Defective AMPA-mediated synaptic transmission and morphology in human neurons with hemizygous SHANK3 deletion engrafted in mouse prefrontal cortex

Chiola, Simone; Napan, Kandy L; Wang, Yueqi; Lazarenko, Roman M.; Armstrong, Celeste J.; Cui, Jun; Shcheglovitov, Aleksandr

doi:10.1038/s41380-021-01023-2

Cited by 15 publications

(13 citation statements)

References 77 publications

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“…Stem cell-derived neurons were generated from embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) using a previously described approach ( Shcheglovitov et al, 2013 ; Chiola et al, 2021 ). Stem cell lines used in this study include previously validated control lines: H9 (ESC line), and Coriell (iPSCs were derived from Coriell GM07492 fibroblasts and provided by Carl Ernst as a kind gift; Bell et al, 2018 ).…”

Section: Methodsmentioning

confidence: 99%

Secreted Reporter Assay Enables Quantitative and Longitudinal Monitoring of Neuronal Activity

Santos

Chiola

Yang

et al. 2021

eNeuro

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The ability to measure changes in neuronal activity in a quantifiable and precise manner is of fundamental importance to understand neuron development and function.Repeated monitoring of neuronal activity of the same population of neurons over several days is challenging and, typically, low-throughput. Here, we describe a new biochemical reporter assay that allows for repeated measurements of neuronal activity in a cell type-specific manner. We coupled activity-dependent elements from the Arc/Arg3.1 gene with a secreted reporter, Gaussia luciferase, to quantify neuronal activity without sacrificing the neurons. The reporter predominantly senses calcium and NMDA receptor-dependent activity. By repeatedly measuring the accumulation of the reporter in cell media, we can profile the developmental dynamics of neuronal activity in cultured neurons from male and female mice. The assay also allows for longitudinal analysis of pharmacological treatments, thus distinguishing acute from delayed responses. Moreover, conditional expression of the reporter allows for monitoring cell type-specific changes. This simple, quantitative, cost-effective, automatable, and cell type-specific activity reporter is a valuable tool to study the development of neuronal activity in normal and disease-model conditions, and to identify small molecules or protein factors that selectively modulate the activity of a specific population of neurons.

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Section: Methodsmentioning

confidence: 99%

Secreted Reporter Assay Enables Quantitative and Longitudinal Monitoring of Neuronal Activity

Santos

Chiola

Yang

et al. 2021

eNeuro

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“…Xenotransplantation of human stem cell-derived neurons has been used most often to study neurodegenerative disorders and develop cell-based therapies for Parkinson’s disease [ 87 ], Alzheimer’s disease [ 88 ], stroke [ 89 ], epilepsy [ 90 ], and spinal cord injury [ 91 ]. Fewer studies have used xenotransplantation to investigate connectivity in neurodevelopmental disorders [ 92 , 93 ]. Upon transplantation of iPSC-derived cortical excitatory neurons from Down syndrome (DS) patients into the somatosensory cortex of adult mice, longitudinal in vivo GCaMP6 imaging showed that DS neurons develop more stable dendritic spines and significantly increased spine density than isogenic control neurons [ 92 ].…”

Section: Xenotransplantation and Organoid Models For Studying Connect...mentioning

confidence: 99%

“…Transplantation of SHANK3-deficient human cortical neurons into the mouse prefrontal cortex revealed that SHANK3 deficiency causes different synaptic deficits at different developmental stages [ 93 ]. Initially, the strength of excitatory AMPA receptor-mediated synaptic currents is reduced.…”

