SCN1A intronic variants impact on Nav1.1 protein expression and sodium channel function, and associated with epilepsy phenotypic severity.

High-throughput sequencing has identified numerous intronic variants in the SCN1A gene in epilepsy patients. Abnormal mRNA splicing caused by these variants can lead to significant phenotypic differences, but the mechanisms of epileptogenicity and phenotypic differences remain unknown. Two variants, c.4853-1 G>C and c.4853-25 T>A, were identified in intron 25 of SCN1A, which were associated with severe Dravet syndrome (DS) and mild focal epilepsy with febrile seizures plus (FEFS+), respectively. The impact of these variants on protein expression, electrophysiological properties of sodium channels and their correlation with epilepsy severity was investigated through plasmid construction and transfection based on the aberrant spliced mRNA. We found that the expression of truncated mutant proteins was significantly reduced on the cell membrane, and retained in the cytoplasmic endoplasmic reticulum. The mutants caused a decrease in current density, voltage sensitivity, and an increased vulnerability of channel, leading to a partial impairment of sodium channel function. Notably, the expression of DS-related mutant protein on the cell membrane was higher compared to that of FEFS+-related mutant, whereas the sodium channel function impairment caused by DS-related mutant was comparatively milder than that caused by FEFS+-related mutant. Our study suggests that differences in protein expression levels and altered electrophysiological properties of sodium channels play important roles in the manifestation of diverse epileptic phenotypes. The presence of intronic splice site variants may result in severe phenotypes due to the dominant-negative effects, whereas non-canonical splice site variants leading to haploinsufficiency could potentially cause milder phenotypes.

Early developmental alterations of CA1 pyramidal cells in Dravet syndrome.

Dravet Syndrome (DS) is most often caused by heterozygous loss-of-function mutations in the voltage-gated sodium channel gene SCN1A (Nav1.1), resulting in severe epilepsy and neurodevelopmental impairment thought to be cause by reduced interneuron excitability. However, recent studies in mouse models suggest that interneuron dysfunction alone does not completely explain all the cellular and network impairments seen in DS. Here, we investigated the development of the intrinsic, synaptic, and network properties of CA1 pyramidal cells in a DS model prior to the appearance of overt seizures. We report that CA1 pyramidal cell development is altered by heterozygous reduction of Scn1a, and propose that this is explained by a period of reduced intrinsic excitability in early postnatal life, during which Scn1a is normally expressed in hippocampal pyramidal cells. We also use a novel ex vivo model of homeostatic plasticity to show an instability in homeostatic response during DS epileptogenesis. This study provides evidence for the early effects of Scn1a haploinsufficiency in pyramidal cells in contributing to the pathophysiology of DS.

POLR3B de novo variants are a rare cause of infantile myoclonic epilepsy.

PURPOSE: To report on a new phenotype in a patient carrying a novel, undescribed de novo variant in POLR3B, affected by generalized myoclonic epilepsy and neurodevelopmental disorder, without neuropathy. It is known that biallelic pathogenic variants in POLR3B cause hypomyelinating leukodystrophy-8, and heterozygous de novo variants are described in association to a phenotype characterized by predominantly demyelinating sensory-motor peripheral neuropathy, ataxia, spasticity, intellectual disability and epilepsy, in which the peripheral neuropathy is often the main clinical presentation. METHODS: We collected clinical, electrophysiological and neuroimaging data from the affected subject and performed a Trio-Clinical Exome Sequencing. RESULTS: We detected a de novo novel heterozygous missense variant c.1132A>G in POLR3B (NM_018082.6) that was considered as likely pathogenic following ACMG criteria. We also consulted our custom genomic database of a total of 1485 patients that were genetically analysed from 2018 for epilepsy, and found no other de novo variants in the POLR3B gene. CONCLUSION: We hypothesize a possible genotype-phenotype correlation, particularly regarding epilepsy. We also provide a review of the literature about the previously described POLR3B heterozygous patients, with particular attention to the epileptic phenotype, underlining the association between POLR3B and early onset myoclonic epilepsy, which can represent the main manifestation of the disease at its onset.

Epigenetic insights into GABAergic development in Dravet Syndrome iPSC and therapeutic implications.

