Effect of dietary zinc supplementation on the gastrointestinal microbiota and host gene expression in the Shank3B−/− mouse model of autism spectrum disorder

AbstractShank genes are implicated in ~1% of people with autism and mice with Shank3 knock out mutations exhibit autism-like behaviours. Zinc deficiency and gastrointestinal problems can be common among people with autism, and zinc is a key element required for SHANK protein function and gut development. In Shank3B−/− mice, a supplementary zinc diet reverses autism behaviours. We hypothesise that dietary zinc may alter the gut microbiome, potentially affecting the gut-microbiome-brain axis, which may contribute to changes in autism-like behaviours. To test this, four types of gastrointestinal samples (ileum, caecum, colon, faecal) were collected from wild-type and knock-out Shank3B−/− mice on either control or supplemented-zinc diets. Cage, genotype and zinc diet each contributed significantly to bacterial community variation (accounting for 12.8%, 3.9% and 2.3% of the variation, respectively). Fungal diversity differed significantly between wild-type and knock-out Shank3B−/− mice on the control zinc diet, and the fungal biota differed among gut locations. RNA-seq analysis of host (mouse) transcripts revealed differential expression of genes involved in host metabolism that may be regulated by the gut microbiota and genes involved in anti-microbial interactions. By utilising the Shank3B−/− knock-out mouse model we were able to examine the influence of – and interactions between – dietary zinc and ASD-linked host genotype. These data broaden understanding of the gut microbiome in autism and pave the way towards potential microbial therapeutics for gastrointestinal problems in people with autism.ImportancePreviously, supplemental dietary zinc in the Shank3B−/− mouse model of autism spectrum disorder resulted in observations of ASD behaviours reversal; in this study we also used the Shank3B−/− mouse model to examine the influence of – and interaction between – dietary zinc and ASD-linked host genotype. Sample location along the gastrointestinal tract, genotype and zinc diet explained some of the variation in the microbiota data, with notable bacterial differences between treatment groups. Differential expression of host genes between treatment groups, including antimicrobial interaction genes and gut microbiota-regulated host metabolism genes, suggests that the interplay between gut microbes, the gastrointestinal tract and the brain may play a major role towards the observed amelioration of ASD behaviours seen previously with supplemented dietary zinc. These results widen the scope towards manipulating both dietary zinc and the microbiota itself to ameliorate ASD-related behaviours and associated gastrointestinal issues.

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Effect of Dietary Zinc Supplementation on the Gastrointestinal Microbiota and Host Gene Expression in the Shank3B-/- Mouse Model of Autism Spectrum Disorder

10.21203/rs.3.rs-864668/v1 ◽

2021 ◽

Author(s):

Giselle C Wong ◽

Yewon Jung ◽

Kevin Lee ◽

Chantelle Fourie ◽

Kim M. Handley ◽

...

Keyword(s):

