Since the discovery of somatic reprogramming, human induced pluripotent stem cells (hiPSCs) have been exploited to model a variety of neurological and psychiatric disorders. Because hiPSCs represent an almost limitless source of patient-derived neurons that retain the genetic variations thought to contribute to disease etiology, they have been heralded as a patient-specific platform for high throughput drug screening. However, the utility of current protocols for generating neurons from hiPSCs remains limited by protracted differentiation timelines and heterogeneity of the neuronal phenotypes produced. Neuronal induction via the forced expression of exogenous transcription factors rapidly induces defined populations of functional neurons from fibroblasts and hiPSCs. Here, we describe an adapted protocol that accelerates maturation of functional excitatory neurons from hiPSC-derived neural progenitor cells (NPCs) via lentiviral transduction of Neurogenin 2 (using both mNgn2 and hNGN2). This methodology, relying upon a robust and scalable starting population of hiPSC NPCs, should be readily amenable to scaling for hiPSC-based high-throughput drug screening.
SUMMARY Gain-of-function (GOF) variants in K + channels cause severe childhood epilepsies, but there are no mechanisms to explain how increased K + currents lead to network hyperexcitability. Here, we introduce a human Na + -activated K + (K Na ) channel variant ( KCNT1 -Y796H) into mice and, using a multiplatform approach, find motor cortex hyperexcitability and early-onset seizures, phenotypes strikingly similar to those of human patients. Although the variant increases K Na currents in cortical excitatory and inhibitory neurons, there is an increase in the K Na current across subthreshold voltages only in inhibitory neurons, particularly in those with non-fast-spiking properties, resulting in inhibitory-neuron-specific impairments in excitability and action potential (AP) generation. We further observe evidence of synaptic rewiring, including increases in homotypic synaptic connectivity, accompanied by network hyperexcitability and hypersynchronicity. These findings support inhibitory-neuron-specific mechanisms in mediating the epileptogenic effects of KCNT1 channel GOF, offering cell-type-specific currents and effects as promising targets for therapeutic intervention.
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