Dravet syndrome (DS) is a devastating early-onset refractory epilepsy syndrome caused by variants in the SCN1A gene. A disturbed GABAergic interneuron function is implicated in the progression to DS but the underlying developmental and pathophysiological mechanisms remain elusive, in particularly at the chromatin level. Induced pluripotent stem cells (iPSCs) derived from DS cases and healthy donors were used to model disease-associated epigenetic abnormalities of GABAergic development. Chromatin accessibility was assessed at multiple time points (Day 0, Day 19, Day 35, and Day 65) of GABAergic differentiation. Additionally, the effects of the commonly used anti-seizure drug valproic acid (VPA) on chromatin accessibility were elucidated in GABAergic cells. The distinct dynamics in the chromatin profile of DS iPSC predicted accelerated early GABAergic development, evident at D19, and diverged further from the pattern in control iPSC with continued differentiation, indicating a disrupted GABAergic maturation. Exposure to VPA at D65 reshaped the chromatin landscape at a variable extent in different iPSC-lines and rescued the observed dysfunctional development of some DS iPSC-GABA. The comprehensive investigation on the chromatin landscape of GABAergic differentiation in DS-patient iPSC offers valuable insights into the epigenetic dysregulations associated with interneuronal dysfunction in DS. Moreover, the detailed analysis of the chromatin changes induced by VPA in iPSC-GABA holds the potential to improve the development of personalized and targeted anti-epileptic therapies.

Navigating Dravet syndrome in Spain: A cross-sectional study of diagnosis, management, and care coordination.

OBJECTIVES: Dravet syndrome (DS) is a rare form of refractory epilepsy that begins in the first year of life. Approximately 85% of patients have a mutation in the SCN1A gene, which encodes a voltage-gated sodium channel. The main objective of the present work was to assess the degree of knowledge of DS among Spanish primary care (PC) professionals, the communication flow between them and the pediatric neurologists (PNs), and the services available and resources offered to patients in Spain when searching for a diagnosis and adequate treatment. METHODS: Two anonymized online surveys on DS diagnosis and patient management in PC were conducted with Spanish PC pediatricians (PCPs) and caregivers of DS patients in Spain. RESULTS: Most PCPs are aware of genetic epilepsy but lack full knowledge of DS and patient advocacy groups (PAGs). Access to epilepsy treatments varies among regions, with many referrals to hospitals and pediatric neurologists. Diagnosis is often delayed, with misdiagnoses and frequent emergency room (ER) visits. Treatment involves multiple drugs, and sodium channel blockers are used, which are contraindicated in DS treatment. Improved training, resources, and communication are needed for early diagnosis. SIGNIFICANCE: To improve the care and treatment of DS patients in Spain, early diagnosis is required and, possibly, specific efforts aimed at identifying patients in adulthood, generating socio-sanitary structures that integrate social and health services to provide comprehensive care, taking into account the different features and comorbidities of the disease. PLAIN LANGUAGE SUMMARY: Dravet syndrome (DS) is a form of genetic epilepsy that starts within the first year of life. We present a study showing that, while family doctors are aware of genetic epilepsies, many don't have a complete understanding of DS. Unfortunately, getting the right diagnosis can take a long time, leading to unnecessary visits to the emergency room. Patients often need several medications, and sometimes they're given drugs that aren't recommended for DS. The takeaway is that training for doctors, more resources, and improved communication could help creating better healthcare systems and therefore give easier access to the right therapies.

A tool for Dravet syndrome-associated neuropsychiatric comorbidities evaluation (DANCE).