Autism Spectrum Disorder ◽

Mouse Model ◽

Gut Microbiota ◽

Gut Microbiome ◽

Autism Spectrum ◽

Wild Type ◽

Dietary Zinc ◽

Knock Out ◽

Gastrointestinal Problems ◽

Host Metabolism

Abstract Background: Shank genes are implicated in ~1% of people with autism and mice with Shank3 knock out mutations exhibit autism-like behaviours. Zinc deficiency and gastrointestinal problems can be common among people with autism, and zinc is a key element required for SHANK protein function and gut development. In Shank3B-/- mice, a supplementary zinc diet reverses autism behaviours. We hypothesise that dietary zinc may alter the gut microbiome, potentially affecting the gut-microbiota-brain axis, which may contribute to changes in autism-like behaviours. Methods: Four types of gastrointestinal samples (ileum, caecum, colon, faecal) were collected from wild-type and knock-out Shank3B-/- mice on either control or supplemented-zinc diets. Bacterial 16S rRNA gene and fungal ITS2 genomic region amplicons were sequenced on the Illumina MiSeq platform and RNA on the Illumina HiSeq platform.Results: Cage, genotype and zinc diet each contributed significantly to bacterial community variation (accounting for 12.8%, 3.9% and 2.3% of the variation, respectively). Fungal diversity differed significantly between wild-type and knock-out Shank3B-/- mice on the control zinc diet, and the fungal biota differed among gut locations. RNA-seq analysis of host (mouse) transcripts revealed differential expression of genes involved in host metabolism that may be regulated by the gut microbiota and genes involved in anti-microbial interactions. Limitations: This study used the Shank3B-/- mouse model of autism spectrum disorder. Heterozygous and homozygous Shank3 gene mutations are found in 1% of the ASD population, only homozygous Shank3 mice were utilised in this study. Any translational conclusions should consider these limitations.Conclusions: By utilising the Shank3B-/- knock-out mouse model we were able to examine the influence of – and interactions between – dietary zinc and ASD-linked host genotype. Differential expression of host antimicrobial interaction genes as well as gut microbiota-regulated host metabolism genes among the treatment groups, suggests that the interplay between gut microbes, the gastrointestinal tract and the brain may play a major role towards the observed amelioration of ASD behaviours seen previously with supplemented dietary zinc. These data broaden understanding of the gut microbiome in autism and pave the way towards potential microbial therapeutics for gastrointestinal problems in people with autism.

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Dysregulation of DNA methylation during development as a potential mechanisms contributing to obsessive compulsive disorders and autism

10.31234/osf.io/udfya ◽

2019 ◽

Author(s):

German I. Todorov ◽

Karthikeyan Mayilvahanan ◽

David Ashurov ◽

Catarina Cunha

Keyword(s):

Mouse Model ◽

Autism Spectrum ◽

Obsessive Compulsive ◽

Cancer Treatments ◽

Knock Out ◽

Treatment Priority ◽

Underlying Mechanisms ◽

Knock Out Mice ◽

Obsessive Compulsive Disorders ◽

Compulsive Disorders

Autism Spectrum Disorder (ASD) is a pervasive developmental disorder, that is raising at a concerning rate. However, underlying mechanisms are still to be discovered. Obsessions and compulsions are the most debilitating aspect of these disorders (OCD), and they are the treatment priority for patients. SAPAP3 knock out mice present a reliable mouse model for repetitive compulsive behavior and are mechanistically closely related to the ASD mouse model Shank3 on a molecular level and AMPA receptor net effect. The phenotype of SAPAP3 knock out mice is obsessive grooming that leads to self-inflicted lesions by 4 months of age. Recent studies have accumulated evidence, that epigenetic mechanisms are important effectors in psychiatric conditions such as ASD and OCD. Methylation is the most studied mechanism, that recently lead to drug developments for more precise cancer treatments. We injected SAPAP3 mice with an epigenetic demethylation drug RG108 during pregnancy and delayed the onset of the phenotype in the offspring by 4 months. This result gives us clues about possible mechanism involved in OCD and ASD. Additionally, it shows that modulation of methylation mechanisms during development might be explored as a preventative treatment in the cases of high inherited risk of certain mental health conditions.

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Gut Microbiome-Based Analysis of Lipid A Biosynthesis in Individuals with Autism Spectrum Disorder: An In Silico Evaluation

Nutrients ◽

10.3390/nu13020688 ◽

2021 ◽

Vol 13 (2) ◽

pp. 688

Author(s):

Abdulkadir Yusif Maigoro ◽

Soojin Lee

Keyword(s):

