Large-scale cellular-resolution gene profiling in human neocortex reveals species-specific molecular signatures.

Although there have been major advances in elucidating the functional biology of the human brain, relatively little is known of its cellular and molecular organization. Here we report a large-scale characterization of the expression of ∼1,000 genes important for neural functions by in situ hybridization at a cellular resolution in visual and temporal cortices of adult human brains. These data reveal diverse gene expression patterns and remarkable conservation of each individual gene's expression among individuals (95%), cortical areas (84%), and between human and mouse (79%). A small but substantial number of genes (21%) exhibited species-differential expression. Distinct molecular signatures, comprised of genes both common between species and unique to each, were identified for each major cortical cell type. The data suggest that gene expression profile changes may contribute to differential cortical function across species, and in particular, a shift from corticosubcortical to more predominant corticocortical communications in the human brain.

Pten Mutations Alter Brain Growth Trajectory and Allocation of Cell Types through Elevated ß-Catenin Signaling.

Abnormal patterns of head and brain growth are a replicated finding in a subset of individuals with autism spectrum disorder (ASD). It is not known whether risk factors associated with ASD and abnormal brain growth (both overgrowth and undergrowth) converge on common biological pathways and cellular mechanisms in the developing brain. Heterozygous mutations in PTEN (PTEN(+/-)), which encodes a negative regulator of the PI3K-Akt-mTOR pathway, are a risk factor for ASD and macrocephaly. Here we use the developing cerebral cortex of Pten(+/-) mice to investigate the trajectory of brain overgrowth and underlying cellular mechanisms. We find that overgrowth is detectable from birth to adulthood, is driven by hyperplasia, and coincides with excess neurons at birth and excess glia in adulthood. ß-Catenin signaling is elevated in the developing Pten(+/-) cortex, and a heterozygous mutation in Ctnnb1 (encoding ß-catenin), itself a candidate gene for ASD and microcephaly, can suppress Pten(+/-) cortical overgrowth. Thus, a balance of Pten and ß-catenin signaling regulates normal brain growth trajectory by controlling cell number, and imbalance in this relationship can result in abnormal brain growth. SIGNIFICANCE STATEMENT: We report that Pten haploinsufficiency leads to a dynamic trajectory of brain overgrowth during development and altered scaling of neuronal and glial cell populations. ß-catenin signaling is elevated in the developing cerebral cortex of Pten haploinsufficient mice, and a heterozygous mutation in ß-catenin, itself a candidate gene for ASD and microcephaly, suppresses Pten(+/-) cortical overgrowth. This leads to the new insight that Pten and ß-catenin signaling act in a common pathway to regulate normal brain growth trajectory by controlling cell number, and disruption of this pathway can result in abnormal brain growth.

Pten haploinsufficient mice show broad brain overgrowth but selective impairments in autism-relevant behavioral tests.

Accelerated head and brain growth (macrocephaly) during development is a replicated biological finding in a subset of individuals with autism spectrum disorder (ASD). However, the relationship between brain overgrowth and the behavioral and cognitive symptoms of ASD is poorly understood. The PI3K-Akt-mTOR pathway regulates cellular growth; several genes encoding negative regulators of this pathway are ASD risk factors, including PTEN. Mutations in PTEN have been reported in individuals with ASD and macrocephaly. We report that brain overgrowth is widespread in Pten germline haploinsufficient (Pten(+/-)) mice, reflecting Pten mRNA expression in the developing brain. We then ask if broad brain overgrowth translates into general or specific effects on the development of behavior and cognition by testing Pten(+/-) mice using assays relevant to ASD and comorbidities. Deficits in social behavior were observed in both sexes. Males also showed abnormalities related to repetitive behavior and mood/anxiety. Females exhibited circadian activity and emotional learning phenotypes. Widespread brain overgrowth together with selective behavioral impairments in Pten(+/-) mice raises the possibility that most brain areas and constituent cell types adapt to an altered trajectory of growth with minimal impact on the behaviors tested in our battery; however, select areas/cell types relevant to social behavior are more vulnerable or less adaptable, thus resulting in social deficits. Probing dopaminergic neurons as a candidate vulnerable cell type, we found social behavioral impairments in mice with Pten conditionally inactivated in dopaminergic neurons that are consistent with the possibility that desynchronized growth in key cell types may contribute to ASD endophenotypes.