Section: Xenotransplantation and Organoid Models For Studying Connect...mentioning

confidence: 99%

iPSC toolbox for understanding and repairing disrupted brain circuits in autism

2021

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Over the past decade, tremendous progress has been made in defining autism spectrum disorder (ASD) as a disorder of brain connectivity. Indeed, whole-brain imaging studies revealed altered connectivity in the brains of individuals with ASD, and genetic studies identified rare ASD-associated mutations in genes that regulate synaptic development and function. However, it remains unclear how specific mutations alter the development of neuronal connections in different brain regions and whether altered connections can be restored therapeutically. The main challenge is the lack of preclinical models that recapitulate important aspects of human development for studying connectivity. Through recent technological innovations, it is now possible to generate patient- or mutation-specific human neurons or organoids from induced pluripotent stem cells (iPSCs) and to study altered connectivity in vitro or in vivo upon xenotransplantation into an intact rodent brain. Here, we discuss how deficits in neurodevelopmental processes may lead to abnormal brain connectivity and how iPSC-based models can be used to identify abnormal connections and to gain insights into underlying cellular and molecular mechanisms to develop novel therapeutics.

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“…Cultured neurons lack a physiologically relevant environment. Grafting of human neurons into rodent brains has been used to study human neuronal development and functionality 26,38 . We transplanted control and NLGN3 R451C-mutant excitatory human neurons expressing mCherry and mVenus, respectively, together into the hippocampus of neonatal (postnatal day 0-3) RAG2 immunocompromised mice (Fig.…”

Section: The Nlgn3 R451c-mutation Increases Excitatory Synaptic Connectivity Of Human Neurons Transplanted Into Mouse Brainmentioning

confidence: 99%

“…The development of efficient methods for generating neurons from human pluripotent stem cells [20][21][22][23] has provided an opportunity to use human neurons form modeling neuropsychiatric disorders [24][25][26][27] . Notably, mutations in the NRXN1 28,29 and in NLGN4 30,31 genes in human neurons have uncovered intriguing functional changes that appear to be specific to human neurons.…”

Section: Introduction (1500)mentioning

confidence: 99%

Analyses of the Autism-associated Neuroligin-3 R451C Mutation in Human Neurons Reveals a Gain-of-Function Synaptic Mechanism

Wang

Mirabella

Dai

et al. 2021

Preprint

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Mutations in many synaptic genes are associated with autism spectrum disorders (ASDs), suggesting that synaptic dysfunction is a key driver of ASD pathogenesis. Among these mutations, the R451C-substitution in the NLGN3 gene that encodes the postsynaptic adhesion molecule Neuroligin-3 is noteworthy because it was the first specific mutation linked to ASDs. In mice, the corresponding Nlgn3 R451C-knockin mutation recapitulates social interaction deficits of ASD patients and produces synaptic abnormalities, but the impact of the NLGN3 R451C-mutation on human neurons has not been investigated. Here, we generated human knock-in neurons with the NLGN3 R451C-mutation. Strikingly, analyses of NLGN3 R451C-mutant neurons revealed that the R451C-mutation decreased NLGN3 protein levels but enhanced the strength of excitatory synapses without affecting inhibitory synapses. No significant cell death and endoplasmic reticulum stress were detected. Importantly, the augmentation of excitatory transmission was confirmed in vivo with human neurons transplanted into mouse forebrain. Using single-cell RNA-seq experiments with co-cultured excitatory and inhibitory NLGN3 R451C-mutant neurons, we identified differentially expressed genes in relatively mature human neurons that corresponded to synaptic gene expression networks. Moreover, gene ontology and enrichment analyses revealed convergent gene networks associated with ASDs and other mental disorders. Our findings suggest that the NLGN3 R451C-mutation induces a gain-of-function enhancement in excitatory synaptic transmission that may contribute to the pathophysiology of ASDs.

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Defective AMPA-mediated synaptic transmission and morphology in human neurons with hemizygous SHANK3 deletion engrafted in mouse prefrontal cortex

Cited by 15 publications

References 77 publications

Secreted Reporter Assay Enables Quantitative and Longitudinal Monitoring of Neuronal Activity

Secreted Reporter Assay Enables Quantitative and Longitudinal Monitoring of Neuronal Activity

iPSC toolbox for understanding and repairing disrupted brain circuits in autism

Analyses of the Autism-associated Neuroligin-3 R451C Mutation in Human Neurons Reveals a Gain-of-Function Synaptic Mechanism

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