Autism Spectrum Disorder ◽

Gut Microbiome ◽

In Silico ◽

Lipid A ◽

Autism Spectrum ◽

Spectrum Disorder ◽

Synthesis Rate ◽

Special Focus ◽

Control Subjects ◽

Gastrointestinal Problems

The link between autism spectrum disorder (ASD) and the gut microbiome has received much attention, with special focus on gut–brain-axis immunological imbalances. Gastrointestinal problems are one of the major symptoms of ASD and are thought to be related to immune dysregulation. Therefore, in silico analysis was performed on mined data from 36 individuals with ASD and 21 control subjects, with an emphasis on lipid A endotoxin-producing bacteria and their lipopolysaccharide (LPS) metabolic pathways. Analysis of enzyme distribution among the 15 most abundant genera in both groups revealed that almost all these genera utilized five early-stage enzymes responsible for catalyzing the nine conserved lipid A synthesis steps. However, Haemophilus and Escherichia, which were significantly more abundant in individuals with ASD than in the control subjects, possess a complete set of essential lipid A synthesis enzymes. Furthermore, the 10 genera with the greatest increase in individuals with ASD showed high potential for producing late-stage lipid A products. Collectively, these results suggested that the synthesis rate of immunogenic LPS end products is likely to increase in individuals with ASD, which may be related to their gastrointestinal symptoms and elevated inflammatory conditions.

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4225 Evaluation of Neurotransmitters in Channelopathy-Related Epilepsy

Journal of Clinical and Translational Science ◽

10.1017/cts.2020.298 ◽

2020 ◽

Vol 4 (s1) ◽

pp. 95-95

Author(s):

Sunita N Misra ◽

Theresa M. Czech ◽

Jennifer A. Kearney

Keyword(s):

Mouse Model ◽

Chart Review ◽

Retrospective Chart Review ◽

Brain Regions ◽

Autism Spectrum ◽

Wild Type ◽

Voltage Gated Sodium Channels ◽

Study Population ◽

Genotype A

OBJECTIVES/GOALS: Variants in voltage-gated sodium channels (VGSC) are a common cause of severe early onset epilepsy. Changes in CSF neurotransmitters (NT) were identified in 2 cases of VGSC-related epilepsy. Here we investigate NT changes in patients and a novel mouse model of VGSC-related epilepsy. METHODS/STUDY POPULATION: We conducted a single site IRB approved retrospective chart review of patients with VGSC-related epilepsy who underwent CSF NT testing for diagnostic purposes. In parallel, we examined NT levels from the brains of wild-type (WT) and a novel VGSC-related epilepsy mouse model after obtaining IACUC approval. We rapidly isolated forebrain, cortex, striatum, and brainstem from 5-6 animals per sex and genotype. A combination of HPLC with electrochemical detection and mass spectrometry were used to quantify NT levels from brain samples. RESULTS/ANTICIPATED RESULTS: We identified 10 patients with VGSC-related epilepsy who received CSF NT testing. Two of these patients had abnormal NT results including changes to dopamine (DA) or serotonin (5-HT) metabolites. We analyzed NT levels from four brain regions from male and female WT and VGSC-related epilepsy mice. We anticipate that most of the NT levels will be similar to WT, however subtle changes in the DA or 5-HT metabolites may be seen in VGSC-related epilepsy. DISCUSSION/SIGNIFICANCE OF IMPACT: Patients with VGSC-related epilepsy often have autism spectrum disorder, sleep, and movement disorders. Understanding the role of aberrant NT levels in VGSC-related epilepsy may provide additional therapeutic targets that address common neuropsychological comorbidities as well as seizures.

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Changes in the Fluorescence Tracking of NaV1.6 Protein Expression in a BTBR T+Itpr3tf/J Autistic Mouse Model

Neural Plasticity ◽

10.1155/2019/4893103 ◽

2019 ◽

Vol 2019 ◽

pp. 1-12 ◽

Cited By ~ 3

Author(s):

Musaad A. Alshammari ◽

Mohammad R. Khan ◽

Fawaz Alasmari ◽

Abdulaziz O. Alshehri ◽

Rizwan Ali ◽

...

