Behavioral indices of ongoing pain are largely unchanged in male mice with tissue or nerve injury-induced mechanical hypersensitivity

Abstract

Despite the impact of chronic pain on the quality of life in patients, including changes to affective state and daily life activities, rodent preclinical models rarely address this aspect of chronic pain. To better understand the behavioral consequences of the tissue and nerve injuries typically used to model neuropathic and inflammatory pain in mice, we measured home cage and affective state behaviors in animals with spared nerve injury, chronic constriction injury (CCI), or intraplantar complete Freund's adjuvant. Mechanical hypersensitivity is prominent in each of these conditions and persists for many weeks. Home cage behavior was continuously monitored for 16 days in a system that measures locomotion, feeding, and drinking, and allows for precise analysis of circadian patterns. When monitored after injury, animals with spared nerve injury and complete Freund's adjuvant behaved no differently from controls in any aspect of daily life. Animals with CCI were initially less active, but the difference between CCI and controls disappeared by 2 weeks after injury. Further, in all pain models, there was no change in any measure of affective state. We conclude that in these standard models of persistent pain, despite the development of prolonged hypersensitivity, the mice do not have significantly altered "quality of life." As alteration in daily life activities is the feature that is so disrupted in patients with chronic pain, our results suggest that the models used here do not fully reflect the human conditions and point to a need for development of a murine chronic pain model in which lifestyle changes are manifest.

Figure 1

Mechanical hypersensitivity after nerve injury or inflammation. 50% withdrawal thresholds (in grams) were obtained using the von Frey up-down method in mice (A) after SNI (red bars) or CCI (blue), and in sham (grey) and naïve (white) controls or (B) after saline (grey) or CFA (orange) injection. (C) Mechanical thresholds were normalized to the naïve groups for the SNI and CCI experiments. For all, repeated measures ANOVA showed a significant effect of treatment with Bonferroni post-test differences between naïve/sham and CCI groups indicated with: * <0.05, ** <0.01, *** <0.001; post-hoc differences between naïve/sham and SNI groups: + <0.05, ++ <0.01, +++ <0.001

Figure 2

Home cage movement, feeding and drinking in mice with SNI, CCI and CFA. (A) Total movement (in meters traveled) on day 1 of monitoring was significantly decreased in Balb/c mice with CCI (blue) and sham (grey) compared to naïve (white) controls, but not with either SNI (red) strain or Balb CFA (orange). (B) Average total movement over days 3–17 after injury was also significantly decreased in mice with CCI, but was similar to controls in all other experimental and sham groups. (C) Total daily movement in the dark (i.e. night; a, c, e, g) and light (b, d, f, h) cycles over days 3 to 17 was very similar to controls across the entire period for Balb/c mice with SNI (c, d) and CFA (e, f) and C57Bl/6 with SNI (g, h), but was significantly decreased in the early days after injury in the light cycle (b; repeated measures ANOVA, p=0.024) and for a longer period in the dark cycle (a; repeated measures ANOVA, p=0.002). Post-test differences are all indicated as less than 0.05 for ease of reading, though many are much smaller. Average daily food (D) and water (E) intake were no different in any group. Na: Naïve control; Sh: Sham control; Sa: Saline control. Post-test differences between CCI and Naive: *; differences between Sham and Naïve: +.

Figure 3

Average distance traveled over circadian time. (A) Raster plots of a naïve mouse activity in Balb/c (top) and C57Bl/6 (bottom) mice over 12 days, with green representing periods of locomotor activity; orange, eating; and blue, drinking. (B) Time budgets of naïve, sham and CCI groups of mice in the early and late monitoring periods. Mouse time budgets include inactive (grey), locomotion (green), eating (orange), drinking (blue, less than 1% of the day, so not visible in the graph), and “other” (red, mouse is active, but not engaged in other activities). Inset numbers are percent of time in each activity. (C) Average movement (in distance traveled) binned by hour on days 3–7 (a, c, e) or days 13–17 (b, d, f). SNI Balb/c (c-d) and C57Bl/6 (e-f) animals did not show any differences in their circadian pattern in either the early or late days of the experiment. Average hourly distance traveled in the initial monitoring days by animals with CCI were significantly decreased from controls during the first half of the dark cycle (a; p< 0.0001), but not significantly different in the later days of monitoring (b; p=0.0505).

Figure 4

Short-duration tests of daily activity: (A) Distance traveled in the open field one month after injury was not different in any pain model in Balb/c mice (CCI n=8–9, p=0.99; SNI n=11–12, p=0.12; CFA n=6–7, p=0.07) or (B) Bl/6 mice (CCI n=8–9, p=0.09; SNI n=18–19, p=0.82; CFA n=8–9, p=1.0). (C) Distance traveled in the open field on days 3 and 7 after surgery did not differ among any of the experimental and control groups (n=8, Day 3 p=0.23, Day 7 p=0.97). (D) Mice with SNI or CCI performed as well after injury as before on the rotarod test when previously trained (n=5, p=0.62), (E) but significantly worse 3 days after surgery (n=8, p=0.006), if untrained. (E) Time spent climbing at 3 weeks after injury in Bl/6 mice with SNI or Balb/c mice with CFA did not differ from controls (SNI n=7–8, p=0.82; CFA n=10, p=1.0).

Figure 5

Behavioral measures of affective state at one month after injury. (A) Time spent in the center area of the open field in C57Bl/6 mice (bottom) with CCI (n=8–9, p=0.67), SNI (n=18–19, p=0.69) and CFA (n=8–9, p=0.58) was not different from controls. Balb/c animals (top) with CCI (n=8–9, p=0.71) did not differ from controls, but there was a significant increase in the time spent in the center in Balb/c mice with SNI (n=11–12, p=0.017) and CFA (n=6–7, p=0.073). (B) There was also little difference between experimental groups and controls in the time spent in the open areas of the elevated zero maze, in Balb/c (top) animals (CCI p=0.18, SNI p=0.33, CFA p=0.48) and in C57Bl/6 (bottom) animals (SNI p=0.44, CFA p=0.58) except for those with CCI that spent significantly more time in the open areas (p=0.017). (C) The number of marbles buried on week 5 in Bl/6 SNI animals or Balb CFA animals did not differ from control nor did Balb CCI or Bl/6 SNI differ from their controls at week 7.

Figure 6

Open field behavior in short-term models of sickness and pain. (A-B) Distance traveled in the 30-minute open field test 2 hours after injection was severely decreased in animals injected with LPS, but not in those with intraplantar formalin in either (A) C57Bl/6 or (B) Balb/c animals (n=8/grp). (C-D) Sick animals also spent very little time in the center area compared to saline injected animals in (C) Bl/6 or (D) Balb/c strains. While formalin injected Balb/c animals showed a trend to less time in the center, the difference was not statistically significant (p=0.1). Sal: saline control; Frmln: formalin-injected group.