Cognitive and neuroinflammatory consequences of mild repeated stress are exacerbated in aged mice

Abstract

Peripheral immune stimulation as well as certain types of psychological stress increases brain levels of inflammatory cytokines such as interleukin-1beta (IL-1beta), IL-6 and tumor necrosis factor alpha (TNFalpha). We have demonstrated that aged mice show greater increases in central inflammatory cytokines, as well as greater cognitive deficits, compared to adults in response to peripheral lipopolysaccharide (LPS) administration. Because aged mice are typically more sensitive to systemic stressors such as LPS, and certain psychological stressors induce physiological responses similar to those that follow LPS, we hypothesized that aged mice would be more sensitive to the physiological and cognitive effects of mild stress than adult mice. Here, adult (3-5 months) and aged (22-23 months) male BALB/c mice were trained in the Morris water maze for 5 days. Mice were then exposed to a mild restraint stress of 30 min before being tested in a working memory version of the water maze over a 3-day period. On day 4 mice were stressed and then killed for collection of blood and brain. In a separate group of animals, mice were killed immediately after one, two or three 30 min restraint sessions and blood was collected for peripheral corticosterone and cytokine protein measurement, and brains were dissected for central cytokine mRNA measurement. Stress disrupted spatial working memory in both adult and aged mice but to a much greater extent in the aged mice. In addition, aged mice showed an increase in stress-induced expression of hippocampal IL-1beta mRNA and MHC class II protein compared to non-stressed controls while expression in adult mice was unaffected by stress. These data show that aged mice are more sensitive to both the cognitive and inflammatory effects of mild stress than are adult mice and suggest a possible role for IL-1beta.

Figure 1

Change in body weight in adult (n=20/group) and aged (n=12/group) mice over 4 days of 30 min restraint stress. ^**p<0.01, ^*** p<0.001 different from non-stressed controls. ⁺⁺p<0.01, ⁺⁺⁺ p<0.001 different from adult stressed mice.

Figure 2

Distance swam (D), latency to platform (E) and swim speed (F) in adult and aged mice in the water maze over 5d acquisition (left panels) and 3 days of stress (right panels). Mice were trained in the water maze for 5d with a static platform position. During testing, mice were exposed to a 30 min restraint stress and then tested in the water maze for 3 days. Left panel: means with different letters are at least p<0.05 different from each other. Right panel: ^* p<0.05, ^**p<0.01 different from non-stressed controls. ⁺ p<0.05, ⁺⁺ p<0.01, ⁺⁺⁺ p<0.001, different from adult stressed mice.

Figure 3

Inflammatory cytokine mRNA in the hippocampus (A) and hypothalamus (B), hippocampal MHC II mRNA (C) and plasma CORT (D) after stress in adult and aged mice that swam in the water maze. Mice were stressed and then tested in the water maze over 3 days. On the fourth day, mice were stressed a final time and then immediately killed for collection of blood and brain. Means with different letters are different from each other (p<0.05).

Figure 4

Correlation of IL-1β mRNA and Distance Swam in Aged (A) and Adult (B) stressed and non-stressed mice.

Figure 5

Hippocampal IL-1β (A), TNFα (B) and MHC II (C) mRNA, and hypothalamic IL-1β (D) and TNFα (E) mRNA, and plasma CORT (F) in adult and aged mice after acute and repeated stress (n=4–6 per group). Adult and aged mice were exposed to 1, 2 or 3 30 min restraint stress sessions and immediately killed for collection of blood and brain. Means with different letters are different from each other (p<0.05).

Figure 6

Immunohistochemical staining for MHC class II in the dentate gyrus of adult and aged, stressed and non-stressed mice at 40x magnification. Adult and aged mice were exposed to a single 30 min restraint stress session and then immediately after stress, brains were collected. Pictures are representative photomicrophraphs of the dentate gyrus of an adult and aged mouse exposed to stress or control. Arrows point to examples of positive MHC II staining. Scale bar = 25µm.