Elsevier

Psychoneuroendocrinology

Volume 49, November 2014, Pages 119-129
Psychoneuroendocrinology

Dietary supplementation of omega-3 fatty acids rescues fragile X phenotypes in Fmr1-Ko mice

https://doi.org/10.1016/j.psyneuen.2014.07.002Get rights and content

Summary

Omega-3 polyunsaturated fatty acids (n-3 PUFAs) are known to critically influence brain development and functions. Dietary supplementation with n-3 PUFAs has been suggested as a non-pharmacological therapy for a number of developmental disorders, e.g., autistic spectrum disorders (ASD), but human studies so far have led to conflicting results. Furthermore, it has been hypothesized that the therapeutic impact of n-3 PUFAs on these disorders might be explained by their anti-inflammatory properties and their promoting effects on synaptic function and plasticity, but no clear evidence has been produced in this direction. We evaluated the impact of n-3 PUFA dietary supplementation in a mouse model of fragile X syndrome (FXS), i.e., a major developmental disease and the most frequent monogenic cause of ASD. Fmr1-KO and wild-type mice were provided with a diet enriched or not with n-3 PUFAs from weaning until adulthood when they were tested for multiple FXS-like behaviors. The brain expression of several cytokines and of brain-derived neurotrophic factor (BDNF) was concomitantly assessed as inflammatory and synaptic markers. n-3 PUFA supplementation rescued most of the behavioral abnormalities displayed by Fmr1-KO mice, including alterations in emotionality, social interaction and non-spatial memory, although not their deficits in social recognition and spatial memory. n-3 PUFAs also rescued most of the neuroinflammatory imbalances of KOs, but had a limited impact on their BDNF deficits. These results demonstrate that n-3 PUFAs dietary supplementation, although not a panacea, has a considerable therapeutic value for FXS and potentially for ASD, suggesting a major mediating role of neuroinflammatory mechanisms.

Introduction

Fragile X syndrome (FXS) is a pervasive developmental disorder caused by the lack of FMRP (fragile X mental retardation protein), coded by the FMR1 X-linked gene and playing a pivotal role in synaptic function and plasticity (Verkerk et al., 1991). FXS is characterized by a constellation of behavioral abnormalities, mostly recapitulated by its murine model (The Dutch Belgian Fragile X Consortium, 1994). Fmr1-KO mice indeed exhibit FXS-like behavioral phenotypes, including social and emotional alterations, as well as cognitive deficits (for a review, see Tsiouris and Brown (2004)). This mouse line therefore represents a valuable tool to test novel therapeutic strategies for FXS, since no treatment has been univocally identified to date.

Dietary supplementation of omega-3 polyunsaturated fatty acids (n-3 PUFAs) has been suggested as a potential therapy for a number of neurological and psychiatric pathologies, including some developmental disorders. For example, the administration of n-3 PUFAs seems to attenuate the behavioral abnormalities of autistic (Amminger et al., 2007, Meiri et al., 2009), attention-deficit and hyperactivity disorder (Johnson et al., 2012) and Rett (De Felice et al., 2012) patients who often present low erythrocyte and plasma omega-3/omega-6 ratios (Bell et al., 2010, Young et al., 2004). Since n-3 PUFAs are known to be crucial for brain development and functionality, it is likely that any insufficiency could play a central role in the etiopathology of these and other developmental disorders. Furthermore, n-3 PUFAs exert multiple brain and peripheral effects (Martinez-Victoria and Yago, 2012, Rustan et al., 1997), having anti-inflammatory properties (Kelley and Rudolph, 2000, Laye, 2010) and promoting synaptic function and plasticity (Heinrichs, 2010, Zhang et al., 2011), i.e., acting in two domains that are severely affected in most developmental disorders, including FXS (Ashwood et al., 2010, Marek et al., 2012, Yuskaitis et al., 2010). Yet, to our knowledge the potential link between n-3 PUFAs and FXS has never been investigated.