Ectopic expression of the striatal-enriched GTPase Rhes elicits cerebellar degeneration and an ataxia phenotype in Huntington's disease.

Huntington's disease (HD) is caused by an expansion of glutamine repeats in the huntingtin protein (mHtt) that invokes early and prominent damage of the striatum, a region that controls motor behaviors. Despite its ubiquitous expression, why certain brain regions, such as the cerebellum, are relatively spared from neuronal loss by mHtt remains unclear. Previously, we implicated the striatal-enriched GTPase, Rhes (Ras homolog enriched in the striatum), which binds and SUMOylates mHtt and increases its solubility and cellular cytotoxicity, as the cause for striatal toxicity in HD. Here, we report that Rhes deletion in HD mice (N171-82Q), which express the N-terminal fragment of human Htt with 82 glutamines (Rhes(-/-)/N171-82Q), display markedly reduced HD-related behavioral deficits, and absence of lateral ventricle dilatation (secondary to striatal atrophy), compared to control HD mice (N171-82Q). To further validate the role of GTPase Rhes in HD, we tested whether ectopic Rhes expression would elicit a pathology in a brain region normally less affected in HD. Remarkably, ectopic expression of Rhes in the cerebellum of N171-82Q mice, during the asymptomatic period led to an exacerbation of motor deficits, including loss of balance and motor incoordination with ataxia-like features, not apparent in control-injected N171-82Q mice or Rhes injected wild-type mice. Pathological and biochemical analysis of Rhes-injected N171-82Q mice revealed a cerebellar lesion with marked loss of Purkinje neuron layer parvalbumin-immunoreactivity, induction of caspase 3 activation, and enhanced soluble forms of mHtt. Similarly reintroducing Rhes into the striatum of Rhes deleted Rhes(-/-)Hdh(150Q/150Q) knock-in mice, elicited a progressive HD-associated rotarod deficit. Overall, these studies establish that Rhes plays a pivotal role in vivo for the selective toxicity of mHtt in HD.

Comparing synaptic proteomes across five mouse models for autism reveals converging molecular similarities including deficits in oxidative phosphorylation and Rho GTPase signaling.

Specific and effective treatments for autism spectrum disorder (ASD) are lacking due to a poor understanding of disease mechanisms. Here we test the idea that similarities between diverse ASD mouse models are caused by deficits in common molecular pathways at neuronal synapses. To do this, we leverage the availability of multiple genetic models of ASD that exhibit shared synaptic and behavioral deficits and use quantitative mass spectrometry with isobaric tandem mass tagging (TMT) to compare their hippocampal synaptic proteomes. Comparative analyses of mouse models for Fragile X syndrome (Fmr1 knockout), cortical dysplasia focal epilepsy syndrome (Cntnap2 knockout), PTEN hamartoma tumor syndrome (Pten haploinsufficiency), ANKS1B syndrome (Anks1b haploinsufficiency), and idiopathic autism (BTBR+) revealed several common altered cellular and molecular pathways at the synapse, including changes in oxidative phosphorylation, and Rho family small GTPase signaling. Functional validation of one of these aberrant pathways, Rac1 signaling, confirms that the ANKS1B model displays altered Rac1 activity counter to that observed in other models, as predicted by the bioinformatic analyses. Overall similarity analyses reveal clusters of synaptic profiles, which may form the basis for molecular subtypes that explain genetic heterogeneity in ASD despite a common clinical diagnosis. Our results suggest that ASD-linked susceptibility genes ultimately converge on common signaling pathways regulating synaptic function and propose that these points of convergence are key to understanding the pathogenesis of this disorder.