Keyword(s):

Mouse Model ◽

Western Blot ◽

Neuronal Excitability ◽

Autism Spectrum ◽

Wild Type ◽

Western Blot Assay ◽

Critical Determinant ◽

Molecular Features ◽

Voltage Gated Sodium Channels ◽

Wild Type Control

The axon initial segment (AIS), the site of action potential initiation in neurons, is a critical determinant of neuronal excitability. Growing evidence indicates that appropriate recruitment of the AIS macrocomplex is essential for synchronized firing. However, disruption of the AIS structure is linked to the etiology of multiple disorders, including autism spectrum disorder (ASD), a condition characterized by deficits in social communication, stereotyped behaviors, and very limited interests. To date, a complete understanding of the molecular components that underlie the AIS in ASD has remained elusive. In this research, we examined the AIS structure in a BTBR T+Itpr3tf/J mouse model (BTBR), a valid model that exhibits behavioral, electrical, and molecular features of autism, and compared this to the C57BL/6J wild-type control mouse. Using Western blot studies and high-resolution confocal microscopy in the prefrontal frontal cortex (PFC), our data indicate disrupted expression of different isoforms of the voltage-gated sodium channels (NaV) at the AIS, whereas other components of AIS such as ankyrin-G and fibroblast growth factor 14 (FGF14) and contactin-associated protein 1 (Caspr) in BTBR were comparable to those in wild-type control mice. A Western blot assay showed that BTBR mice exhibited a marked increase in different sodium channel isoforms in the PFC compared to wild-type mice. Our results provide potential evidence for previously undescribed mechanisms that may play a role in the pathogenesis of autistic-like phenotypes in BTBR mice.

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The Brain-Gut-Microbiome System: Pathways and Implications for Autism Spectrum Disorder

Nutrients ◽

10.3390/nu13124497 ◽

2021 ◽

Vol 13 (12) ◽

pp. 4497

Author(s):

Michelle A. Chernikova ◽

Genesis D. Flores ◽

Emily Kilroy ◽

Jennifer S. Labus ◽

Emeran A. Mayer ◽

...

Keyword(s):

Autism Spectrum Disorder ◽

Gut Microbiota ◽

Gut Microbiome ◽

Fecal Microbiota Transplantation ◽

Fecal Microbiota ◽

Autism Spectrum ◽

Spectrum Disorder ◽

Preclinical Research ◽

Gastrointestinal Problems ◽

The Brain

Gastrointestinal dysfunction is one of the most prevalent physiological symptoms of autism spectrum disorder (ASD). A growing body of largely preclinical research suggests that dysbiotic gut microbiota may modulate brain function and social behavior, yet little is known about the mechanisms that underlie these relationships and how they may influence the pathogenesis or severity of ASD. While various genetic and environmental risk factors have been implicated in ASD, this review aims to provide an overview of studies elucidating the mechanisms by which gut microbiota, associated metabolites, and the brain interact to influence behavior and ASD development, in at least a subgroup of individuals with gastrointestinal problems. Specifically, we review the brain-gut-microbiome system and discuss findings from current animal and human studies as they relate to social-behavioral and neurological impairments in ASD, microbiota-targeted therapies (i.e., probiotics, fecal microbiota transplantation) in ASD, and how microbiota may influence the brain at molecular, structural, and functional levels, with a particular interest in social and emotion-related brain networks. A deeper understanding of microbiome-brain-behavior interactions has the potential to inform new therapies aimed at modulating this system and alleviating both behavioral and physiological symptomatology in individuals with ASD.

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Effects of gene by microbiome interactions on behavioral and neurobiological phenotypes in a mouse model for autism spectrum disorder

10.1101/2020.04.29.065821 ◽

2020 ◽

Author(s):

Aya Osman ◽

Nicholas L. Mervosh ◽

Ana N. Strat ◽

Katherine R. Meckel ◽

Tanner J. Euston ◽

...

Keyword(s):

