Neuroinflammation and disruption in working memory in aged mice after acute stimulation of the peripheral innate immune system

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Abstract

Acute cognitive disorders are common in elderly patients with peripheral infections but it is not clear why. Here, we injected old and young mice with Escherichia coli lipopolysaccharide (LPS) to mimic an acute peripheral infection and separated the hippocampal neuronal cell layers from the surrounding hippocampal tissue by laser capture microdissection and measured mRNA for several inflammatory cytokines (IL-1β, IL-6, and TNFα) that are known to disrupt cognition. The results showed that old mice had an increased inflammatory response in the hippocampus after LPS compared to younger cohorts. Immunohistochemistry further showed more microglial cells in the hippocampus of old mice compared to young adults, and that more IL-1β-positive cells were present in the dentate gyrus and in the CA1, CA2, and CA3 regions of LPS-treated old mice compared to young adults. In a test of cognition that required animals to effectively integrate new information with a preexisting schema to complete a spatial task, we found that hippocampal processing is more easily disrupted in old animals than in younger ones when the peripheral innate immune system is stimulated. Collectively, the results suggest that aging can facilitate neurobehavioral complications associated with peripheral infections probably by allowing the over expression of inflammatory cytokines in brain areas that mediate cognitive processing.

Introduction

Old age is associated with increased brain inflammation (Ye and Johnson, 1999, Lee et al., 2000, Richwine et al., 2005) and stimulation of the peripheral innate immune system with lipopolysaccharide (LPS) leads to an increased inflammatory response in the brain of healthy aged mice (Godbout et al., 2005). This is noteworthy because an increased inflammatory response in the aged brain may cause cognitive deficits that are highly prevalent in elderly patients. For example, acute cognitive impairment (i.e., delirium) is common in elderly emergency department patients (Wofford et al., 1996a, Chiovenda et al., 2002) and frequently occurs in patients with infections unrelated to the CNS (Wofford et al., 1996a). Cognitive impairment complicates diagnosis and is associated with reductions in self-care, increased hospitalization and delayed recovery (Johnston et al., 1987). The severity of dementia is related to mortality of infectious disease, indicating an insidious relationship between infection and cognition, whereby infection in the elderly induces cognitive impairment, and cognitive impairment exacerbates the infection.

Because of the dysregulated linkage between the peripheral immune system and brain in old mice we hypothesized that elevated levels of inflammatory cytokines in the hippocampi of aged animals would be associated with greater deficits in hippocampal-mediated learning and memory after peripheral immune activation. We focused on the hippocampus because it is sensitive to the insults of aging and is likely involved in acute cognitive disorders that are evident in elderly patients with systemic infections. Further, inflammatory cytokines can impair synaptic plasticity in the dentate gyrus (DG) and CA regions of the hippocampus (Bellinger et al., 1993, Cunningham et al., 1996, Tancredi et al., 2000, Hellstrom et al., 2005, Shaw et al., 2005, Perry et al., 2007) as well as disrupt hippocampal-dependent learning and memory (Palin et al., 2004, Rosi et al., 2005, Tanaka et al., 2006). A recent study reported that older rats infected with Escherichia coli had increased hippocampal IL-1β and deficits in hippocampal-dependent memory and learning that were not evident in similarly infected younger cohorts (Barrientos et al., 2006).

In the present study, using a mouse model and a working memory version of the water maze, we found that hippocampal processing is more easily disrupted in old animals than in younger ones after peripheral immune activation. This outcome is independent of a generalized effect on memory consolidation (Sparkman et al., 2005a, Barrientos et al., 2006), and demonstrates an important and novel aspect of infection-related cognitive impairments that is consistent with previous studies of hippocampal dysfunction (Squire, 1992, Avital et al., 2003). Furthermore, using laser capture microdissection (LCM) and immunohistochemistry (IHC), we showed the LPS-induced disruption in working memory in old mice was associated with increased expression of inflammatory cytokines in the hippocampal neuronal cell layers (DG and CA regions) and in surrounding hippocampal tissue. The results suggest that aging can influence the neurobehavioral complications associated with systemic infections probably by allowing the over expression of inflammatory cytokines in brain areas that mediate cognitive processing.

Section snippets

Animals

Young (3–6 months old) and old (22–24 months old) male BALB/c mice from our in-house specific-pathogen-free colony were used. Mice were housed in polypropylene cages and maintained at 23 °C under a reverse phase 12 h light–dark cycle with ad libitum access to water and rodent chow. At the end of each study, mice were examined postmortem for gross signs of disease (e.g. splenomeglia and tumors). Data from mice determined to be unhealthy were excluded from analysis. All procedures were in

Results

Young (3–6 months old) and old (22–24 month old) mice were injected i.p. with saline or LPS and 4 h later they were killed and mRNA encoding several inflammatory cytokines was measured in their hippocampi by real-time PCR. LCM was used to separate the neuronal cell layers (i.e., DG and CA regions) from surrounding hippocampal tissue, thus providing a better analysis of the microenvironment of hippocampal neurons which are known to be sensitive to insults of aging and inflammatory cytokines. Fig.

Discussion

The present study demonstrates that old mice have more microglia that stain for tomato lectin and higher basal levels of several inflammatory cytokine mRNAs in the hippocampus than do young adults. Furthermore, when the peripheral innate immune system was stimulated by LPS to mimic an acute Gram-negative bacterial infection, the expression of inflammatory cytokines in the hippocampus was higher in old mice than in young ones. Using a working memory version of the water maze that required

Acknowledgments

We thank Dr. James Zachary for assisting with LCM and Dr. Masaaki Nakai (University of Illinois) for helping with IHC staining. Dr. Colm Cunningham at University of Southampton (U.K.) also provided helpful advice on IHC staining. This research was supported by NIH grants AG16710 (R.W.J.), AG023580 (R.W.J.), MH069148 (R.W.J.) and DK64862 (G.G.F.). J.B.B. and J.P.G. were supported by a Ruth L. Kirchstein NRSA Postdoctoral Fellowship.

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    Please see Brief Commnetary by Marina Lynch on page 299 of this issue.

    1

    Current address: Department of Molecular Virology, Immunology and Medical Genetics, The Ohio State University, Columbus, OH 43210, USA.

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