Haploinsufficiency for Pten and Serotonin transporter cooperatively influences brain size and social behavior.

Autism spectrum disorder (ASD) is a heterogeneous group of neurodevelopmental disorders that share deficits in sociability, communication, and restrictive and repetitive interests. ASD is likely polygenic in origin in most cases, but we presently lack an understanding of the relationships between ASD susceptibility genes and the neurobiological and behavioral phenotypes of ASD. Two genes that have been implicated as conferring susceptibility to ASD are PTEN and Serotonin transporter (SLC6A4). The PI3K and serotonin pathways, in which these genes respectively act, are both potential biomarkers for ASD diagnosis and treatment. Biochemical evidence exists for an interaction between these pathways; however, the relevance of this for the pathogenesis of ASD is unclear. We find that Pten haploinsufficient (Pten(+/-)) mice are macrocephalic, and this phenotype is exacerbated in Pten(+/-); Slc6a4(+/-) mice. Furthermore, female Pten(+/-) mice are impaired in social approach behavior, a phenotype that is exacerbated in female Pten(+/-); Slc6a4(+/-) mice. While increased brain size correlates with decreased sociability across these genotypes in females, within each genotype increased brain size correlates with increased sociability, suggesting that epigenetic influences interact with genetic factors in influencing the phenotype. These findings provide insight into an interaction between two ASD candidate genes during brain development and point toward the use of compound mutant mice to validate biomarkers for ASD against biological and behavioral phenotypes.

Pten haploinsufficiency causes desynchronized growth of brain areas involved in sensory processing.

How changes in brain scaling relate to altered behavior is an important question in neurodevelopmental disorder research. Mice with germline Pten haploinsufficiency (Pten +/-) closely mirror the abnormal brain scaling and behavioral deficits seen in humans with macrocephaly/autism syndrome, which is caused by PTEN mutations. We explored whether deviation from normal patterns of growth can predict behavioral abnormalities. Brain regions associated with sensory processing (e.g., pons and inferior colliculus) had the biggest deviations from expected volume. While Pten +/- mice showed little or no abnormal behavior on most assays, both sexes showed sensory deficits, including impaired sensorimotor gating and hyporeactivity to high-intensity stimuli. Developmental analysis of this phenotype showed sexual dimorphism for hyporeactivity. Mapping behavioral phenotypes of Pten +/- mice onto relevant brain regions suggested abnormal behavior is likely when associated with relatively enlarged brain regions, while unchanged or relatively decreased brain regions have little predictive value.

Dyrk1a Mutations Cause Undergrowth of Cortical Pyramidal Neurons via Dysregulated Growth Factor Signaling.

BACKGROUND: Mutations in DYRK1A are a cause of microcephaly, autism spectrum disorder, and intellectual disability; however, the underlying cellular and molecular mechanisms are not well understood. METHODS: We generated a conditional mouse model using Emx1-cre, including conditional heterozygous and homozygous knockouts, to investigate the necessity of Dyrk1a in the cortex during development. We used unbiased, high-throughput phosphoproteomics to identify dysregulated signaling mechanisms in the developing Dyrk1a mutant cortex as well as classic genetic modifier approaches and pharmacological therapeutic intervention to rescue microcephaly and neuronal undergrowth caused by Dyrk1a mutations. RESULTS: We found that cortical deletion of Dyrk1a in mice causes decreased brain mass and neuronal size, structural hypoconnectivity, and autism-relevant behaviors. Using phosphoproteomic screening, we identified growth-associated signaling cascades dysregulated upon Dyrk1a deletion, including TrkB-BDNF (tyrosine receptor kinase B-brain-derived neurotrophic factor), an important regulator of ERK/MAPK (extracellular signal-regulated kinase/mitogen-activated protein kinase) and mTOR (mammalian target of rapamycin) signaling. Genetic suppression of Pten or pharmacological treatment with IGF-1 (insulin-like growth factor-1), both of which impinge on these signaling cascades, rescued microcephaly and neuronal undergrowth in neonatal mutants. CONCLUSIONS: Altogether, these findings identify a previously unknown mechanism through which Dyrk1a mutations disrupt growth factor signaling in the developing brain, thus influencing neuronal growth and connectivity. Our results place DYRK1A as a critical regulator of a biological pathway known to be dysregulated in humans with autism spectrum disorder and intellectual disability. In addition, these data position Dyrk1a within a larger group of autism spectrum disorder/intellectual disability risk genes that impinge on growth-associated signaling cascades to regulate brain size and connectivity, suggesting a point of convergence for multiple autism etiologies.