BACKGROUND: Dravet syndrome (DS) is a rare and severe form of epilepsy that begins in infancy, which is primarily caused by pathogenic variants in the SCN1A gene. DS is characterized by prolonged and frequent drug-resistant seizures, as well as developmental delays and behavioral problems. The identification of these comorbidities is based on clinical interview and relies on healthcare professionals (HCPs) experience. METHODS: We assembled a group of expert HCPs and caregivers to create a screening checklist for assessing DS-Associated Neuropsychiatric Comorbidities (DANC). The checklist includes questions related to cognitive and psychiatric domains, motor skills, and the impact of DS on families' daily lives. We administered the checklist to 24 caregivers of DS patients from Belgium, France, and Spain. After piloting, we obtained feedback from expert HCPs and caregivers to refine the checklist. RESULTS: DS patients showed a wide array of neuropsychiatric symptoms related to DS. The most common cognitive domains reported were attention difficulties and multitasking problems (18/24 caregivers), and impulsivity (17/24), while the most common psychiatric symptoms were temper tantrums (14/24), mood swings (13/24) and autism spectrum disorder (12/24). Balance and coordination problem have been reported in almost all patients with a statement of only 4/23 with complete mobility. Most patients were dependent on others for self-care and eating, and presented sleeping disturbances. Caregivers reported high levels of stress in the family unit, both between siblings and parents. Results show that the main concerns of parents were the behavior and the cognition of the person with DS. The quantitative feedback results showed good-to-very good scores on usefulness, ease of completion, clarity and comprehensiveness of the checklist. CONCLUSIONS: This pilot study suggests that the DANCE checklist could be a useful screening tool in daily practice for neuropsychiatric comorbidities facilitating their diagnosis and treatment, and empowering both caregivers and patients.

Genotypic and phenotypic characteristics of sodium channel-associated epilepsy in Chinese population.

Variants in voltage-gated sodium channel (VGSC) genes are implicated in seizures, epilepsy, and neurodevelopmental disorders, constituting a significant aspect of hereditary epilepsy in the Chinese population. Through retrospective analysis utilizing next-generation sequencing (NGS), we examined the genotypes and phenotypes of VGSC-related epilepsy cases from a cohort of 691 epilepsy subjects. Our findings revealed that 5.1% of subjects harbored VGSC variants, specifically 22 with SCN1A, 9 with SCN2A, 1 with SCN8A, and 3 with SCN1B variants; no SCN3A variants were detected. Among these, 14 variants were previously reported, while 21 were newly identified. SCN1A variant carriers predominantly presented with Dravet Syndrome (DS) and Genetic Epilepsy with Febrile Seizures Plus (GEFS + ), featuring a heightened sensitivity to fever-induced seizures. Statistically significant disparities emerged between the SCN1A-DS and SCN1A-GEFS+ groups concerning seizure onset and genetic diagnosis age, incidence of status epilepticus, mental retardation, anti-seizure medication (ASM) responsiveness, and familial history. Notably, subjects with SCN1A variants affecting the protein's pore region experienced more frequent cluster seizures. All SCN2A variants were of de novo origin, and 88.9% of individuals with SCN2A variations exhibited cluster seizures. This research reveals a significant association between variations in VGSC-related genes and the clinical phenotype diversity of epilepsy subjects in China, emphasizing the pivotal role of NGS screening in establishing accurate disease diagnoses and guiding the selection of ASM.

Fine mapping and candidate gene analysis of Dravet syndrome modifier loci on mouse chromosomes 7 and 8.

Dravet syndrome is a developmental and epileptic encephalopathy (DEE) characterized by intractable seizures, comorbidities related to developmental, cognitive, and motor delays, and a high mortality burden due to sudden unexpected death in epilepsy (SUDEP). Most Dravet syndrome cases are attributed to SCN1A haploinsufficiency, with genetic modifiers and environmental factors influencing disease severity. Mouse models with heterozygous deletion of Scn1a recapitulate key features of Dravet syndrome, including seizures and premature mortality; however, severity varies depending on genetic background. Here, we refined two Dravet survival modifier (Dsm) loci, Dsm2 on chromosome 7 and Dsm3 on chromosome 8, using interval-specific congenic (ISC) mapping. Dsm2 was complex and encompassed at least two separate loci, while Dsm3 was refined to a single locus. Candidate modifier genes within these refined loci were prioritized based on brain expression, strain-dependent differences, and biological relevance to seizures or epilepsy. High priority candidate genes for Dsm2 include Nav2, Ptpn5, Ldha, Dbx1, Prmt3 and Slc6a5, while Dsm3 has a single high priority candidate, Psd3. This study underscores the complex genetic architecture underlying Dravet syndrome and provides insights into potential modifier genes that could influence disease severity and serve as novel therapeutic targets.

Preictal dysfunctions of inhibitory interneurons paradoxically lead to their rebound hyperactivity and to low-voltage-fast onset seizures in Dravet syndrome.

