Dietary n-3 PUFAs Deficiency Increases Vulnerability to Inflammation-Induced Spatial Memory Impairment

Abstract

Dietary n-3 polyunsaturated fatty acids (PUFAs) are critical components of inflammatory response and memory impairment. However, the mechanisms underlying the sensitizing effects of low n-3 PUFAs in the brain for the development of memory impairment following inflammation are still poorly understood. In this study, we examined how a 2-month n-3 PUFAs deficiency from pre-puberty to adulthood could increase vulnerability to the effect of inflammatory event on spatial memory in mice. Mice were given diets balanced or deficient in n-3 PUFAs for a 2-month period starting at post-natal day 21, followed by a peripheral administration of lipopolysaccharide (LPS), a bacterial endotoxin, at adulthood. We first showed that spatial memory performance was altered after LPS challenge only in n-3 PUFA-deficient mice that displayed lower n-3/n-6 PUFA ratio in the hippocampus. Importantly, long-term depression (LTD), but not long-term potentiation (LTP) was impaired in the hippocampus of LPS-treated n-3 PUFA-deficient mice. Proinflammatory cytokine levels were increased in the plasma of both n-3 PUFA-deficient and n-3 PUFA-balanced mice. However, only n-3 PUFA-balanced mice showed an increase in cytokine expression in the hippocampus in response to LPS. In addition, n-3 PUFA-deficient mice displayed higher glucocorticoid levels in response to LPS as compared with n-3 PUFA-balanced mice. These results indicate a role for n-3 PUFA imbalance in the sensitization of the hippocampal synaptic plasticity to inflammatory stimuli, which is likely to contribute to spatial memory impairment.

Figure 1

LPS treatment induces spatial memory impairment in n-3 PUFA-deficient mice. Spatial memory was assessed in the Y-maze paradigm for n-3 PUFA-deficient and n-3 PUFA-balanced mice 24 h after saline or LPS treatment. The graph shows the time spent (in seconds) in the novel or the familiar arm after a 30-min ITI. Only n-3 PUFA-deficient mice injected with LPS exhibited a random exploration of the two arms, as a marker of memory impairment. Data are expressed as mean±SEM (**p<0.01). N-3 PUFA-deficient/saline: n=9, n-3 PUFA-deficient/LPS: n=10, n-3 PUFA-balanced/saline n=10, n-3 PUFA-balanced/LPS n=8.

Figure 2

LPS treatment impairs LTD in the hippocampus of n-3 PUFA-deficient mice and decreases spine density of hippocampal neurons in both n-3 PUFA-balanced and n-3 PUFA-deficient mice. (a) High frequency stimulation (HFS) of CA1 stratum radiatum layer induced a long-term potentiation (LTP) of evoked fEPSP recorded in CA1 stratum radiatum layer, in n-3 PUFA-balanced mice injected with either saline (n=13 slices in 10 mice, white symbols) or LPS (n=16 slices in 14 mice, grey symbols), as well as in n-3 PUFA-deficient mice injected with either saline (n=18 slices in 16 mice, black symbols) or LPS (n=16 slices in 14 mice, grey symbols). Average time courses of mean fEPSP were normalized to baseline. Low frequency stimulation (LFS) of CA1 stratum radiatum layer induced a long-term depression (LTD) of evoked fEPSP recorded in CA1 stratum radiatum layer, in n-3 PUFA-balanced mice injected with either saline (n=13 slices in 12 mice, white symbols) or LPS (n=8 slices in 8 mice, grey symbols), as well as in n-3 PUFA-deficient mice injected with saline (n=7 slices in 7 mice, black symbols). However, LTD was impaired in LPS-treated n-3 PUFA-deficient mice (n=7 slices in 7 mice, grey symbols). Average time courses of mean fEPSP were normalized to baseline (*p<0.05). Last panel represents fEPSP (as a percentage of the baseline) 45 min after the protocol of stimulation. Data are expressed as mean±SEM *p<0.05, **p<0.01, paired t-test between baseline and mean normalized fEPSP 45 min post induction); (b) Left panel: representative images of Golgi staining from all conditions, scale bar=10 μm. Right panel: Quantification of the number of spines on dendritic trees of pyramidal neurons of the hippocampus (CA1 region). Data are presented as the number of spines per 10 μm. The first two graphs show the average number of spines on the apical and basal dendrites respectively. N-3 PUFA-deficient/saline: n=3, n-3 PUFA-deficient/LPS: n= 3, n-3 PUFA-balanced/saline n=3, n-3 PUFA-balanced/LPS n=5.

Figure 3

N-3 PUFA deficiency alters LPS-induced hippocampal production of cytokines. (a) Kinetics of expression of IL-1β, IL-6, TNF-α, and IL-10 mRNA in response to saline or LPS injection (ratio LPS/saline, arbitrary unit). Data are expressed as mean±SEM (n-3 PUFA-deficient vs n-3 PUFA-balanced mice, *p<0.05; ***p<0.001). n=4–6 mice per goup; (b) Kinetics of expression of CXCL10 mRNA in response to saline or LPS injection (ratio LPS/saline, arbitrary unit). Data are expressed as mean±SEM (n-3 PUFA-deficient vs n-3 PUFA-balanced mice, ***p<0.001). n=4–6 mice per goup. (c) GPR 84 mRNA expression in the hippocampus 24 h post saline or LPS injection. Data are expressed as mean±SEM (treatment effect, **p=0.0022). n=4–5 mice per group.

Figure 4

LPS treatment similarly induces sickness behavior and cytokine release, but not corticosterone, in the plasma of n-3 PUFA balanced and n-3 PUFA-deficient mice. (a) Plasma concentration of the pro-inflammatory cytokines IL-1β, IL-6, and TNF-α and the anti-inflammatory cytokine IL-10 in response to saline or LPS injection (in pg/ml). Measures were performed 2, 6, 12, and 24 h post treatment. Data are expressed as mean±SEM. * n-3 PUFA-balanced/saline vs n-3 PUFA-balanced/LPS; ^$ n-3 PUFA-deficient/saline vs n-3 PUFA-deficient/LPS, ^# n-3 PUFA-deficient/LPS vs n-3 PUFA-balanced/LPS (*^,#p<0.05; ^$$,##p<0.01; ***^{,$$$, ###}p<0.001). n=5–6 mice per group; (b) Weight loss and food intake of mice fed an n-3 PUFA-deficient or n-3 PUFA-balanced diet 24 h after saline or LPS treatment. All mice developed a sickness behavior of same amplitude in response to LPS injection. Data are expressed as mean±SEM (***p<0.001). n=6 per group; (c) Left panel: Plasma concentration of corticosterone in response to saline or LPS in n-3 PUFA-deficient and n-3 PUFA-balanced animals (hormonal concentration expressed as ng/ml). Measures were performed 2, 6, 12, and 24 h post treatment. Data are expressed as mean±SEM. * n-3 PUFA-balanced/saline vs n-3 PUFA-balanced/LPS; ^$ n-3 PUFA-deficient/saline vs n-3 PUFA-deficient/LPS, ^# n-3 PUFA-deficient/LPS vs n-3 PUFA-balanced LPS, ^£ n-3 PUFA-deficient/saline vs n-3 PUFA-balanced/saline (^£p<0.05; ***^{,$$$, ###}p<0.001); Right panel: LPS/saline ratio of corticosterone concentration (^***p<0.001). n=5 mice per group.