Connecting Genotype with Behavioral Phenotype in Mouse Models of Autism Associated with PTEN Mutations.

A subset of individuals with autism spectrum disorder (ASD) and macrocephaly carry mutations in the gene PTEN. Animal models, particularly mice, have been helpful in establishing a causal role for Pten mutations in autism-relevant behavioral deficits. These models are a useful tool for investigating neurobiological mechanisms of these behavioral phenotypes and developing potential therapeutic interventions. Here we provide an overview of various genetic mouse models that have been used to characterize behavioral phenotypes caused by perturbation of Pten We discuss convergent and divergent phenotypes across models with the aim of highlighting a set of behavioral domains that are sensitive to the effects of Pten mutation and that may provide useful readouts for translational and basic neuroscience research.

Social Behavior Is Modulated by Valence-Encoding mPFC-Amygdala Sub-circuitry.

The prefrontal cortex and amygdala are anatomical substrates linked to both social information and emotional valence processing, but it is not known whether sub-circuits in the medial prefrontal cortex (mPFC) that project to the basolateral amygdala (BLA) are recruited and functionally contribute to social approach-avoidance behavior. Using retrograde labeling of mPFC projections to the BLA, we find that BLA-projecting neurons in the infralimbic cortex (IL) are preferentially activated in response to a social cue as compared with BLA-projecting neurons in the prelimbic cortex (PL). Chemogenetic interrogation of these sub-circuits shows that activation of PL-BLA or inhibition of IL-BLA circuits impairs social behavior. Sustained closed-loop optogenetic activation of PL-BLA circuitry induces social impairment, corresponding to a negative emotional state as revealed by real-time place preference behavioral avoidance. Reactivation of foot shock-responsive PL-BLA circuitry impairs social behavior. Altogether, these data suggest a circuit-level mechanism by which valence-encoding mPFC-BLA sub-circuits shape social approach-avoidance behavior.

Elevated protein synthesis in microglia causes autism-like synaptic and behavioral aberrations.

Mutations that inactivate negative translation regulators cause autism spectrum disorders (ASD), which predominantly affect males and exhibit social interaction and communication deficits and repetitive behaviors. However, the cells that cause ASD through elevated protein synthesis resulting from these mutations remain unknown. Here we employ conditional overexpression of translation initiation factor eIF4E to increase protein synthesis in specific brain cells. We show that exaggerated translation in microglia, but not neurons or astrocytes, leads to autism-like behaviors in male mice. Although microglial eIF4E overexpression elevates translation in both sexes, it only increases microglial density and size in males, accompanied by microglial shift from homeostatic to a functional state with enhanced phagocytic capacity but reduced motility and synapse engulfment. Consequently, cortical neurons in the mice have higher synapse density, neuroligins, and excitation-to-inhibition ratio compared to control mice. We propose that functional perturbation of male microglia is an important cause for sex-biased ASD.

Genetic Suppression of mTOR Rescues Synaptic and Social Behavioral Abnormalities in a Mouse Model of Pten Haploinsufficiency.

