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In the mammalian brain, information is believed to be encoded at the cellular level through alterations in synaptic weights. Furthermore, changes in synaptic strength are correlated with structural changes at dendritic spines, such as growth and shrinkage, which may serve to shape inputs into functional domains and increase the computational power of neurons. Neuroanatomical alterations in dendritic spines have been described in humans with intellectual disability, further supporting the relationship between neuronal structure and function. Fragile X Syndrome (FXS) is the most common single-gene neurodevelopmental disorder, and a hallmark feature of this disorder is the increased density of long spines in several brain regions including the hippocampus. Identification of FXS spines as filopodia-like has led to the theory that these spines are immature, and that altered spine development underlies the cognitive dysfunction in this disorder. However, the functional capacity of the long spines observed in FXS is not well understood. For my thesis work, I used two photon imaging, glutamate uncaging and electrophysiology to perform a high-resolution characterization of dendritic spine structure, function, and plasticity in the hippocampus of the FXS mouse model in order to determine what gives rise to these alterations and how this contributes to the observed neuronal dysfunction in this disorder. From my dissertation research, I find that while Fmr1 KO neurons have region-specific alterations in both dendrite and spine morphology, the functional responses of single synapses in FXS mutant neurons are grossly normal. FXS spines respond proportionally to increased levels of glutamate release, and the linear relationship between structure and function is preserved at these synapses. In addition, structural plasticity, both growth and shrinkage, at single inputs is similar in magnitude to control neurons following synaptic potentiation and depression, respectively. However, upon more detailed examination of structural plasticity, either at single or multiple inputs, I find several deficits. First, following structural plasticity, I observe aberrant heterosynaptic plasticity in Fmr1 KO neurons, where unstimulated mutant spines located in close proximity to activated spines become significantly larger compared to neighboring spines in control neurons, which showed no significant change in size. Next, competition for mGluR-LTD does not occur in Fmr1 KO neurons, leading to an increase in spines that undergo spine shrinkage. I conclude from this work that while spine morphology is altered in FXS, spines develop with functional synapses that have the capacity to express bidirectional forms of structural plasticity. However, these spines undergo abnormal structural plasticity across stimulated inputs, leading to the expression of aberrant heterosynaptic structural plasticity. As activity is integrated across a dendritic branch, such excess plasticity observed in Fmr1 KO neurons could contribute to the altered spine morphology as well as cognitive dysfunction observed in FXS.

No description available.

Fragile X syndrome (FXS) is identified by abnormal dendrite morphology and altered synaptic protein expression. Astrocyte secreted factors such as Tenascin C (TNC), may contribute to the synaptic changes, including maturation of the synapse. TNC is a known endogenous ligand of toll-like receptor 4 (TLR4) that has been shown to induce the expression of pro-inflammatory cytokines such as interleukin-6 (IL-6). At the molecular level, elevated IL-6 promotes excitatory synapse formation and increases dendrite spine length. With these molecular changes linked to the phenotype of FXS, we examined the expression and the mechanism of the endogenous TLR4 activator TNC, and its downstream target IL-6 in astrocytes from the FMR1 KO mouse model. Secreted TNC and IL-6 were significantly increased in FMR1 KO astrocytes. Exogenous TNC and lipopolysaccharide (LPS) stimulation of TLR4 induced secreted IL-6, whereas the antagonist of TLR4 (LPS-RS) had an opposing effect. Cortical protein expression of TNC and IL-6 were also significantly elevated in the postnatal FMR1 KO mouse. These results identify TNC as an endogenous ligand of TLR4, capable of effecting IL-6 secretion by astrocytes. In addition, there was an increase in the number of VGLUT1/PSD95 positive synaptic puncta of both WT and FMR1 KO neurons when plated with astrocyte conditioned media from FMR1 KO astrocytes, compared to those plated with media from wild type astrocytes. By assessing the cellular mechanisms involved, a novel therapeutic option could be made available to target abnormalities of synaptic function seen in FXS. / Thesis / Master of Science (MSc) / Autism spectrum disorders (ASDs) are neurodevelopmental disorders which arise from genetic and environmental factors. In the brain, a type of cell called the astrocyte is responsible for proper brain growth and development. Astrocytes release factors that promote inflammation, causing disruption of brain functions that control learning, memory and behaviour. Such factors released by astrocytes are capable of binding to their receptors, in turn impacting downstream targets, which have physiological effects. This research used various biological and genetic techniques to determine if the mechanism of an astrocyte-specific factor called Tenascin C (TNC) is impaired in the Fragile X mouse model. In a normal astrocyte, TNC with its binding partner is able to release molecules responsible for inflammation. Such molecules have been shown to increase the number synapses, where neurons and astrocytes exchange information, to control brain function. This proposed research would be the first to determine a role for TNC in ASDs. By assessing the cellular mechanisms involved between TNC and its binding partner, a novel therapeutic option could be made available in ASDs.