Autism Spectrum Disorder ◽

Environmental Factors ◽

Mouse Model ◽

Translational Research ◽

Gut Microbiome ◽

Neurodevelopmental Disorder ◽

Autism Spectrum ◽

Spectrum Disorder ◽

Research Strategies ◽

Stage Of Development

AbstractAutism spectrum disorder (ASD) is a serious neurodevelopmental disorder with a very high prevalence rate and a chronic disease course beginning in early childhood. Despite the tremendous burden of ASD, there are currently no disease-modifying treatments. Like many neuropsychiatric illnesses ASD has a complex pathophysiology driven by genetic and environmental factors. There is interest in identifying modifiable environmental factors as potential translational research strategies for development of therapeutics for ASD. A rapidly growing body of research demonstrates that the resident bacteria of the gastrointestinal tract, collectively the gut microbiome, have profound influence on brain and behavior. This gut-brain signaling pathway is highly relevant to ASD as the microbiome begins to form at birth, is heavily influenced by environmental factors throughout early life, and begins to stabilize at the same stage of development that symptoms of ASD begin to develop. To investigate potential gene x microbiome interactions in a model of ASD, we utilized mutant mice carrying a deletion of the ASD-associated Shank3 gene (Shank3KO), which clinically manifests as Phelan-McDermid syndrome, as a model for genetic risk of ASD. Analysis of the gut microbiome of Shank3KO mice demonstrated genotype effects on both microbiome composition and metabolite production. Behaviorally, Shank3KO mice demonstrate decreased social interactions and have altered anxiety and compulsive-like behaviors. Disruption of the microbiome with broad spectrum antibiotics lead to an exacerbation of all behavioral phenotypes in Shank3KO mice. Additionally, we found that Shank3KO mice had markedly increased changes in gene expression in the prefrontal cortex following microbiome depletion. Taken together, our results suggest a gene x microbiome interaction in this mouse model for ASD and raise the possibility that targeting the microbiome may be a valid translational research strategy in developing therapeutics for ASD.

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Changes in Receptive Field Size in Syngap1 +/- Mouse Model of Autism Spectrum Disorder

Proceedings of IMPRS ◽

10.18060/25850 ◽

2021 ◽

Vol 4 (1) ◽

Author(s):

Evan Anderson ◽

Charles A Martin ◽

Maria Dadarlat

Keyword(s):

Autism Spectrum Disorder ◽

Intellectual Disability ◽

Receptive Field ◽

Mouse Model ◽

Neural Activity ◽

Autism Spectrum ◽

Field Size ◽

Wild Type ◽

Receptive Field Size ◽

Field Area

Background and Hypothesis: Autism spectrum disorder (ASD) is a form of intellectual disability with impairments in social functioning and cognition. Mutations within the SynGAP1 gene are associated with ASD due to over activation of RAS-GTP causing insertion of AMPA receptors onto the post synaptic membrane, and thus early maturation of dendrites. These mutations lead to an excitatory/inhibitory imbalance within the brain, and patients often present with intellectual disability, seizures, and issues of cognition. Research has shown that Syngap1 +/- mice have decreased cortical gray matter brain volume throughout areas involved in the visual system pathway. However, hierarchal visual processing has not been well characterized in a Syngap1 +/- mouse model of autism. Project Methods: Using 64 and 128 channel microelectrodes, we recorded the neural activity within V1 of four mice. Neural activity was recorded in response to visual stimuli - testing receptive field size between two wild-type and two Syngap1 +/- mice. Data was run through spike sorting algorithms to identify neurons. Receptive field area for each neuron was then calculated and compared between the two genotypes. Results: The receptive field areas of Syngap1 +/- mice were statistically larger (p < 0.01) compared to wild type mice. Syngap1 +/- mice had a mean receptive field area of 450.5 visual degrees (±443.7) and WT mice had a mean receptive field area of 261.1 visual degrees (±187.21). Most of the neurons within Syngap +/- had no distinct receptive field, 67.9%, while only 25.7% of wild type neurons lacked a distinct receptive field. Conclusion and Potential Impact: Overall, Syngap1 +/- mice had larger, and less distinct receptive fields compared to wild type mice. Deficiencies in the Syngap1 protein may impair refinement of visual stimuli. Understanding the mechanism of response of missing SynGAP1 can inform directed therapies and interventions to treat patients with the missing gene and manage their intellectual disability.

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Oxytocin receptors influence the development and maintenance of social behavior in zebrafish (Danio rerio).

10.1101/2021.06.26.449566 ◽

2021 ◽

Author(s):

Anja Gemmer ◽

Kristina Mirkes ◽

Lukas Anneser ◽

Tim Eilers ◽

Caroline Kibat ◽

...