Heterozygous mutations in PTEN, which encodes a negative regulator of the mTOR and ß-catenin signaling pathways, cause macrocephaly/autism syndrome. However, the neurobiological substrates of the core symptoms of this syndrome are poorly understood. Here, we investigate the relationship between cerebral cortical overgrowth and social behavior deficits in conditional Pten heterozygous female mice (Pten cHet) using Emx1-Cre, which is expressed in cortical pyramidal neurons and a subset of glia. We found that conditional heterozygous mutation of Ctnnb1 (encoding ß-catenin) suppresses Pten cHet cortical overgrowth, but not social behavioral deficits, whereas conditional heterozygous mutation of Mtor suppresses social behavioral deficits, but not cortical overgrowth. Neuronal activity in response to social cues and excitatory synapse markers are elevated in the medial prefrontal cortex (mPFC) of Pten cHet mice, and heterozygous mutation in Mtor, but not Ctnnb1, rescues these phenotypes. These findings indicate that macroscale cerebral cortical overgrowth and social behavioral phenotypes caused by Pten haploinsufficiency can be dissociated based on responsiveness to genetic suppression of Ctnnb1 or Mtor. Furthermore, neuronal connectivity appears to be one potential substrate for mTOR-mediated suppression of social behavioral deficits in Pten haploinsufficient mice. Autism Res 2019, 12: 1463-1471. © 2019 International Society for Autism Research, Wiley Periodicals, Inc. LAY SUMMARY: A subgroup of individuals with autism display overgrowth of the head and the brain during development. Using a mouse model of an autism risk gene, Pten, that displays both brain overgrowth and social behavioral deficits, we show here that that these two symptoms can be dissociated. Reversal of social behavioral deficits in this model is associated with rescue of abnormal synaptic markers and neuronal activity.

Pten haploinsufficiency disrupts scaling across brain areas during development in mice.

Haploinsufficiency for PTEN is a cause of autism spectrum disorder and brain overgrowth; however, it is not known if PTEN mutations disrupt scaling across brain areas during development. To address this question, we used magnetic resonance imaging to analyze brains of male Pten haploinsufficient (Pten+/-) mice and wild-type littermates during early postnatal development and adulthood. Adult Pten+/- mice display a consistent pattern of abnormal scaling across brain areas, with white matter (WM) areas being particularly affected. This regional and WM enlargement recapitulates structural abnormalities found in individuals with PTEN haploinsufficiency and autism. Early postnatal Pten+/- mice do not display the same pattern, instead exhibiting greater variability across mice and brain regions than controls. This suggests that Pten haploinsufficiency may desynchronize growth across brain regions during early development before stabilizing by maturity. Pten+/- cortical cultures display increased proliferation of glial cell populations, indicating a potential substrate of WM enlargement, and provide a platform for testing candidate therapeutics. Pten haploinsufficiency dysregulates coordinated growth across brain regions during development. This results in abnormally scaled brain areas and associated behavioral deficits, potentially explaining the relationship between PTEN mutations and neurodevelopmental disorders.

Multiple roles for apoptosis facilitating condensation of the Drosophila ventral nerve cord.

At the end of embryogenesis, the ventral nerve cord (VNC) of Drosophila undergoes a shape change, termed condensation. During condensation the length of the VNC shortens by 25%, a process dependent on extracellular matrix deposited by hemocytes, an intact cytoskeleton of glia and neurons and neural activity. Here we show that cell death contributes to nerve cord shortening. Firstly, apoptosis occurs at the interface of the epidermis and the nerve cord where it plays a role in the separation of these two tissues. Separation precedes condensation and in conditions where separation is prevented, condensation fails. Secondly, many cells undergo apoptosis within VNC during condensation. This cell death is localized mainly to the posterior part of the nerve cord where more than half of all cell death occurs. Preventing apoptosis either in neurons or glia partially inhibits VNC shortening during condensation. Despite the importance of midline glia in axon tract development, preventing midline glia cell death results in normal hatching and adult formation. We find that undead midline glia are eliminated from the midline and become mispositioned or expelled from the nervous system. We suggest that this represent a form of pattern repair that operates to reduce the impact of the additional cells.