Fragile X Syndrome (FXS) is caused by the instability of a CGG-repeated tract at the 5’ end of the Fmr1 transcript. This instability causes silencing of the gene coding for FMRP. Higher levels of reactive oxygen species, lipid peroxidation, and protein oxidation within brain tissue have been found to be associated with the disease. These imbalances, along with altered levels of components of the glutathione system, provide evidence for increased oxidative stress. Astrocytes, glial cells within the brain, have many functions within neurodevelopment. Specifically, they regulate growth and synaptic contacts of neurons, regulate the level of excitability of synapses, and protect neurons at high levels of activity. To protect neurons from oxidative stress, astrocytes maintain oxidative homeostasis through their mitochondrial electron transport and antioxidant systems. This study examines the relationship between oxidative stress and FXS by assessing mitochondrial function and the antioxidant system of astrocytes. Using the Fmr1 knockout (KO) mouse model, mitochondrial respiration, and reactive oxygen species (ROS) production was analyzed in cultured cortical astrocytes. Astrocytes collected from male and female mice were analyzed under both normoxic and hypoxic conditions. In addition, western blots were conducted on both cortical tissue and cultured cortical astrocytes to determine potential differences in enzyme expression. Results indicate elevations of leak state respiration and ROS production in Fmr1 KO cultured cortical astrocytes alongside alterations in antioxidant and NADPH-oxidase expression. Characterization of mitochondrial function and the antioxidant system of astrocytes will be highly valuable to the understanding of glial roles during brain development and could provide future insight to direct clinically relevant studies of FXS and other neurodevelopment disorders. / Thesis / Master of Science (MSc) / Fragile X Syndrome (FXS) is the most common genetic cause of intellectual disability. It is characterized by the loss of FMRP, an important protein in brain development. Within the FXS brain there is evidence of oxidative stress. The cells that maintain oxidative homeostasis in the brain are astrocytes. Astrocytes are glial cells important for brain development. This thesis evaluated astrocytes' ability to maintain oxidative homeostasis in the FXS cortex. The findings of this thesis provide important insights into our understanding of FXS pathology and will help direct clinically relevant studies of FXS and other neurodevelopmental disorders.

Yes / Group I metabotropic glutamate receptor (mGluR) dependent long-term depression (LTD) is a major form of synaptic plasticity underlying learning and memory. The molecular mechanisms involved in mGluR-LTD have been investigated intensively for the last two decades. In this 60th anniversary special issue article, we review the recent advances in determining the mechanisms that regulate the induction, transduction and expression of mGluR-LTD in the hippocampus, with a focus on the mitogen-activated protein kinase (MAPK) pathways. In particular we discuss the requirement of p38 MAPK and extracellular signal-regulated kinase 1/2 (ERK 1/2) activation. The recent advances in understanding the signaling cascades regulating mGluR-LTD are then related to the cognitive impairments observed in neurological disorders, such as fragile X syndrome and Alzheimer's disease.

The human body is home to complex microbial communities that are fundamental to our physiology. Utilizing mouse models, behavior assays, gene expression analyses, and probiotic interventions, this research explores the intricate relationship between the gut microbiome, the central nervous system, and the immune system. I discuss a strain of Escherichia coli Nissle engineered to produce serotonin, revealing its impact on gut function and immune response. I also evaluate butyrate’s potential to alleviate symptoms of Fragile X Syndrome, highlighting the gut-brain axis. Additionally, I study a strain of Bifidobacterium adolescentis and its role in metabolizing bile acids and modulating host immune cells and stress. Collectively, these studies address the complex interplay between the gut microbiome and host health and behavior, illuminating the therapeutic potential of microbiome manipulation and setting the stage for novel interventions in neurodevelopmental disorders and immune function regulation.

