Abstract
BackgroundFragile X syndrome (FXS), a neurodevelopmental disorder, is a leading monogenetic cause of intellectual disability and autism spectrum disorder. Notwithstanding the extensive studies using rodent and other pre-clinical models of FXS, which have provided detailed mechanistic insights into the pathophysiology of this disorder, it is only relatively recently that human stem cell-derived neurons have been employed as a model system to further our understanding of the pathophysiological events that may underlie FXS. Our study assesses the physiological properties of human pluripotent stem cell-derived cortical neurons lacking fragile X mental retardation protein (FMRP).MethodsElectrophysiological whole-cell voltage- and current-clamp recordings were performed on two control and three FXS patient lines of human cortical neurons derived from induced pluripotent stem cells. In addition, we also describe the properties of an isogenic pair of lines in one of which FMR1 gene expression has been silenced.ResultsNeurons lacking FMRP displayed bursts of spontaneous action potential firing that were more frequent but shorter in duration compared to those recorded from neurons expressing FMRP. Inhibition of large conductance Ca2+-activated K+ currents and the persistent Na+ current in control neurons phenocopies action potential bursting observed in neurons lacking FMRP, while in neurons lacking FMRP pharmacological potentiation of voltage-dependent Na+ channels phenocopies action potential bursting observed in control neurons. Notwithstanding the changes in spontaneous action potential firing, we did not observe any differences in the intrinsic properties of neurons in any of the lines examined. Moreover, we did not detect any differences in the properties of miniature excitatory postsynaptic currents in any of the lines.ConclusionsPharmacological manipulations can alter the action potential burst profiles in both control and FMRP-null human cortical neurons, making them appear like their genetic counterpart. Our studies indicate that FMRP targets that have been found in rodent models of FXS are also potential targets in a human-based model system, and we suggest potential mechanisms by which activity is altered.
Highlights
Fragile X syndrome (FXS), a neurodevelopmental disorder, is a leading monogenetic cause of intellectual disability and autism spectrum disorder
Given the large number of fragile X mental retardation protein (FMRP) targets that are associated with synaptic function and the well-described synaptic and network properties that are dysregulated in pre-clinical models of FXS, our present study focuses on assessing the physiological properties of two control (CON1, CON2) and three FXS patient lines (FXS1, FXS2, FXS3) of human cortical neurons derived from induced pluripotent stem cells
Absence of FMRP does not affect differentiation efficiency of FMRP-lacking neurons Fibroblast-derived induced pluripotent stem cells (iPSCs) were generated from two healthy individuals (CON1 and CON2), three FXS patients lacking FMRP (FXS1, FXS2 and FXS3) and one isogenic embryonic stem cell (FMR1+/y; FMR1 gene genetically deleted (FMR1−/y)) pair where the FMR1 gene was deleted using CRISPR/Cas9-mediated genome editing [24]
Summary
Fragile X syndrome (FXS), a neurodevelopmental disorder, is a leading monogenetic cause of intellectual disability and autism spectrum disorder. It is not always the case that all neuronal populations show hyperexcitability; a recent study using mouse primary cortical neurons demonstrated that loss of FMRP did not affect the basal neuronal excitability [15] while a study using foetal rat visual cortex showed the Fmr null neurons to be hypoexcitable [16]. Such results which may be taken by some to be conflicting but could illustrate the complexity of studying FXS pathophysiology in rodent models
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