Epilepsies have numerous specific mechanisms. The understanding of neural dynamics leading to seizures is important for disclosing pathological mechanisms and developing therapeutic approaches. We investigated electrographic activities and neural dynamics leading to convulsive seizures in patients and mouse models of Dravet syndrome (DS), a developmental and epileptic encephalopathy in which hypoexcitability of GABAergic neurons is considered to be the main dysfunction. We analyzed EEGs from DS patients carrying a SCN1A pathogenic variant, as well as epidural electrocorticograms, hippocampal local field potentials, and hippocampal single-unit neuronal activities in Scn1a+/- and Scn1aRH/+ DS mice. Strikingly, most seizures had low-voltage-fast onset in both patients and mice, which is thought to be generated by hyperactivity of GABAergic interneurons, the opposite of the main pathological mechanism of DS. Analyzing single-unit recordings, we observed that temporal disorganization of the firing of putative interneurons in the period immediately before the seizure (preictal) precedes the increase of their activity at seizure onset, together with the entire neuronal network. Moreover, we found early signatures of the preictal period in the spectral features of hippocampal and cortical field potential of Scn1a mice and of patients' EEG, which are consistent with the dysfunctions that we observed in single neurons and that allowed seizure prediction. Therefore, the perturbed preictal activity of interneurons leads to their hyperactivity at the onset of generalized seizures, which have low-voltage-fast features that are similar to those observed in other epilepsies and are triggered by hyperactivity of GABAergic neurons. Preictal spectral features may be used as predictive seizure biomarkers.

Modulating Endoplasmic Reticulum Chaperones and Mutant Protein Degradation in GABRG2(Q390X) Associated with Genetic Epilepsy with Febrile Seizures Plus and Dravet Syndrome.

A significant number of patients with genetic epilepsy do not obtain seizure freedom, despite developments in new antiseizure drugs, suggesting a need for novel therapeutic approaches. Many genetic epilepsies are associated with misfolded mutant proteins, including GABRG2(Q390X)-associated Dravet syndrome, which we have previously shown to result in intracellular accumulation of mutant GABAA receptor γ2(Q390X) subunit protein. Thus, a potentially promising therapeutic approach is modulation of proteostasis, such as increasing endoplasmic reticulum (ER)-associated degradation (ERAD). To that end, we have here identified an ERAD-associated E3 ubiquitin ligase, HRD1, among other ubiquitin ligases, as a strong modulator of wildtype and mutant γ2 subunit expression. Overexpressing HRD1 or knockdown of HRD1 dose-dependently reduced the γ2(Q390X) subunit. Additionally, we show that zonisamide (ZNS)-an antiseizure drug reported to upregulate HRD1-reduces seizures in the Gabrg2+/Q390X mouse. We propose that a possible mechanism for this effect is a partial rescue of surface trafficking of GABAA receptors, which are otherwise sequestered in the ER due to the dominant-negative effect of the γ2(Q390X) subunit. Furthermore, this partial rescue was not due to changes in ER chaperones BiP and calnexin, as total expression of these chaperones was unchanged in γ2(Q390X) models. Our results here suggest that leveraging the endogenous ERAD pathway may present a potential method to degrade neurotoxic mutant proteins like the γ2(Q390X) subunit. We also demonstrate a pharmacological means of regulating proteostasis, as ZNS alters protein trafficking, providing further support for the use of proteostasis regulators for the treatment of genetic epilepsies.

Opposing effects of the purinergic P2X7 receptor on seizures in neurons and microglia in male mice.