Despite its attractiveness as a non-pharmacological treatment, with the crucial advantage of high acceptability by children and their families, n-3 PUFAs dietary supplementation still needs more convincing evidence as a potential therapy for developmental disorders, no effect having been reported in some studies (Bent et al., 2011, Gillies et al., 2012, Milte et al., 2013). This weakness may be due to the limitations of human studies, i.e., the high clinical, genetic and environmental variability and the limited size of the samples (Gillies et al., 2012), as well as the difficulty to start the treatment at early clinical phases (Richardson, 2006). All these limitations can be overcome by animal models, offering at the same time the possibility of easily performing neurobiological analyses.

Here we tested whether dietary supplementation with n-3 PUFAs could rescue the FXS-like behavioral phenotype of Fmr1-KO mice and eliminate the KO abnormalities in inflammatory and synaptic function. Fmr1-KOs and their wild type littermates were provided with either a control diet or one enriched with n-3 PUFAs from weaning until adulthood, i.e., from 3 weeks to 3 months of age. This time window was chosen since (i) it covers the interval between the first appearance of FXS-like behavioral abnormalities, i.e., at the juvenile age, and their full expression at adulthood (Pietropaolo et al., 2011), (ii) it includes the developmental phase of adolescence, characterized by marked levels of neurobehavioral plasticity and responsiveness to environmental manipulations (Spear, 2000). Adult mice were then tested for multiple behavioral FXS-like phenotypes, including emotional, social, and cognitive deficits. The expression of selected neuroinflammatory markers (IL-1β, IL-6, IL-10, TNFα, CD11b, CD45) and modulators of neuroplasticity (brain-derived neurotrophic factor, BDNF) were evaluated in cortical and hippocampal areas, i.e., those brain regions where FMRP normally is most abundant (Bakker et al., 2000, Khandjian, 1999). Furthermore, we analyzed some additional parameters linked to energy balance and lipid metabolism to check potential differences in the nutritional impact of dietary enrichment on WT and KO mice, i.e., assessing food intake, body weight, body and erythrocyte fat composition and blood levels of leptin.

Section snippets

Animals

Fmr1-KOs and their wild type littermates (C57BL/6J background) were bred in the animal facility of the University of Bordeaux, as described in details elsewhere (Pietropaolo et al., 2011). At 8 days of age, tail samples were collected for genotype assessment by classical PCR (The Dutch Belgian Fragile X Consortium, 1994). Only males were tested, since only limited behavioral deficits have been reported in female Fmr1-KOs (Qin et al., 2005). A total of 47 male mice were used, including 23 kept

Sociability and preference for social novelty in the three compartments test

No difference in general exploration was observed during the first habituation trial (data not shown).

On trial 2, all groups showed a preference for the social versus the non-social stimulus [area effect: F(1,43) = 45.35, p < 0.0001, Fig. 1A] and this effect did not differ among groups. KO mice showed an overall reduced exploration of both areas containing the novel stimuli and this effect was eliminated by n-3 PUFAs supplementation [genotype × diet: F(1,43) = 7.30, p < 0.01; Fig. 1B].

On trial 3, only WT

Discussion

The present study demonstrated that n-3 PUFAs dietary supplementation improves most of the FXS-like phenotypes displayed by Fmr1-KO mice (Table 2). n-3 PUFAs dietary enrichment was able to eliminate the behavioral abnormalities shown by Fmr1-KO mice, with the exception of the deficits in social recognition and spontaneous alternation. The enriched diet also corrected most of the neuroinflammatory alterations of KO animals, but not the deficits in BDNF brain expression. Furthermore, n-3 PUFAs

Role of the funding source

The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Conflict of interest

None declared.

Acknowledgements

This work was supported by grants from the March of Dimes (12-FY05-1198), CNRS, and the University of Bordeaux to W.E. Crusio, and of the INRA and Région Aquitaine to S. Layé.

We thank Rafael Pineau and his team for expert animal care.

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