Improved Scalability of Neuron-Based Phenotypic Screening Assays for Therapeutic Discovery in Neuropsychiatric Disorders.

There is a pressing need to improve approaches for drug discovery related to neuropsychiatric disorders (NSDs). Therapeutic discovery in neuropsychiatric disorders would benefit from screening assays that can measure changes in complex phenotypes linked to disease mechanisms. However, traditional assays that track complex neuronal phenotypes, such as neuronal connectivity, exhibit poor scalability and are not compatible with high-throughput screening (HTS) procedures. Therefore, we created a neuronal phenotypic assay platform that focused on improving the scalability and affordability of neuron-based assays capable of tracking disease-relevant phenotypes. First, using inexpensive laboratory-level automation, we industrialized primary neuronal culture production, which enabled the creation of scalable assays within functioning neural networks. We then developed a panel of phenotypic assays based on culturing of primary neurons from genetically modified mice expressing HTS-compatible reporters that capture disease-relevant phenotypes. We demonstrated that a library of 1,280 compounds was quickly screened against both assays using only a few litters of mice in a typical academic laboratory setting. Finally, we implemented one assay in a fully automated high-throughput academic screening facility, illustrating the scalability of assays designed using this platform. These methodological improvements simplify the creation of highly scalable neuron-based phenotypic assays designed to improve drug discovery in CNS disorders.

A function for Egf receptor signaling in expanding the developing brain in Drosophila.

BACKGROUND: In invertebrates and vertebrates, neural midline cells secrete signals that pattern the central nervous system (CNS). However, an important part of the developing insect brain, involved in functions such as olfaction and feeding behavior, is positioned lateral to the foregut and lacks neural cells at the midline. Could the foregut substitute for neural midline cells and secrete signals that pattern this part of the brain? RESULTS: In Drosophila embryos, the neural midline marker Single-minded is expressed in foregut cells adjacent to the brain, as are members of the Egf receptor signaling pathway. Removing the function of these molecules results in aberrant proliferation and reduced size in the brain lateral to the foregut. CONCLUSIONS: Cells of the brain lateral to the foregut receive an Egf signal from the midline and proliferate in response. A likely source of this signal is the foregut. These findings raise the possibility that the brain lateral to the foregut is an evolutionarily recent addition to the arthropod brain, and that the anterior boundary of the brain neural midline is a conserved feature in bilaterally symmetric animals.

Forebrain depletion of Rheb GTPase elicits spatial memory deficits in mice.

The precise molecular and cellular events responsible for age-dependent cognitive dysfunctions remain unclear. We report that Rheb (ras homolog enriched in brain) GTPase, an activator of mammalian target of rapamycin (mTOR), regulates memory functions in mice. Conditional depletion of Rheb selectively in the forebrain of mice obtained from crossing Rhebf/f and CamKIICre results in spontaneous signs of age-related memory loss, that is, spatial memory deficits (T-maze, Morris water maze) without affecting locomotor (open-field test), anxiety-like (elevated plus maze), or contextual fear conditioning functions. Partial depletion of Rheb in forebrain was sufficient to elicit memory defects with little effect on the neuronal size, cortical thickness, or mammalian target of rapamycin activity. Rheb depletion, however, increased the levels of beta-site amyloid precursor protein cleaving enzyme 1 (BACE1), a protein elevated in aging and Alzheimer's disease. Overall, our study demonstrates that forebrain Rheb promotes aging-associated cognitive defects. Thus, molecular understanding of Rheb pathway in brain may provide new therapeutic targets for aging and/or Alzheimer's disease-associated memory deficits.

Hyperconnectivity of prefrontal cortex to amygdala projections in a mouse model of macrocephaly/autism syndrome.