BACKGROUND: The purinergic ATP-gated P2X7 receptor (P2X7R) is increasingly recognized to contribute to pathological neuroinflammation and brain hyperexcitability. P2X7R expression has been shown to be increased in the brain, including both microglia and neurons, in experimental models of epilepsy and patients. To date, the cell type-specific downstream effects of P2X7Rs during seizures remain, however, incompletely understood. METHODS: Effects of P2X7R signaling on seizures and epilepsy were analyzed in induced seizure models using male mice including the kainic acid model of status epilepticus and pentylenetetrazole model and in male and female mice in a genetic model of Dravet syndrome. RNA sequencing was used to analyze P2X7R downstream signaling during seizures. To investigate the cell type-specific role of the P2X7R during seizures and epilepsy, we generated mice lacking exon 2 of the P2rx7 gene in either microglia (P2rx7:Cx3cr1-Cre) or neurons (P2rx7:Thy-1-Cre). To investigate the protective potential of overexpressing P2X7R in GABAergic interneurons, P2X7Rs were overexpressed using adeno-associated virus transduction under the mDlx promoter. RESULTS: RNA sequencing of hippocampal tissue from wild-type and P2X7R knock-out mice identified both glial and neuronal genes, in particular genes involved in GABAergic signaling, under the control of the P2X7R following seizures. Mice with deleted P2rx7 in microglia displayed less severe acute seizures and developed a milder form of epilepsy, and microglia displayed an anti-inflammatory molecular profile. In contrast, mice lacking P2rx7 in neurons showed a more severe seizure phenotype when compared to epileptic wild-type mice. Analysis of single-cell expression data revealed that human P2RX7 expression is elevated in the hippocampus of patients with temporal lobe epilepsy in excitatory and inhibitory neurons. Functional studies determined that GABAergic interneurons display increased responses to P2X7R activation in experimental epilepsy. Finally, we show that viral transduction of P2X7R in GABAergic interneurons protects against evoked and spontaneous seizures in experimental temporal lobe epilepsy and in mice lacking Scn1a, a model of Dravet syndrome. CONCLUSIONS: Our results suggest a dual and opposing action of P2X7R in epilepsy and suggest P2X7R overexpression in GABAergic interneurons as a novel therapeutic strategy for acquired and, possibly, genetic forms of epilepsy.

Microglia Mitigate Neuronal Activation in a Zebrafish Model of Dravet Syndrome.

It has been known for a long time that epileptic seizures provoke brain neuroinflammation involving the activation of microglial cells. However, the role of these cells in this disease context and the consequences of their inflammatory activation on subsequent neuron network activity remain poorly understood so far. To fill this gap of knowledge and gain a better understanding of the role of microglia in the pathophysiology of epilepsy, we used an established zebrafish Dravet syndrome epilepsy model based on Scn1Lab sodium channel loss-of-function, combined with live microglia and neuronal Ca2+ imaging, local field potential (LFP) recording, and genetic microglia ablation. Data showed that microglial cells in scn1Lab-deficient larvae experiencing epileptiform seizures displayed morphological and biochemical changes characteristic of M1-like pro-inflammatory activation; i.e., reduced branching, amoeboid-like morphology, and marked increase in the number of microglia expressing pro-inflammatory cytokine Il1ß. More importantly, LFP recording, Ca2+ imaging, and swimming behavior analysis showed that microglia-depleted scn1Lab-KD larvae displayed an increase in epileptiform seizure-like neuron activation when compared to that seen in scn1Lab-KD individuals with microglia. These findings strongly suggest that despite microglia activation and the synthesis of pro-inflammatory cytokines, these cells provide neuroprotective activities to epileptic neuronal networks, making these cells a promising therapeutic target in epilepsy.

[Correlation between clinical phenotypes and genotypes among 46 children with SCN1A-related developmental epileptic encephalopathy].

OBJECTIVE: To explore the correlation between clinical phenotypes and genotypes among 46 children with SCN1A-related developmental epileptic encephalopathy (DEE). METHODS: Clinical data of 46 children with DEE and SCN1A variants identified at the Guangzhou Women and Children's Medical Center between January 2018 and June 2022 were collected. The children were grouped based on their age of onset, clinical manifestations, neurodevelopmental status, and results of genetic testing. The correlation between SCN1A genotypes and clinical phenotypes was analyzed. RESULTS: Among the 46 patients, 2 children (4.35%) had developed the symptoms before 3 months of age, 42 (91.30%) were between 3 to 9 months, and 2 cases (4.35%) were after 10 months. Two cases (4.35%) presented with epilepsy of infancy with migrating focal seizures (EIMFS), while 44 (95.7%) had presented with Dravet syndrome (DS), including 28 cases (63.6%) with focal onset (DS-F), 13 cases (29.5%) with myoclonic type (DS-M), 1 case (2.27%) with generalized type (DS-G), and 2 cases (4.55%) with status epilepticus type (DS-SE). Both of the two EIMFS children had severe developmental delay, and among the DS patients, 7 cases had normal development, while the remaining had developmental delay. A total of 44 variants were identified through genetic sequencing, which included 16 missense variants and 28 truncating variants. All EIMFS children had carried the c.677C>T (p.Thr226Met) missense variant. In the DS group, there was a significant difference in the age of onset between the missense variants group and the truncating variants group (P < 0.05). Missense variants were more common in D1 (7/15, 46.7%) and pore regions (8/15, 53.3%), while truncating variants were more common in D1 (12/28, 42.9%). Children with variants outside the pore region were more likely to develop myoclonic seizures. CONCLUSION: The clinical phenotypes of DEE are diverse. There is a difference in the age of onset between individuals with truncating and missense variants in the SCN1A gene. Missense variants outside the pore region are associated with a higher incidence of myoclonic seizures.

