Default Patterning Produces Pan-cortical Glutamatergic and CGE/LGE-like GABAergic Neurons from Human Pluripotent Stem Cells.

Default differentiation of human pluripotent stem cells has been promoted as a model of cortical development. In this study, a developmental transcriptome analysis of default-differentiated hPSNs revealed a gene expression program resembling in vivo CGE/LGE subpallial domains and GABAergic signaling. A combination of bioinformatic, functional, and immunocytochemical analysis further revealed that hPSNs consist of both cortical glutamatergic and CGE-like GABAergic neurons. This study provides a comprehensive characterization of the heterogeneous group of neurons produced by default differentiation and insight into future directed differentiation strategies.

Effects of Ethanol on Cellular Composition and Network Excitability of Human Pluripotent Stem Cell-Derived Neurons.

BACKGROUND: Prenatal alcohol exposure (PAE) in animal models results in excitatory-inhibitory (E/I) imbalance in neocortex due to alterations in the GABAergic interneuron (IN) differentiation and migration. Thus, E/I imbalance is a potential cause for intellectual disability in individuals with fetal alcohol spectrum disorder (FASD), but whether ethanol (EtOH) changes glutamatergic and GABAergic IN specification during human development remains unknown. Here, we created a human cellular model of PAE/FASD and tested the hypothesis that EtOH exposure during differentiation of human pluripotent stem cell-derived neurons (hPSNs) would cause the aberrant production of glutamatergic and GABAergic neurons, resulting in E/I imbalance. METHODS: We applied 50 mM EtOH daily to differentiating hPSNs for 50 days to model chronic first-trimester exposure. We used quantitative polymerase chain reaction, immunocytochemical, and electrophysiological analysis to examine the effects of EtOH on hPSN specification and functional E/I balance. RESULTS: We found that EtOH did not alter neural induction nor general forebrain patterning and had no effect on the expression of markers of excitatory cortical pyramidal neurons. In contrast, our data revealed highly significant changes to levels of transcripts involved with IN precursor development (e.g., GSX2, DLX1/2/5/6, NR2F2) as well as mature IN specification (e.g., SST, NPY). Interestingly, EtOH did not affect the number of GABAergic neurons generated nor the frequency or amplitude of miniature excitatory and inhibitory postsynaptic currents. CONCLUSIONS: Similar to in vivo rodent studies, EtOH significantly and specifically altered the expression of genes involved with IN specification from hPSNs, but did not cause imbalances of synaptic excitation-inhibition. Thus, our findings corroborate previous studies pointing to aberrant neuronal differentiation as an underlying mechanism of intellectual disability in FASD. However, in contrast to rodent binge models, our chronic exposure model suggests possible compensatory mechanisms that may cause more subtle defects of network processing rather than gross alterations in total E/I balance.

Differential roles of microglia and monocytes in the inflamed central nervous system.

In the human disorder multiple sclerosis (MS) and in the model experimental autoimmune encephalomyelitis (EAE), macrophages predominate in demyelinated areas and their numbers correlate to tissue damage. Macrophages may be derived from infiltrating monocytes or resident microglia, yet are indistinguishable by light microscopy and surface phenotype. It is axiomatic that T cell-mediated macrophage activation is critical for inflammatory demyelination in EAE, yet the precise details by which tissue injury takes place remain poorly understood. In the present study, we addressed the cellular basis of autoimmune demyelination by discriminating microglial versus monocyte origins of effector macrophages. Using serial block-face scanning electron microscopy (SBF-SEM), we show that monocyte-derived macrophages associate with nodes of Ranvier and initiate demyelination, whereas microglia appear to clear debris. Gene expression profiles confirm that monocyte-derived macrophages are highly phagocytic and inflammatory, whereas those arising from microglia demonstrate an unexpected signature of globally suppressed cellular metabolism at disease onset. Distinguishing tissue-resident macrophages from infiltrating monocytes will point toward new strategies to treat disease and promote repair in diverse inflammatory pathologies in varied organs.

Deletion of PTEN produces autism-like behavioral deficits and alterations in synaptic proteins.

Many genes have been implicated in the underlying cause of autism but each gene accounts for only a small fraction of those diagnosed with autism. There is increasing evidence that activity-dependent changes in neuronal signaling could act as a convergent mechanism for many of the changes in synaptic proteins. One candidate signaling pathway that may have a critical role in autism is the PI3K/AKT/mTOR pathway. A major regulator of this pathway is the negative repressor phosphatase and tensin homolog (PTEN). In the current study we examined the behavioral and molecular consequences in mice with neuron subset-specific deletion of PTEN. The knockout (KO) mice showed deficits in social chamber and social partition test. KO mice demonstrated alterations in repetitive behavior, as measured in the marble burying test and hole-board test. They showed no changes in ultrasonic vocalizations emitted on postnatal day 10 or 12 compared to wildtype (WT) mice. They exhibited less anxiety in the elevated-plus maze test and were more active in the open field test compared to WT mice. In addition to the behavioral alterations, KO mice had elevation of phosphorylated AKT, phosphorylated S6, and an increase in S6K. KO mice had a decrease in mGluR but an increase in total and phosphorylated fragile X mental retardation protein. The disruptions in intracellular signaling may be why the KO mice had a decrease in the dendritic potassium channel Kv4.2 and a decrease in the synaptic scaffolding proteins PSD-95 and SAP102. These findings demonstrate that deletion of PTEN results in long-term alterations in social behavior, repetitive behavior, activity, and anxiety. In addition, deletion of PTEN significantly alters mGluR signaling and many synaptic proteins in the hippocampus. Our data demonstrates that deletion of PTEN can result in many of the behavioral features of autism and may provide insights into the regulation of intracellular signaling on synaptic proteins.