same mouse in response to CRD (45 mmHg, 10 s). When both raw EMG (upper line) and ICP (second line from the bottom) signals are analyzed in Spike 2 by computing “DC Remove” 1 s to exclude all slow events >2 s (i.e. colonic smooth muscle contractions) and “root mean square (RMS) amplitude” to extract the area AUC of the signal, the resulting EMG and phasic ICP (pICP) signals (middle lines) show an obvious similarity in waveform. (C) Representative EMG and ICP traces at 45 and 60 mmHg CRDs. The VMR is polyphasic, covering the whole distension period with bouts of activity at the beginning and end of the distension period; hence VMR was measured over the 10-s distension.

Figure 2

(A) Ascending phasic CRD results in similar VMR changes as monitored simultaneously by EMG (filled squares) and ICP (open circles) in conscious mice equipped with EMG electrodes in the abdominal wall and single housed for 5 days post surgery. Broken lines represent the linear correlation curves p = 0.0284 for EMG and p = 0.0108 for ICP. (B) Correlation between EMG and ICP responses to ascending phasic CRD in mice. Broken lines represent the 95% confidence intervals, dotted line represents the baseline. Data are expressed as mean ± SEM of AUC from n = 9 mice.

Figure 3

The muscarinic receptor antagonist, atropine did not alter the VMR to CRD monitored with ICP in naïve mice. A first set of ascending phasic CRD was performed in 10 mice and after 45 min, atropine (1 mg/kg) was injected subcutaneously and 15 min later, a second CRD was performed. Data are expressed as mean ± SEM of VMR expressed as % of 60 mmHg response to the first CRD.

Figure 4

The analgesic buprenorphine inhibited the VMR to ascending CRD monitored with ICP in naïve mice. A first set of ascending phasic CRD was performed and 45 min later, buprenorphine (0.3 mg/kg) or saline was injected i.p. and after 15 min, a second CRD was performed. Data are expressed as mean ± SEM, of 9–10 mice per group. *P < 0.05 compared with baseline (pooled groups of the first CRD) or i.p. vehicle.

Figure 5

Influence of repeated intermittent WAS on BW and defecation in mice subjected to EMG electrode implantation and SH post surgery (S-SH) compared with mice having no surgery and group-housed (NS-GH) or having no surgery and single-housed (NS-SH). (A) Daily BW changes in S-SH, NS-SH and NS-GH mice exposed daily to 1 h of WAS for 10 days. Each point represents the mean ± SEM for n = 13–18 mice. (B) S-SH mice exhibit an increased average defecation in response to repeated WAS compared to NS-GH and NS-SH mice. In contrast, S-SH, NS-GH and NS-SH mice when monitored in non-stressful conditions (basal non-stressful defecation in home cage for 1 h following a quick bedding change) exhibit similar defecation rates. Each bar represents the mean ± SEM of number of mice indicated in the bars; *P < 0.05 vs. S-SH control group, ⁺P < 0.05 vs. all other groups. (C) Time course of defecation response to 1-h WAS in NS-GH, S-SH and NS-SH mice. Each point represents the mean ± SEM for n = 13–18 mice. The grey band represents the mean ± SEM basal defecation of mice under non-stressful conditions during 10 days (pooled values of NS-GH, n = 4; S-SH, n = 5 and NS-SH, n = 13). *P < 0.05 vs. FPO at day 1 for respective groups, ^#P < 0.05 vs. mean basal non-stressful defecation, ⁺P < 0.05 compared with NS-GH + WAS group, ^$P < 0.05 vs. NS-SH + WAS group.

Figure 6

Repeated WAS alters differently the visceral sensitivity to CRD in S-SH, NS-GH, or NS-SH mice. (A) S-SH mice exhibit an increased VMR to CRD at 60 mmHg compared to baseline. (B) NS-GH mice exhibit a decreased VMR to CRD at 30, 45, and 60 mmHg compared to baseline. (C) NS-SH mice show no significant changes in VMR post WAS compared to baseline. (D) S-SH mice exhibit an increased VMR to CRD at 30, 45, and 60 mmHg vs. NS-GH mice as monitored 24 h after the last session of repeated WAS (1 h/day for 10 days). Data are expressed as mean ± SEM, n = 10–18 per group as given in graphs. *P < 0.05 vs. baseline, ⁺P < 0.05 vs. S-SH mice.