[Progressive myoclonic epilepsy: a retrospective study of newly-diagnosed adult patients from a single center].

OBJECTIVE: To retrospectively analyze the clinical phenotype and pathogenic variants in patients with Progressive myoclonus epilepsy (PME). METHODS: Clinical data and results of genetic testing for 11 patients diagnosed with PME at the Department of Neurology, the First Affiliated Hospital of Zhejiang University School of Medicine from June 2017 to December 2022 were collected and analyzed. RESULTS: All of the patients, including 4 males and 7 females, had predominant action myoclonus. Three patients had myoclonus as the initial manifestation, whilst eight were diagnosed through genetic testing, including three cases with NEU1 gene variants, two with EPM2A gene variants (1 was novel), one with MT-TK gene variant, one with ATN1 gene variant, and one with CSTB gene variant. No pathogenic variant was identified in the remaining three cases. Among the eight patients with a genetic diagnosis, three were diagnosed with sialidosis, two with Lafora disease, one with Dentatorubral-pallidoluysian atrophy (DRPLA), one with Unverricht-Lundborg disease (ULD), and one with Myoclonic epilepsy with ragging red fibers (MERRF). CONCLUSION: Compared with pediatric patients, adult patients with PME represent a distinct subtype with slower progression and milder cognitive impairment.

Genetic exploration of Dravet syndrome: two case report.

BACKGROUND: Dravet syndrome is an infantile-onset developmental and epileptic encephalopathy (DEE) characterized by drug resistance, intractable seizures, and developmental comorbidities. This article focuses on manifestations in two Indonesian children with Javanese ethnicity who experienced Dravet syndrome with an SCN1A gene mutation, presenting genetic analysis findings using next-generation sequencing. CASE PRESENTATION: We present a case series involving two Indonesian children with Javanese ethnicity whom had their first febrile seizure at the age of 3 months, triggered after immunization. Both patients had global developmental delay and intractable seizures. We observed distinct genetic findings in both our cases. The first patient revealed heterozygous deletion mutation in three genes (TTC21B, SCN1A, and SCN9A). In our second patient, previously unreported mutation was discovered at canonical splice site upstream of exon 24 of the SCN1A gene. Our patient's outcomes improved after therapeutic evaluation based on mutation findings When comparing clinical manifestations in our first and second patients, we found that the more severe the genetic mutation discovered, the more severe the patient's clinical manifestations. CONCLUSION: These findings emphasize the importance of comprehensive genetic testing beyond SCN1A, providing valuable insights for personalized management and tailored therapeutic interventions in patients with Dravet syndrome. Our study underscores the potential of next-generation sequencing in advancing genotype-phenotype correlations and enhancing diagnostic precision for effective disease management.

Integrating Proteomics and Transcriptomics Reveals the Potential Pathways of Hippocampal Neuron Apoptosis in Dravet Syndrome Model Mice.

An important component contributing to the onset of epilepsy is the death of hippocampal neurons. Several studies have shown that Dravet syndrome model mice: Scn1a KO mice have a high number of apoptotic neurons following seizures, but the precise mechanism underlying this remains unclear. The aim of this research was to elucidate the potential molecular mechanism of neuronal apoptosis in Scn1a KO mice by integrating proteomics and transcriptomics, with the ultimate goal of offering better neuroprotection. We found that apoptotic processes were enriched in both proteomic and transcriptomic GO analyses, and KEGG results also indicated that differential proteins and genes play a role in neurotransmission, the cell cycle, apoptosis, and neuroinflammation. Then, we examined the upstream and downstream KGML interactions of the pathways to determine the relationship between the two omics, and we found that the HIF-1 signaling pathway plays a significant role in the onset and apoptosis of epilepsy. Meanwhile, the expression of the apoptosis-related protein VHL decreased in this pathway, and the expression of p21 was upregulated. Therefore, this study suggests that VHL/HIF-1α/p21 might be involved in the apoptosis of hippocampal neurons in Scn1a KO mice.

