Pathogenic SCN2A variants cause early-stage dysfunction in patient-derived neurons

Abstract

Pathogenic heterozygous variants in SCN2A, which encodes the neuronal sodium channel NaV1.2, cause different types of epilepsy or intellectual disability (ID)/autism without seizures. Previous studies using mouse models or heterologous systems suggest that NaV1.2 channel gain-of-function typically causes epilepsy, whereas loss-of-function leads to ID/autism. How altered channel biophysics translate into patient neurons remains unknown. Here, we investigated iPSC-derived early-stage cortical neurons from ID patients harboring diverse pathogenic SCN2A variants [p.(Leu611Valfs*35); p.(Arg937Cys); p.(Trp1716*)], and compared them to neurons from an epileptic encephalopathy patient [p.(Glu1803Gly)] and controls. ID neurons consistently expressed lower NaV1.2 protein levels. In neurons with the frameshift variant, NaV1.2 mRNA and protein levels were reduced by ~ 50%, suggesting nonsense-mediated decay and haploinsufficiency. In other ID neurons, only protein levels were reduced implying NaV1.2 instability. Electrophysiological analysis revealed decreased sodium current density and impaired action potential (AP) firing in ID neurons, consistent with reduced NaV1.2 levels. By contrast, epilepsy neurons displayed no change in NaV1.2 levels or sodium current density, but impaired sodium channel inactivation. Single-cell transcriptomics identified dysregulation of distinct molecular pathways including inhibition of oxidative phosphorylation in neurons with SCN2A haploinsufficiency, and activation of calcium signaling and neurotransmission in epilepsy neurons. Together, our patient iPSC-derived neurons reveal characteristic sodium channel dysfunction consistent with biophysical changes previously observed in heterologous systems. Additionally, our model links the channel dysfunction in ID to reduced NaV1.2 levels and uncovers impaired AP firing in early-stage neurons. The altered molecular pathways may reflect a homeostatic response to NaV1.2 dysfunction and can guide further investigations.

Crosstalk among WEE1 Kinase, AKT, and GSK3 in Nav1.2 Channelosome Regulation

“…Because WEE1 exerts specific regulation on Nav1.2 but not Nav1.6 channels, unbalanced levels of WEE1 could perturb the developmental switch between Nav1.2 and Nav1.6 isoforms, delaying or accelerating neuronal maturation with consequences for synaptic integration and plasticity. Both WEE1 and FGF14 have been associated with schizophrenia and other neurodevelopmental disorders [42][43][44][45][46][47].Thus, WEE1 may be part of a signaling pathway, including FGF14 and Nav1.2, that if perturbed, could contribute to endophenotypes related to neurodevelopmental disorders such as schizophrenia, autism spectrum disorders and SCN2A channelopathies [28][29][30][31][32][33][34][35].…”

“…Overall, these findings indicate that the FGF14/Nav1.2 signalosome involves a connecting mode of WEE1-dependent AKT/GSK3 signaling pathways. This study could provide insights into the signaling mechanisms underlying neurodevelopmental disorders associated with Nav1.2 channelopathies [28][29][30][31][32][33][34][35], aiding in the development of targeted therapies for these conditions.…”

A deep learning framework for automated and generalized synaptic event analysis

“…Spontaneous EPSCs were recorded from human iPSC-derived neurons in a similar fashion, but ACSF consisted of (in mM): 135 NaCl, 10 HEPES, 10 D-glucose, 5 KCl, 2 CaCl 2 , 1 MgCl 2 . We recorded from 8-week-old neuronal cultures as described in (Asadollahi et al, 2023). Synaptic events were observed in ∼ 51% of neurons.…”

Scn2a insufficiency alters spontaneous neuronal Ca2+ activity in somatosensory cortex during wakefulness

“… 12 Recently, patient-derived neurons which expressed a stop variant were found to mediate smaller current and fire less AP than isogenic controls. 16 However, the consequences of these in vitro cellular changes on brain network remains unclear.…”