Multiple autism risk genes converge on the regulation of mTOR signalling, which is a key effector of neuronal growth and connectivity. We show that mTOR signalling is dysregulated during early postnatal development in the cerebral cortex of germ-line heterozygous Pten mutant mice (Pten+/-), which model macrocephaly/autism syndrome. The basolateral amygdala (BLA) receives input from subcortical-projecting neurons in the medial prefrontal cortex (mPFC). Analysis of mPFC to BLA axonal projections reveals that Pten+/- mice exhibit increased axonal branching and connectivity, which is accompanied by increased activity in the BLA in response to social stimuli and social behavioural deficits. The latter two phenotypes can be suppressed by pharmacological inhibition of S6K1 during early postnatal life or by reducing the activity of mPFC-BLA circuitry in adulthood. These findings identify a mechanism of altered connectivity that has potential relevance to the pathophysiology of macrocephaly/autism syndrome and autism spectrum disorders featuring dysregulated mTOR signalling.

Autism-relevant behaviors are minimally impacted by conditional deletion of Pten in oxytocinergic neurons.

Germline heterozygous mutations in Pten (phosphatase and tensin homolog) are associated with macrocephaly and autism spectrum disorders (ASD). Pten germline heterozygous (Pten+/- ) mice approximate these mutations, and both sexes show widespread brain overgrowth and impaired social behavior. Strikingly similar behavior phenotypes have been reported in oxytocin (Oxt) and/or oxytocin receptor (OxtR) knockout mice. Thus, we hypothesized that the behavioral phenotypes of germline Pten+/- mice may be caused by reduced Pten function in Oxt-expressing cells. To investigate this, we tested mice in which Pten was conditionally deleted using oxytocin-Cre (Oxt-Cre+ ; PtenloxP/+ , Oxt-Cre+ ; PtenloxP/loxP ) on a battery including assays of social, repetitive, depression-like, and anxiety-like behaviors. Minimal behavioral abnormalities were found; decreased anxiety-like behavior in the open field test in Oxt-Cre+ ; PtenloxP/loxP males was the only result that phenocopied germline Pten+/- mice. However, Oxt cell size was dramatically increased in Oxt-Cre+ ; PtenloxP/loxP mice in adulthood. Thus, conditional deletion of Pten using Oxt-Cre has a profound effect on Oxt cell structure, but not on ASD-relevant behavior. We interpret these results as inconsistent with our starting hypothesis that reduced Pten function in Oxt-expressing cells causes the behavioral deficits observed in germline Pten+/- mice. Autism Res 2016, 9: 1248-1262. © 2016 International Society for Autism Research, Wiley Periodicals, Inc.

Zika virus infection during the period of maximal brain growth causes microcephaly and corticospinal neuron apoptosis in wild type mice.

Zika virus (ZIKV) infection in pregnant women has been established as a cause of microcephaly in newborns. Here we test the hypothesis that neurodevelopmental stages when the brain is undergoing rapid growth are particularly vulnerable to the effects of ZIKV infection. We injected ZIKV intracranially into wild type C57BL/6 mice at two different time points: early postnatal development, when the brain is growing at its maximal rate, and at weaning, when the brain has largely reached adult size. Both time points showed widespread immunoreactivity for ZIKV and cleaved caspase 3 (CC3, a marker of apoptosis) throughout the brain. However, in early postnatal ZIKV injected mice, some brain areas and cell types display particularly large increases in apoptosis that we did not observe in older animals. Corticospinal pyramidal neurons, a cell type implicated in human microcephaly associated with ZIKV infection, are an example of one such cell type. Proliferating cells in the ventricular zone stem cell compartment are also depleted. These findings are consistent with the hypothesis that periods of rapid brain growth are especially susceptible to neurodevelopmental effects of ZIKV infection, and establish a valuable model to investigate mechanisms underlying neurodevelopmental effects of ZIKV infection and explore candidate therapeutics.