Pair housing reverses post-stroke depressive behavior in mice

Abstract

Social isolation (SI) has been linked epidemiologically to high rates of morbidity and mortality following stroke. In contrast, strong social support enhances recovery and lowers stroke recurrence. However, the mechanism by which social support influences stroke recovery has not been adequately explored. The goal of this study was to examine the effect of post-stroke pair housing and SI on behavioral phenotypes and chronic functional recovery in mice. Young male mice were paired for 14 days before a 60 min transient middle cerebral artery occlusion (MCAO) or sham surgery and assigned to various housing environments immediately after stroke. Post-stroke mice paired with either a sham or stroke partner showed significantly higher (P<0.05) sociability after MCAO than isolated littermates. Sociability deficits worsened over time in isolated animals. Pair-housed mice showed restored sucrose consumption (P<0.05) and reduced immobility in the tail suspension test compared to isolated cohorts. Pair-housed stroked mice demonstrated significantly reduced cerebral atrophy after 6 weeks (17.5 ± 1.5% in PH versus 40.8 ± 1.3% in SI; P<0.001). Surprisingly, total brain arginase-1, a marker of a M2 "alternatively activated" myeloid cells was higher in isolated mice. However, a more detailed assessment of cellular expression showed a significant increase in the number of microglia that co-labeled with arginase-1 in the peri-infarct region in PH stroke mice compared to SI mice. Pair housing enhances sociability and reduces avolitional and anhedonic behavior. Pair housing reduced serum IL-6 and enhanced peri-infarct microglia arginase-1 expression. Social interaction reduces post-stroke depression and improves functional recovery.

Keywords: Arginase-1; IL-6; Ischemic stroke; Post stroke depression; Sociability; Social isolation.

Fig. 1

Schematic of experimental design and behavioural testing (Cohort 2).

Fig. 2

A significant interaction was found between stroke and housing condition (F (1, 39) = 6.026; P=0.0187) in mice after 6 weeks of stroke (cohort 1). Moreover, there was a significant difference between ISO and PH mice (P=0.0185) (Fig 2 a). b) A two-factor interaction ‘group versus days’ in the repeated measure two-way ANOVA followed by Bonferroni post hoc tests showed an overall significant difference between the PH and SI groups (F 3,24 = 5.413 p< 0.01)

Fig. 3

A significant interaction was found between stroke and housing conditions during sniffing time between test and stranger mice (F 1,21=5.884, P<0.05). However there was no main effect of either stroke or housing condition alone in a two way ANOVA followed by Bonferroni post hoc tests.

Fig. 4

Effect of sucrose consumption in mice a) A two way ANOVA with bonferroni correction for multiple comparisons found no significant interaction between housing condition and stroke in sucrose consumption in cohort1 at 6 weeks after stroke. However, there was a significant difference (P<0.05) between ISO and PH group independent of stroke in the consumption of sucrose. b) In repeated testing of sucrose consumption test a two-factor interaction ‘group versus Days’ in the repeated measure two-way ANOVA followed by Bonferroni post hoc tests showed a significant interaction between groups and days (F18, 102= 1.740, p<0.05). The consumption of sucrose increased progressively in both SI and PH groups.

Fig. 5

Immobility duration was significantly decreased in post stroke PH mice compared to ISO mice (* P<0.05). Interestingly, sham mice pair housed with stroke partner also showed a significant increase in immobility duration (**P<0.01, two-way ANOVA with Bonferroni multiple comparison test). A significant interaction (F1, 21,=13.81, P<0.01) was also found between housing condition and stroke in two way ANOVA.

Fig. 6

(a) Stroke mice paired with healthy partner had significantly less cerebral atrophy. (***P<0.001). (b) Representative coronal section of post stroke ISO and PH brains stained with Cresyl violet.

Fig. 7

Post stroke PH mice regained their initial body weight after 6 weeks. However, isolated mice did not return to their initial weight even after 6 weeks.

Fig. 8

IL-6 levels were measured in serum collected from mice prior to sacrifice. A two way ANOVA test followed by multiple comparisons after bonferroni correction showed no significant difference between ST-PH and ST-ISO group i.e., there was no stroke effect nor any interaction between housing and stroke (F 1, 36 = 0.02826, P = 0.8674). However there was a significant difference (P<0.05) between ISO and PH group regardless of stroke i.e., isolation increased IL-6 levels.

Fig. 9

Representative mosaic image of mouse brain section captured at 10x magnification for GFAP expression at 6 weeks after MCAO. Double immunostaioning of GFAP (red) and DAPI (blue) showed that GFAP⁺ cell were activated at the site of infraction in both stroke groups. Dotted lines show that dense GFAP expression (Glial scar) was confined to the striatal region in ST-PH mice while in strokes-ISO mice it was widely expressed in both striatum and cortex (n=4 per group). SI sham mice also showed higher baseline activity of GFAP.

Fig. 10

Representative mosaic image of mouse brain section (adjacent to the section used for GFAP expression) captured at 10x magnification for Iba-1 expression. Brain section was taken after 6 weeks of stroke. Double immunostaioning of Iba-1 (red) and DAPI (blue) showed that Iba-1⁺ cell were dense and appeared “activated” at the site of infraction in both stroke groups (n=4 per group). Unlike GFAP, Iba-1 expression was not widespread in ipsilateral hemisphere in the ST-ISO group, and was confined to damaged areas as shown by dotted lines.

Fig 11

a) Arginase-1 expression after stroke a) Overall expression of arginase-1 was significantly increased (F 1, 14 = 18.05, P = 0.0008)) in ST-ISO group as compared to other groups in the lysate of ipsilateral brain (n=5 in each group). b) Immunofluorescent staining for Iba1 (red), arginase-1 (green) and DAPI (blue) in the brains of young mice 6 weeks after MCAO (representative of 4 animals). Increased expression of arginase-1 was found in both ST- ISO and ST- PH group as compared to SH-ISO and SH-PH. c) However, higher number of both Iba⁺ and arginase-1⁺ cells ( ) were present in PH house stroke (ST-PH) mice as compared to other groups of mice (*P<0.05 and **P<0.001 vs ST-PH).