OBJECTIVE Neuronal channelopathies cause brain disorders including epilepsy, migraine and ataxia. Despite the development of mouse models, pathophysiological mechanisms for these disorders remain uncertain. One particularly devastating channelopathy is Dravet Syndrome (DS), a severe childhood epilepsy typically caused by de novo dominant mutations in the SCN1A gene encoding the voltage-gated sodium channel Nav1.1. Heterologous expression of mutant channels suggests loss-of-function, raising the quandary of how loss of sodium channels underlying action potentials produces hyperexcitability. Mouse model studies suggest that decreased Nav1.1 function in interneurons causes disinhibition. We sought to determine how mutant SCN1A affects human neurons using the induced pluripotent stem cell (iPSC) method to generate patient-specific neurons. METHODS Forebrain-like pyramidal- and bipolar-shaped neurons are derived from two DS subjects and three human controls by iPSC reprogramming of fibroblasts. DS and control iPSC-derived neurons are compared using whole-cell patch clamp recordings. Sodium current density and intrinsic neuronal excitability are examined. RESULTS Neural progenitors from DS and human control iPSCs display a forebrain identity and differentiate into bipolar- and pyramidal-shaped neurons. DS patient-derived neurons show increased sodium currents in both bipolar- and pyramidal-shaped neurons. Consistent with increased sodium currents, both types of patient-derived neurons show spontaneous bursting and other evidence of hyperexcitability. Sodium channel transcripts are not elevated, consistent with a post-translational mechanism. INTERPRETATION These data demonstrate that epilepsy patient-specific iPSC-derived neurons are useful for modeling epileptic-like hyperactivity. Our findings reveal a previously unrecognized cell-autonomous epilepsy mechanism potentially underlying Dravet Syndrome, and offer a platform for screening new anti-epileptic therapies.

Bovine Kir7.1 clones were obtained from a retinal pigment epithelium (RPE)‐subtracted cDNA library. Human RPE cDNA library screening resulted in clones encoding full‐length human Kir7.1. Northern blot analysis indicated that bovine Kir7.1 is highly expressed in the RPE. Human Kir7.1 channels were expressed in Xenopus oocytes and studied using the two‐electrode voltage‐clamp technique. The macroscopic Kir7.1 conductance exhibited mild inward rectification and an inverse dependence on extracellular K+ concentration ([K+]o). The selectivity sequence based on permeability ratios was K+ (1.0) ≈ Rb+ (0.89) > Cs+ (0.013) > Na+ (0.003) ≈ Li+ (0.001) and the sequence based on conductance ratios was Rb+ (9.5) >> K+ (1.0) > Na+ (0.458) > Cs+ (0.331) > Li+ (0.139). Non‐stationary noise analysis of Rb+ currents in cell‐attached patches yielded a unitary conductance for Kir7.1 of ≈2 pS. In whole‐cell recordings from freshly isolated bovine RPE cells, the predominant current was a mild inwardly rectifying K+ current that exhibited an inverse dependence of conductance on [K+]o. The selectivity sequence based on permeability ratios was K+ (1.0) ≈ Rb+ (0.89) > Cs+ (0.021) > Na+ (0.003) ≈ Li+ (0.002) and the sequence based on conductance ratios was Rb+ (8.9) >> K+ (1.0) > Na+ (0.59) > Cs+ (0.23) > Li+ (0.08). In cell‐attached recordings with Rb+ in the pipette, inwardly rectifying currents were observed in nine of 12 patches of RPE apical membrane but in only one of 13 basolateral membrane patches. Non‐stationary noise analysis of Rb+ currents in cell‐attached apical membrane patches yielded a unitary conductance for RPE Kir of ≈2 pS. On the basis of this molecular and electrophysiological evidence, we conclude that Kir7.1 channel subunits comprise the K+ conductance of the RPE apical membrane.

Voltage-gated Na + channel (VGSC) β1 subunits, encoded by SCN1B, are multifunctional channel modulators and cell adhesion molecules (CAMs). Mutations in SCN1B are associated with the genetic epilepsy with febrile seizures plus (GEFS+) spectrum disorders in humans, and Scn1b-null mice display severe spontaneous seizures and ataxia from postnatal day (P)10. The goal of this study was to determine changes in neuronal pathfinding during early postnatal brain development of Scn1b-null mice to test the hypothesis that these CAM-mediated roles of Scn1b may contribute to the development of hyperexcitability. c-Fos, a protein induced in response to seizure activity, was up-regulated in the Scn1b-null brain at P16 but not at P5. Consistent with this, epileptiform activity was observed in hippocampal and cortical slices prepared from the P16 but not from the P5-P7 Scn1b-null brain. On the basis of these results, we investigated neuronal pathfinding at P5. We observed disrupted fasciculation of parallel fibers in the P5 null cerebellum. Further, P5 null mice showed reduced neuron density in the dentate gyrus granule cell layer, increased proliferation of granule cell precursors in the hilus, and defective axonal extension and misorientation of somata and processes of inhibitory neurons in the dentate gyrus and CA1. Thus, Scn1b is critical for neuronal proliferation, migration, and pathfinding during the critical postnatal period of brain development. We propose that defective neuronal proliferation, migration, and pathfinding in response to Scn1b deletion may contribute to the development of hyperexcitability.β subunit | sodium channel | hippocampus N euronal voltage-gated Na + channels (VGSCs) are composed of one pore-forming α and two β subunits (1). β1 (encoded by SCN1B) modulates channel gating and cell surface expression (2). In addition, β1 is an Ig superfamily cell adhesion molecule (CAM) that participates in cell-cell and cell-matrix adhesion (3). The SCN1B splice variant β1B is a secreted CAM that is expressed predominantly during embryonic brain development (4). SCN1B mutations are associated with the spectrum of genetic epilepsy with febrile seizures plus (GEFS+) (OMIM 604233) epilepsies (5, 6). At the cellular level, SCN1B GEFS+ mutations cause a range of defects, including altered channel availability, disrupted adhesion (7), exclusion of β1 from the axon initial segment (AIS) of pyramidal neurons (8), reduced cell surface expression of β1, and intracellular retention of β1B (4, 9). In addition, a GEFS+ mutation in SCN1A decreases modulation of Na v 1.1 by β1 (10). Together, these results suggest a causal relationship between VGSC α-β1 interactions, cell-cell adhesion, and epilepsy.Scn1b-null mice display severe spontaneous seizures and ataxia from approximately postnatal day (P)10 (11). Scn1b deletion reduces resurgent Na + current (I Na ) in P14-P16 cerebellar granule neurons (CGNs) (12). In contrast, β1 has no effect on I Na in dissociated P14-P16 hippocampal neurons (9). However, CA3 action potentials have ...