Effects of Lactobacillus helveticus on murine behavior are dependent on diet and genotype and correlate with alterations in the gut microbiome
Introduction
The human intestinal tract is a multifaceted environment where up to 1000 different species of bacteria both influence and are affected by the intestinal epithelium and underlying gut immune system (Ashida et al., 2012). Gut microbiota signals through numerous mechanism to impact systemic and intestinal inflammatory states (Madsen et al., 2002a, Madsen et al., 2002b, Mencarelli et al., 2011, Yan et al., 2007), and metabolism (Murphy et al., 2012, Turnbaugh et al., 2008, Turnbaugh et al., 2006). A new paradigm has emerged in the literature suggesting that microbial dysbiosis in the gut may drive diseases in genetically susceptible individuals, including inflammatory bowel disease (IBD) and irritable bowel syndrome (Albenberg et al., 2012, Xavier and Podolsky, 2007). Studies have shown that gut microbes can influence anxiety, depression and memory (Agrawal and Gomez-Pinilla, 2012, Bravo et al., 2011, Li et al., 2009, Maniam and Morris, 2010) through modulation of the gut–brain axis via a number of mechanisms, including the production of various metabolites and neuroactive compounds (Grenham et al., 2011). As depression and anxiety are often associated with gut diseases, (Bercik et al., 2010, Goodhand et al., 2012), understanding the pathways that gut microbes influence to alter brain function and how probiotic effects may be modulated by the presence of active inflammation may help in the design of oral therapies to alleviate these conditions.
Probiotics are microbial organisms that are administered in supplements or foods to enhance the well-being of the host. In numerous studies, live microbes have been shown to be required for probiotic beneficial effects and this may be related to either the release of bioactive molecules or fermentation processes (Anastasovska et al., 2012, Grenham et al., 2011, Wagar et al., 2009). The types of fermentation products that are produced by microbes are dependent upon the substrate available; thus, altering diet can have significant effects on gut fermentation. Furthermore, dietary components can directly affect the intestinal epithelium or underlying immune system during digestion and absorption, which could lead to altered behavior independently of changes in the microbiota. Moreover, the role of the diet in contributing to the effects of probiotics has not been examined, and may significantly affect the ability of specific probiotics to have their beneficial effects.
Lactobacillus helveticus R0052 is a probiotic that was originally isolated from a dairy culture and which has been extensively characterized for its in vitro and in vivo effects in a variety of biological systems (Foster et al., 2011). This strain has been shown to elicit anti-inflammatory responses and to reduce stress in a number of animal models and, in combination with Bifidobacterium longum, also in human studies (Foster et al., 2011, Messaoudi et al., 2011b). The interleukin-10 knockout (IL-10−/−) mouse is a commonly used model of spontaneous, microbial-induced colonic inflammation similar to IBD (Kuhn et al., 1993). This mouse strain remains healthy when kept sterile, but develops severe colitis within 10 weeks in the presence of a normal colonic microbiota (Madsen et al., 2002a, Madsen et al., 2002b). Lactobacillus species, either alone or in combination with other probiotics, have been shown to attenuate intestinal inflammation in IL-10−/− mice (Cazzola et al., 2010, Madsen et al., 1999, Mencarelli et al., 2011, Xia et al., 2011). The aim of this study was to determine if altering the diet of wild-type or IL-10−/− mice to a Western-style diet (high fat, refined sugar) would influence the host response to a probiotic.
Section snippets
Animal model
Animal studies were conducted in accordance with the Canadian Council on Animal Care Guidelines and Policies with approval from the Animal Care and Use Committee at the University of Alberta. Interleukin-10 deficient (IL-10−/−) mice on a 129/SvEv background and wild-type controls were housed under specific pathogen-free conditions and allowed free access to water and food. Mice were fed standard lab diet 5001 chow (29% protein, 55% carbohydrates, 13% fat; 3.8 kCal/g; PMI Nutrition International,
Western diet resulted in an increased weight gain
Wild-type (WT) and IL-10−/− mice ate equivalent calories on the mouse chow and Western diet during the treatment period (Supplementary Table 1), but weight gain was increased in mice fed Western diet (Supplementary Fig. 1). This weight gain in the Western-diet fed mice was reduced by L. helveticus transiently over days 7–14 in WT mice and for the entire study period in IL-10−/− mice (Supplementary Fig. 1). L. helveticus did not alter weight gain in chow-fed mice.
Diet and L. helveticus interact to modulate memory and anxiety-like behavior
The Barnes maze is
Discussion
In this study we demonstrate that a Western-style diet high in fat and refined sugar increases anxiety-like behavior and decreases memory function in mice with underlying low grade inflammation. Effects of L. helveticus on gut morphology and host behavior were dependent upon the diet of the mouse and gut inflammatory state, and correlated with specific changes in the gut microbiota.
While it has been shown that both diet and probiotic bacteria can modulate anxiety and behavior in mice. (Li et
Role of funding source
These studies were supported by the Crohn's and Colitis Foundation of Canada, Canadian Institutes for Health Research, Alberta Innovates, and the Alberta IBD Consortium. The funding sources had no input into the design or execution of these studies.
Contributors
KM and NH designed the study. NH, LK, HB, EP, CO performed the experiments. AT performed the histological analysis. CO and EP undertook the statistical analysis. KM and CO wrote the manuscript. All authors contributed to and approved the final manuscript.
Conflict of interest
The authors all declare no conflict of interest.
Acknowledgements
These studies were supported by the Crohn's and Colitis Foundation of Canada, Canadian Institutes for Health Research, Alberta Innovates, and the Alberta IBD Consortium. The authors would like to thank Matt Emberg and Nic Moore for their technical assistance with these studies.
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