De novo Y1460C missense variant in NaV1.1 impedes the pore region and results in epileptic encephalopathy.

Epilepsy is a common neurological disorder characterized by recurrent unprovoked seizures. SCN1A encodes NaV1.1, a neuronal voltage-gated Na+ channel that is highly expressed throughout the central nervous system. NaV1.1 is localized within the axon initial segment where it plays a critical role in the initiation and propagation of action potentials and neuronal firing, predominantly in γ-amino-butyric-acid (GABA)ergic neurons of the hippocampus. The objective of this study was to characterize a de novo missense variant of uncertain significance in the SCN1A gene of a proband presented with febrile status epilepticus characterized by generalized tonic clonic movements associated with ictal emesis and an abnormal breathing pattern. Screening a gene panel revealed a heterozygous missense variant of uncertain significance in the SCN1A gene, designated c.4379A>G, p.(Tyr1460Cys). The NaV1.1 wild-type (WT) and mutant channel reproduced in vivo and were transfected in HEK 293 cells. Na+ currents were recorded using the whole-cell configuration of the patch-clamp technique. This NaV1.1 variant (Tyr1460Cys) failed to express functional Na+ currents when expressed in HEK293 cells, most probably due to a pore defect of the channel given that the cell surface expression of the channel was normal. Currents generated after co-transfection with functional WT channels exhibited biophysical properties comparable to those of WT channels, which was mainly due to the functional WT channels at the cell surface. The NaV1.1 variant failed to express functional Na+ currents, most probably due to pore impairment and exhibited a well-established loss of function mechanism. The present study highlights the added-value of functional testing for understanding the pathophysiology and potential treatment decisions for patients with undiagnosed developmental epileptic encephalopathy.

NPRL2 Inhibition of mTORC1 Controls Sodium Channel Expression and Brain Amino Acid Homeostasis.

Genetic mutations in nitrogen permease regulator-like 2 (NPRL2) are associated with a wide spectrum of familial focal epilepsies, autism, and sudden unexpected death of epileptics (SUDEP), but the mechanisms by which NPRL2 contributes to these effects are not well known. NPRL2 is a requisite subunit of the GAP activity toward Rags 1 (GATOR1) complex, which functions as a negative regulator of mammalian target of rapamycin complex 1 (mTORC1) kinase when intracellular amino acids are low. Here, we show that loss of NPRL2 expression in mouse excitatory glutamatergic neurons causes seizures before death, consistent with SUDEP in humans with epilepsy. Additionally, the absence of NPRL2 expression increases mTORC1-dependent signal transduction and significantly alters amino acid homeostasis in the brain. Loss of NPRL2 reduces dendritic branching and increases the strength of electrically stimulated action potentials (APs) in neurons. The increased AP strength is consistent with elevated expression of epilepsy-linked, voltage-gated sodium channels in the NPRL2-deficient brain. Targeted deletion of NPRL2 in primary neurons increases the expression of sodium channel Scn1A, whereas treatment with the pharmacological mTORC1 inhibitor called rapamycin prevents Scn1A upregulation. These studies demonstrate a novel role of NPRL2 and mTORC1 signaling in the regulation of sodium channels, which can contribute to seizures and early lethality.

Novel G1481V and Q1491H SCN5A Mutations Linked to Long QT Syndrome Destabilize the Nav1.5 Inactivation State.

BACKGROUND: Nav1.5, which is encoded by the SCN5A gene, is the predominant voltage-gated Na+ channel in the heart. Several mutations of this gene have been identified and reported to be involved in several cardiac rhythm disorders, including type 3 long QT interval syndrome, that can cause sudden cardiac death. We analyzed the biophysical properties of 2 novel variants of the Nav1.5 channel (Q1491H and G1481V) detected in 5- and 12-week-old infants diagnosed with a prolonged QT interval. METHODS: The Nav1.5 wild-type and the Q1491H and G1481V mutant channels were reproduced in vi tr o. Wild-type or mutant channels were cotransfected in human embryonic kidney (HEK) 293 cells with the beta 1 regulatory subunit. Na+ currents were recorded using the whole-cell configuration of the patch-clamp technique. RESULTS: The Q1491H mutant channel exhibited a lower current density, a persistent Na+ current, an enhanced window current due to a +20-mV shift of steady-state inactivation, a +10-mV shift of steady-state activation, a faster onset of slow inactivation, and a recovery from fast inactivation with fast and slow time constants of recovery. The G1481V mutant channel exhibited an increase in current density and a +7-mV shift of steady-state inactivation. The observed defects are characteristic of gain-of-function mutations typical of type 3 long QT interval syndrome. CONCLUSIONS: The 5- and 12-week-old infants displayed prolonged QT intervals. Our analyses of the Q1491H and G1481V mutations correlated with the clinical diagnosis. The observed biophysical dysfunctions associated with both mutations were most likely responsible for the sudden deaths of the 2 infants.

INTRODUCTION: Le canal Nav1.5, codé par le gène SCN5A, est le canal Na+ dépendant du voltage prédominant dans le cœur. Plusieurs mutations de ce gène sont impliquées dans plusieurs anomalies du rythme cardiaque, dont le syndrome du QT long de type 3, qui peut provoquer la mort subite d'origine cardiaque. Nous avons analysé les propriétés biophysiques de deux nouveaux variants du canal Nav1.5 (Q1491H et G1481V) détectés chez deux bébés âgés respectivement de 5 et 12 semaines qui avaient une prolongation de l'intervalle QT. MÉTHODES: Le canal Nav1.5 de type sauvage et les canaux mutants Q1491H et G1481V ont été reproduits in vi tr o. Les canaux de type sauvage ou mutants ont été co-transfectés dans les cellules des reins embryonnaires humains (REH) 293 avec la sous-unité régulatrice bêta 1. Les courants Na+ ont été enregistrés à partir de la configuration en cellule entière via la technique de patch-clamp. RÉSULTATS: Le canal mutant Q1491H montre une densité de courant plus faible, un courant Na+ persistant, un courant fenêtre augmenté en raison d'un changement dép de +20 mV de l'inactivation à l'état stable, un changement de +10 mV de l'activation à l'état stable, une entrée plus rapide de l'inactivation lente et une récupération de l'inactivation rapide avec des constantes de temps rapides et lentes. Le canal mutant G1481V montre une augmentation de la densité de courant et un changement de +7 mV de l'inactivation à l'état stable. Les anomalies observées sont caractéristiques des mutations avec gain de fonction typiques du syndrome du QT long de type 3. CONCLUSIONS: Les deux bébés âgés respectivement de cinq 5 et 12 semaines montraient une prolongation des intervalles QT. Nos analyses des mutations Q1491H et G1481V montrent une corrélation avec le diagnostic clinique. Les dysfonctions biophysiques observées qui sont associées aux deux mutations étaient fort probablement responsables des morts subites des deux bébés.