Résumé : Le syndrome du X fragile (SXF) est la première cause de déficience intellectuelle héréditaire et résulte de mutations dans le gène FMR1 menant à l’absence d’expression de la protéine FMRP. Chez la souris déficiente en FMRP (KO-fmr1), des travaux récents ont montré une suractivation de la voie ERK qui jouerait un rôle important dans la physiopathologie du SXF. La lovastatine est un inhibiteur de la HMG-CoA réductase, utilisée pour traiter l’hypercholestérolémie. Parmi ses effets pléiotropes, elle inhiberait également ERK. Nous avons émis l’hypothèse que la lovastatine pourrait compenser en partie l’absence de FMRP et avoir des effets cognitifs positifs chez les individus avec SXF. Objectifs: Évaluer l’efficacité de la lovastatine à réduire le score global de l’ABC-C (Aberrant Behavior Checklist-Community), ainsi que le score de chaque sous-domaine d’ABC-C. Vérifier l’innocuité de la lovastatine pendant le traitement. Méthodes : Une étude prospective, non-randomisée a été réalisée sur une période de 12 semaines. Seize participants avec SXF (âgés de 10 à 31 ans), ont reçu des doses croissantes de la lovastatine (20 mg et 40 mg). Résultats : Des améliorations ont été observées au niveau du comportement aberrant soit le score total de l’ABC-C (p=0.005), ainsi que les sousdomaines: Hyperactivité (p=0.002), Léthargie(p=0.011), Stéréotypie (p=0.025) et Retrait social (p=0.003). Comme attendu, la lovastatine a été bien toléré sans aucun effet indésirable grave observé. Conclusion : Nos résultats suggèrent que la lovastatine peut avoir des effets bénéfices chez les individus avec le SXF. Une étude randomisée, placébocontrôlée est nécessaire pour valider nos trouvailles. / Abstract : Background: Fragile X Syndrome (FXS) results from dynamic mutations leading ultimately to the absence of expression of the Fragile X Mental Retardation Protein (FMRP). It is characterized by synaptic upregulated protein synthesis and immature dendritic spines associated with altered brain plasticity and cognitive functions. Recent work in Fmr1 knockout mice has shown that lovastatin, an inhibitor of Ras-ERK1/2, normalized hippocampus protein synthesis. We hypothesize that lovastatin, as a diseasemodifying drug, would counterweigh the absence of FMRP and improve brain development and learning abilities. Here we report a phase I study to assess the safety and efficacy of lovastatin in individuals with FXS. Methods: A total of 15 subjects (13 males, 6-31 years old) were treated with escalating doses of lovastatin (up to 40 mg) for three months. Their behaviour were assessed before and after treatment using the Aberrant Behavioral Checklist – Community (ABC-C) total score (primary outcome), as well as domains of the FXS validated version of the ABC-C (secondary outcomes). Results: The treatment was well tolerated and minimal adverse effects were reported. Significant improvement in the primary outcome (P < 0.005), as well as in secondary outcomes, were observed in the majority of the subjects (12/15). Conclusions: We believe long-term sustained treatment with diseased-modifying drugs would be necessary in order to improve behaviour and learning. Lovastatin, well known for its long-term security profile, reveals itself as a strong candidate to that purposes. Nevertheless, a larger placebo-controlled trial of lovastatin in this population is warranted in order to confirm its efficacy.

