Increased resistance of immobilized-stressed mice to infection: Correlation with behavioral alterations
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► Chronic immobilization stress lowers ability to resist bacterial infection, however better bacterial clearance was observed with improved behavior during acute stress.
Introduction
Stress has been defined as a constellation of events comprised of two major components – a stimulus (stressor) that precipitates a reaction in the brain and perception that activates the physiological fight or flight systems in the body (stress response) (Johnson et al., 1992). Stress has been classically categorized as acute stress (defined as stress that lasts for minutes to hours), and chronic stress (lasting for months to years) (Dhabhar and McEwen, 1997). Inescapable stress, commonly called restraint stress or immobilization stress (IS) is an easy and convenient method to induce both psychological (escape reaction) and physical stress (muscle work), resulting in restricted motility and aggression and is believed to be the most severe type of stress in rodent models that is comparable to humans (Romanova et al., 1994, Singh et al., 1999).
Small bouts of stress have been shown to enhance the host’s immune response. However, prolonged periods of stress were found to be detrimental through excess production of neuroendocrine mediators that could dampen the immune responses to invasive pathogens (Radek, 2010). It has been reported that IS resulted in profound depressive behavior in adult rats; but surprisingly those rats that suffered a bacterial infection early in life, had blunted corticosterone responses to the stressor and were remarkably protected from depressive symptoms compared to controls (Bilbo et al., 2008). Although it has been generally accepted that chronic stress often results in immunosuppression while acute stress enhances immune responses (Dhabhar, 2000, Dhabhar and McEwen, 1999), the effects of IS on the host resistance to microbial infection and associated behavioral changes, are not well elucidated. Previous studies have showed that acute stress enhances cutaneous immunity (contact sensitivity) but suppresses systemic cellular immunity (Blecha et al., 1982a, Blecha et al., 1982b, Dhabhar and McEwen, 1996). Various studies have reported that IS exacerbates the development and or progression of many diseases, particularly during adolescence, by mechanisms yet unknown. Moreover, IS has been associated with an increased incidence of infectious diseases, indicating that immunosuppressive attributes of stress translate into significant adverse health effects (Glaser and Kiecolt-Glaser, 2005, Segerstrom and Miller, 2004). IS reportedly also enhances the resistance of mice to sepsis (Wang et al., 2008).
Various studies from time to time have provided inputs on the molecular signaling that links stress-induced neuro-endocrine-immune interactions, suggesting a bi-directional crosstalk which is based on the secretion of cytokines, glucocorticoids, neurotransmitters and neuropeptides (Sternberg, 2006). Studies have revealed that stressed animals had greater mortality from a variety of infections than non-stressed animals and the effects were more prominent when the stressor was administered at the time of microbial challenge (Black, 1994). Data from rodent studies show that stress diminishes vaccine responses, exacerbate viral and bacterial pathogenesis, slows wound healing and alters autoimmune diseases (McCabe et al., 2000, Padgett and Glaser, 2003). It has been reported that IS increases iNOS, COX-2 activities and expression and produces an accumulation of lipid peroxidation products (Madrigal et al., 2002, Madrigal et al., 2003). Also, IS-induced glutathione dysregulation has been reported to be associated with the etiology and progression of several neurotoxic and neurodegenerative diseases (Sayre et al., 2008). Stress-induced corticosteroids have been reported to diminish multiple aspects of the behavioral changes mediated by cytokines, revealing that the behavioral effects in infection and inflammation may depend on stress (Larson and Dunn, 2001). A dysregulation of the cytokine balance could induce depressive symptoms, due to lower levels of anti-inflammatory cytokines and higher levels of proinflammatory cytokines (Dunn, 2006). In a broader perspective, since the immune system is intricately associated with learning, memory, neural plasticity and neurogenesis (Yirmiya and Goshen, 2011), a stress induced disruption could be critical for such processes. The intimate connection between immune and nervous system could also simply and mechanistically explain why stress can influence susceptibility to infection (Fleshner, 2012).
Even though, there is a significant volume of data available regarding IS and its role in infection and neuro-immune interactions, most of the investigations were limited by their choice of relatively benign inflammatory agents such as glycogen, zymosan or killed bacteria. It is unclear, therefore, if the previously reported observations would hold ground if organisms are challenged with living, infectious bacteria. The aim of the current investigation was to test the effect of IS on an in vivo naturalistic model of infection. Viable Escherichia coli (E. coli), a ubiquitous bacteria, was chosen because earlier studies have indicated that a subcutaneous injection E. coli, rather than Staphylococcus aureus, stimulated a more consistent and robust inflammatory response (Campisi et al., 2002). In addition, E. coli is commonly encountered by animals in the wild, and thereby, would serve as a naturalistic infection model.
Section snippets
Animals
Male BALB/c (6–8 weeks) mice were obtained from a registered breeder in our Department and were used for all studies. Mice were housed 4–5 per cage and maintained on a 12 h light:12 h dark cycle (lights on at 08.00 am) in a temperature controlled room (22 ± 2 °C) and food and water were available ad libitum at all times. The experimental protocols were in compliance with the Institutional animal ethics committee (IAEC).
Immobilization stress
Due to the inherent correlation of the circadian rhythm with corticosterone
Effect of acute and chronic E. coli infection on the serum corticosterone concentration in immobilization stressed (IS) mice
The concentrations of corticosterone in mice that were not exposed to stress were very low in the serum. However, exposure to 2 h of restraint stress (acute stress) or to immobilization for longer period of time (chronic stress) significantly increased the serum corticosterone concentrations (P < 0.01). When 2 h immobilized mice were infected with live E. coli (single dose) the corticosterone concentration was decreased significantly in serum than the only stressed mice. However, when chronic
Discussion
Stress can disrupt the homeostasis and balance of organisms and is one of the most important causes of several diseases. Question of whether IS and infection induced biochemical changes in the oxidant status, cytokine levels and cholinergic system were enough to cause alteration in the behavioral responses, has not been well elucidated by previous studies. In this current investigation, we have attempted to answer this question by exploring correlation between IS and infection induced changes
Role of the funding sources
The funding agency had no involvement with the design, execution or data analysis of this study, manuscript preparation, or in the decision to submit the paper for publication.
Contributors
Authors A.B. and B.B. designed the study and designed protocol. S.M., S.C., and A.M. performed all the experiments. K.K. performed the immunoblots. S.M., S.C., A.M. and K.D. managed the literature searches and analyses. S.M. and A.M. undertook the statistical analysis; B.B. and K.D. wrote the manuscript. All authors contributed to and have approved the final manuscript.
Acknowledgments
This study was financially supported by Department of Biotechnology, Ministry of Science and Technology, Government of India, New Delhi under the DBT-Neuroscience scheme. BT/PR1 14196/MED/30/387/2010 September 2011.
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Current address: Department of Psychiatry and Neuroscience, Research Centre of the University Hospital Centre of Quebec, University of Laval, Canada.