A high seizure burden increases several prostaglandin species in the hippocampus of a Scn1a+/- mouse model of Dravet syndrome.

Dravet syndrome is an intractable epilepsy with a high seizure burden that is resistant to current anti-seizure medications. There is evidence that neuroinflammation plays a role in epilepsy and seizures, however few studies have specifically examined neuroinflammation in Dravet syndrome under conditions of a higher seizure burden. Here we used an established genetic mouse model of Dravet syndrome (Scn1a+/- mice), to examine whether a higher seizure burden impacts the number and morphology of microglia in the hippocampus. Moreover, we examined whether a high seizure burden influences classical inflammatory mediators in this brain region. Scn1a+/- mice with a high seizure burden induced by thermal priming displayed a localised reduction in microglial cell density in the granule cell layer and subgranular zone of the dentate gyrus, regions important to postnatal neurogenesis. However, microglial cell number and morphology remained unchanged in other hippocampal subfields. The high seizure burden in Scn1a+/- mice did not affect hippocampal mRNA expression of classical inflammatory mediators such as interleukin 1ß and tumour necrosis factor α, but increased cyclooxygenase 2 (COX-2) expression. We then quantified hippocampal levels of prostanoids that arise from COX-2 mediated metabolism of fatty acids and found that Scn1a+/- mice with a high seizure burden displayed increased hippocampal concentrations of numerous prostaglandins, notably PGF2α, PGE2, PGD2, and 6-K-PGF1A, compared to Scn1a+/- mice with a low seizure burden. In conclusion, a high seizure burden increased hippocampal concentrations of various prostaglandin mediators in a mouse model of Dravet syndrome. Future studies could interrogate the prostaglandin pathways to further better understand their role in the pathophysiology of Dravet syndrome.

Predictors of genetic diagnosis in individuals with developmental and epileptic encephalopathies.

OBJECTIVE: To evaluate the clinical predictors of positive genetic investigation in developmental and epileptic encephalopathies, beyond the influence of Dravet Syndrome. METHODS: The study included 98 patients diagnosed with developmental and epileptic encephalopathies. The patients underwent Sanger sequencing of SCN1A, Chromosomal Microarray Analysis, and Whole Exome Sequencing. The association of clinical variables with a positive genetic test was investigated using univariate and multivariate analysis. RESULTS: Genetic diagnosis was identified in 47 (48 %) patients with developmental and epileptic encephalopathies. Beyond Dravet Syndrome influence, first seizure in the context of fever (p < 0.01), seizures precipitated by temperature (p = 0.04), cognitive regression (p = 0.04), hypotonia (p < 0.01), and focal seizures (p = 0.03) increased the chances of a positive genetic investigation. In contrast, atonic seizures (p = 0.01) and generalized discharges on electroencephalogram (p = 0.02) decreased the chances. Dravet Syndrome was positively associated with a genetic developmental and epileptic encephalopathies etiology (p < 0.01), whereas epilepsy with myoclonic-atonic seizures (p = 0.01), developmental and epileptic encephalopathies with spike-wave activation in sleep (p = 0.04), and Lennox-Gastaut syndrome (p = 0.03) were negatively associated. In multivariate analysis, the first seizure in the context of fever (p < 0.01) and hypotonia (p = 0.02) were positively, and atonic seizures (p = 0.01) were negatively and independently associated with a genetic etiology. CONCLUSION: The predictive variables of genetic investigation in developmental and epileptic encephalopathies are first seizure in the context of fever and hypotonia, whereas atonic seizures decrease the chances of finding a genetic cause for developmental and epileptic encephalopathies. Regarding epileptic syndromes, Dravet Syndrome is highly associated with a positive genetic test, whereas epilepsy with myoclonic-atonic seizures, developmental and epileptic encephalopathies with spike-wave activation in sleep, and Lennox-Gastaut syndrome are rarely associated with a positive genetic investigation.