The fragile X mental retardation protein (FMRP) is an important regulator of protein translation, and a lack of FMRP expression leads to a cognitive disorder known as fragile X syndrome (FXS). Clinical symptoms characterizing FXS include learning impairments and heightened anxiety in response to stressful situations. The Fmr1-/y mouse has previously been shown to have deficits in context discrimination and novel object recognition tasks, which primarily rely on the dentate gyrus (DG) region of the hippocampal formation, but not in the Morris water maze (MWM) or the elevated plus-maze tasks, which primarily depend on the Cornu Ammonis (CA1) region. Furthermore, previous research has demonstrated N-methyl-D-aspartate receptor (NMDAR)-associated synaptic plasticity impairments in the DG but not in the CA1. However, the impact of acute stress on synaptic plasticity in the Fmr1-/y hippocampus has not been examined. The current study sought to extend previous behavioural investigations in the Fmr1-/y mouse, as well as examine the impact of stress on activation of the hypothalamic-pituitary-adrenal (HPA)-axis and on hippocampal synaptic plasticity. To further characterize hippocampus-dependent behaviour in this mouse model, the DG-dependent metric change spatial processing and CA1-dependent temporal order discrimination tasks were evaluated. The results reported here support previous findings and demonstrate that Fmr1-/y mice have performance deficits in the DG-dependent task but not in the CA1-dependent task, suggesting that previously reported subregional differences in NMDAR-associated synaptic plasticity deficits in the hippocampus of the Fmr1-/y mouse model may also manifest as selective behavioural deficits in hippocampus-dependent tasks. In addition, following acute stress, mice lacking FMRP showed a faster elevation of the glucocorticoid corticosterone and a more immediate impairment in long-term potentiation (LTP) in the DG. Stress-induced LTP impairments were rescued by administering the glucocorticoid receptor (GR) antagonist RU38486. Administration of RU38486 also enhanced LTP in Fmr1-/y mice in the absence of acute stress to wild-type levels, and this enhancement was blocked by application of the NMDAR antagonist 2-amino-5-phosphonopentanoic acid. These results suggest that a loss of FMRP results in enhanced GR signalling that may adversely affect NMDAR-dependent synaptic plasticity in the DG. Finally, synaptic plasticity alterations reported in this work were found to be specific to the DG and were unidirectional, i.e., restricted to LTP, as NMDAR- and metabotropic glutamate receptor (mGluR)-LTD were both unaffected by acute stress in the DG or the CA1 regions. This study offers new insights into synaptic plasticity impairments in the Fmr1-/y mouse model, and suggests stress and GRs as important contributors to learning and memory deficits in FXS. / Graduate

L’autisme est un trouble envahissant du développement défini uniquement sur des critères comportementaux et l’âge d’apparition. Le X fragile est une pathologie d’origine monogénique dont 15-25% des patients présente le diagnostique complet de l’autisme et dont de nombreux symptômes chevauchent avec l’autisme. Une souris Fmr1 KO a été créée et validée comme modèle pour le X fragile. A l’instar de la variabilité des phénotypes du X fragile chez l’humain, les données préliminaires montrent que la mutation nulle Fmr1 chez la souris interagit avec l'arrière fond génétique. Les travaux présentés visaient à déterminer les caractéristiques autistiques exprimées par les souris Fmr1 KO, ainsi que l’interaction de la mutation nulle avec le fond génétique (souches C57BL/6J, FVB.129P2tm1Cgr /J et leurs hybrides). Les résultats de ces travaux montrent notamment que les souris Fmr1 KO présentent un évitement initial d’approche sociale, des altérations principalement qualitatives des vocalisations, de l’hyperactivité et une augmentation de l’activité diurne. La mutation interagit avec le fond génétique et les résultats actuels indiquent que les KO de fond FVB.129P2tm1Cgr /J ont le phénotype le plus marqué. / Autism is a pervasive developmental disorder defined by behavioural criteria and age of onset. Fragile X is a disorder due to the silencing of the Fmr1 gene. About 15-25% of Fragile X patients are diagnosed as autistic and many symptoms overlap between the two disorders. A mouse Fmr1 KO was created and validated as a model for Fragile X Syndrome. Preliminary data also show that the null mutation interacts with the genetic background. The work presented in this thesis aimed to determine the autistic features expressed in Fmr1 KO mice, as well as the influence of the genetic background (C57BL/6J and FVB.129P2tm1Cgr/J strains, and their reciprocal hybrids) on the expression of the Fmr1 mutation. Our results show an initial inhibition of social approach in Fmr1 KO mice and a qualitative alteration of ultrasonic vocalizations in isolated pups, as well as an increase in activity, especially during the diurnal period. The Fmr1 mutation interacts with the genetic background and the results indicate that KO on the FVB.129P2tm1Cgr/J background show the most marked phenotype.