Timing matters: the interval between acute stressors within chronic mild stress modifies behavioral and physiologic stress responses in male rats

Abstract

Chronic mild stress can lead to negative health outcomes. Frequency, duration, and intensity of acute stressors can affect health-related processes. We tested whether the temporal pattern of daily acute stressors (clustered or dispersed across the day) affects depression-related physiology. We used a rodent model to keep stressor frequency, duration, and intensity constant, and experimentally manipulated the temporal pattern of acute stressors delivered during the active phase of the day. Adult male Sprague-Dawley rats were exposed to one of three chronic mild stress groups: Clustered: stressors that occurred within 1 hour of each other (n = 21), Dispersed: stressors that were spread out across the active phase (n = 21), and Control: no stressors presented (n = 21). Acute mild stressors included noise, strobe lights, novel cage, cage tilt, wet bedding, and water immersion. Depression-related outcomes included: sucrose preference, body weight, circulating glucocorticoid (corticosterone) concentration after a novel acute stressor and during basal morning and evening times, and endotoxin-induced circulating interleukin-6 concentrations. Compared to control rats, those in the Clustered group gained less weight, consumed less sucrose, had a blunted acute corticosterone response, and an accentuated acute interleukin-6 response. Rats in the Dispersed group had an attenuated corticosterone decline during the active period and after an acute stressor compared to the Control group. During a chronic mild stress experience, the temporal distribution of daily acute stressors affected health-related physiologic processes. Regular exposure to daily stressors in rapid succession may predict more depression-related symptoms, whereas exposure to stressors dispersed throughout the day may predict diminished glucocorticoid negative feedback.

Keywords: Acute pro-inflammatory response; acute stressor dynamics; chronic mild stress; circulating glucocorticoid dynamics; depressive behavior; temporal pattern.

Figure 1.

Study timeline. Order of events during the three phases of the study, relative to the chronic mild stress (CMS) protocol: Pre-CMS, CMS, post-CMS. During pre-CMS, all rats were handled and tested for sucrose preference (SPT; 68–69 days of age). During CMS, rats were exposed to one of three conditions (Control, Dispersed, Clustered), with the latter two groups exposed to three daily 15 min stressors (76–102 days of age). To measure alterations in circadian glucocorticoid regulation, basal serum corticosterone concentration was measured in the morning and evening on the last day of CMS (102 days of age). Post-CMS physiology and behavior were measured in the following order: sucrose preference test (102–103 days of age), innate immune response to lipopolysaccharide (104 days of age), and acute corticosterone response to restraint (107 days of age).

Figure 2.

Body weight gain during chronic mild stress (CMS). Mean body weights (g) from Day 1 to Day 27 in the three stress conditions. Estimated marginal mean and SEM are presented. The number of rats in each group at each time point was 21. *On Day 27, Clustered rat body weights were significantly lower than Control rats (planned pair-wise comparisons, p < .05).

Figure 3.

Sucrose preference at end of chronic mild stress (CMS). Percent sucrose solution consumed over a 24 h period during the sucrose preference test administered on the final day of CMS. Estimated marginal means, which control for pre-CMS sucrose consumption, are shown. Error bars represent SEM and the number of rats per group are in the bars. Clustered rats consumed significantly less sucrose than Control rats (planned pair-wise comparisons, p < .05).

Figure 4.

Circulating glucocorticoid concentration at end and after chronic mild stress (CMS). (a) Mean decline in basal circulating corticosterone concentration during the active period of the last day of CMS for all three groups. Dispersed rats had an attenuated decrease in corticosterone during the active period of the day compared to Control rats (planned pairwise comparisons, p < .05). (b) Corticosterone reactivity to 15 min restraint: Mean change in circulating corticosterone concentration from baseline to 30 min after the onset of 15 min restraint for all three groups. *Clustered rat corticosterone reactivity was significantly attenuated compared to Control rats (planned pair-wise comparisons, p < .05). (c) Corticosterone concentration during recovery phase: Mean circulating corticosterone concentration 105 min after onset of 15-min restraint for all three groups. *Dispersed rats had significantly higher corticosterone concentrations during recovery than Control rats (planned pair-wise comparisons, p < .05). Across all figures, error bars indicate SEM and the number of rats per group are in the bars.

Figure 5.

Circulating interleukin-6 (IL-6) response to lipopolysaccharide (LPS) 2 days after chronic mild stress (CMS). Mean IL-6 concentrations for each group at 0 and 2 h after LPS injection. Error bars indicate SEM. Sample size for each group is in the key. Clustered rats had a significant greater acute IL-6 response to LPS than Control rats (planned pair-wise comparisons, p < .05).