Keyword(s):

Social Behavior ◽

Social Preference ◽

Autism Spectrum ◽

Social Impairment ◽

Wild Type ◽

Knock Out ◽

Oxytocin Receptors ◽

Shoaling Behavior ◽

Kinetics Of

Zebrafish are highly social teleost fish and an excellent model to study social behavior. The neuropeptide Oxytocin is associated different social behaviors as well as disorders resulting in social impairment like autism spectrum disorder. However, how Oxytocin receptor signaling affects the development and expression kinetics of social behavior is not known. In this study we investigated the role of the two oxytocin receptors, Oxtr and Oxtrl, in the development and maintenance of social preference and shoaling behavior in 2- to 8-week-old zebrafish. Using CRISPR/Cas9 mediated oxtr and oxtrl knock-out fish, we found that the development of social preference is accelerated if one of the Oxytocin receptors is knocked-out and that the knock-out fish reach significantly higher levels of social preference. Moreover, oxtr-/- fish showed impairments in the maintenance of social preference. Social isolation prior to testing led to impaired maintenance of social preference in both wild-type and oxtr and oxtrl knock-out fish. Knocking-out one of the Oxytocin receptors also led to increased group spacing and reduced polarization in a 20-fish shoal at 8 weeks post fertilization, but not at 4. These results show that the development and maintenance of social behavior is influenced by the Oxytocin receptors and that the effects are not just pro- or antisocial, but dependent on both the age and social context of the fish.

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Increased axonal bouton stability during learning in the mouse model of MECP2 duplication syndrome

10.1101/186239 ◽

2017 ◽

Author(s):

Ryan T. Ash ◽

Paul G. Fahey ◽

Jiyoung Park ◽

Huda Y. Zoghbi ◽

Stelios M. Smirnakis

Keyword(s):

Motor Learning ◽

Mouse Model ◽

Dendritic Spine ◽

Pyramidal Neuron ◽

Structural Plasticity ◽

Autism Spectrum ◽

Wild Type ◽

Syndromic Autism ◽

Mecp2 Duplication Syndrome ◽

Duplication Syndrome

ABSTRACTMECP2-duplication syndrome is an X-linked form of syndromic autism caused by genomic duplication of the region encoding Methyl-CpG-binding protein 2. Mice overexpressing MECP2 demonstrate altered patterns of learning and memory, including enhanced motor learning. Previous work associated this enhanced motor learning to abnormally increased stability of dendritic spine clusters formed in the apical tuft of corticospinal, area M1, neurons during rotarod training. In the current study, we measure the structural plasticity of axonal boutons in Layer 5 (L5) pyramidal neuron projections to layer 1 of area M1 during motor learning. In wild-type mice we find that during rotarod training, bouton formation rate changes minimally, if at all, while bouton elimination rate doubles. Notably, the observed upregulation in bouton elimination with learning is absent in MECP2-duplication mice. This result provides further evidence of imbalance between structural stability and plasticity in this form of syndromic autism. Furthermore, the observation that axonal bouton elimination doubles with motor learning in wild-type animals contrasts with the increase of dendritic spine consolidation observed in corticospinal neurons at the same layer. This dissociation suggests that different area M1 microcircuits may manifest different patterns of structural synaptic plasticity during motor learning.SIGNIFICANCE STATEMENTAbnormal balance between synaptic stability and plasticity is a feature of several autism spectrum disorders, often corroborated by in vivo studies of dendritic spine turnover. Here we provide the first evidence that abnormally increased stability of axonal boutons, the presynaptic component of excitatory synapses, occurs during motor learning in the MECP2 duplication syndrome mouse model of autism. In contrast, in normal controls, axonal bouton elimination in L5 pyramidal neuron projections to layer 1 of area M1 doubles with motor learning. The fact that axonal projection boutons get eliminated, while corticospinal dendritic spines get consolidated with motor learning in layer 1 of area M1, suggests that structural plasticity manifestations differ across different M1 microcircuits.

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