Characterization of the GABRB2-Associated Neurodevelopmental Disorders.

OBJECTIVE: We aimed to characterize the phenotypic spectrum and functional consequences associated with variants in the gene GABRB2, coding for the γ-aminobutyric acid type A (GABAA ) receptor subunit ß2. METHODS: We recruited and systematically evaluated 25 individuals with variants in GABRB2, 17 of whom are newly described and 8 previously reported with additional clinical data. Functional analysis was performed using a Xenopus laevis oocyte model system. RESULTS: Our cohort of 25 individuals from 22 families with variants in GABRB2 demonstrated a range of epilepsy phenotypes from genetic generalized epilepsy to developmental and epileptic encephalopathy. Fifty-eight percent of individuals had pharmacoresistant epilepsy; response to medications targeting the GABAergic pathway was inconsistent. Developmental disability (present in 84%) ranged from mild intellectual disability to severe global disability; movement disorders (present in 44%) included choreoathetosis, dystonia, and ataxia. Disease-associated variants cluster in the extracellular N-terminus and transmembrane domains 1-3, with more severe phenotypes seen in association with variants in transmembrane domains 1 and 2 and the allosteric binding site between transmembrane domains 2 and 3. Functional analysis of 4 variants in transmembrane domains 1 or 2 (p.Ile246Thr, p.Pro252Leu, p.Ile288Ser, p.Val282Ala) revealed strongly reduced amplitudes of GABA-evoked anionic currents. INTERPRETATION: GABRB2-related epilepsy ranges broadly in severity from genetic generalized epilepsy to developmental and epileptic encephalopathies. Developmental disability and movement disorder are key features. The phenotypic spectrum is comparable to other GABAA receptor-encoding genes. Phenotypic severity varies by protein domain. Experimental evidence supports loss of GABAergic inhibition as the mechanism underlying GABRB2-associated neurodevelopmental disorders. ANN NEUROL 2021;89:573-586.

In vitro neuronal network activity as a new functional diagnostic system to detect effects of Cerebrospinal fluid from autoimmune encephalitis patients.

The intent of this study was to investigate if cerebrospinal fluid (CSF) from autoimmune encephalitis (AE) patients regulates in vitro neuronal network activity differentially to healthy human control CSF (hCSF). To this end, electrophysiological effects of CSF from AE patients or hCSF were measured by in vitro neuronal network activity (ivNNA) recorded with microelectrode arrays (MEA). CSF from patients with either N-methyl-D-aspartate-receptor-antibody (pCSFNMDAR, n = 7) or Leucine-rich-glioma-inactivated-1-Ab (pCSFLGI1, n = 6) associated AE suppressed global spiking activity of neuronal networks by a factor of 2.17 (p < 0.05) or 2.42 (p < 0.05) compared to hCSF. The former also suppressed synchronous network bursting by a factor of 1.93 (p < 0.05) in comparison to hCSF (n = 13). As a functional diagnostic test, this parameter reached a sensitivity of 86% for NMDAR-Ab- and 100% for LGI1-Ab-associated AE vs. hCSF at a specificity of 85%. To explore if modulation at the NMDAR influences effects of hCSF or pathological CSF, we applied the NMDAR-antagonist 2-Amino-5-phosphono-pentanoic acid (AP5). In CSF from NMDAR-Ab-associated AE patients, spike rate reduction by AP5 was more than 2-fold larger than in hCSF (p < 0.05), and network burst rate reduction more than 18-fold (p < 0.01). Recording ivNNA might help discriminating between functional effects of CSF from AE patients and hCSF, and thus could be used as a functional diagnostic test in AE. The pronounced suppression of ivNNA by CSF from NMDAR-Ab-associated AE patients and simultaneous antagonism at the NMDAR by AP5, particularly in burst activity, compared to hCSF plus AP5, confirms that the former contains additional ivNNA-suppressing factors.

Rare coding variants in genes encoding GABAA receptors in genetic generalised epilepsies: an exome-based case-control study.

BACKGROUND: Genetic generalised epilepsy is the most common type of inherited epilepsy. Despite a high concordance rate of 80% in monozygotic twins, the genetic background is still poorly understood. We aimed to investigate the burden of rare genetic variants in genetic generalised epilepsy. METHODS: For this exome-based case-control study, we used three different genetic generalised epilepsy case cohorts and three independent control cohorts, all of European descent. Cases included in the study were clinically evaluated for genetic generalised epilepsy. Whole-exome sequencing was done for the discovery case cohort, a validation case cohort, and two independent control cohorts. The replication case cohort underwent targeted next-generation sequencing of the 19 known genes encoding subunits of GABAA receptors and was compared to the respective GABAA receptor variants of a third independent control cohort. Functional investigations were done with automated two-microelectrode voltage clamping in Xenopus laevis oocytes. FINDINGS: Statistical comparison of 152 familial index cases with genetic generalised epilepsy in the discovery cohort to 549 ethnically matched controls suggested an enrichment of rare missense (Nonsyn) variants in the ensemble of 19 genes encoding GABAA receptors in cases (odds ratio [OR] 2·40 [95% CI 1·41-4·10]; pNonsyn=0·0014, adjusted pNonsyn=0·019). Enrichment for these genes was validated in a whole-exome sequencing cohort of 357 sporadic and familial genetic generalised epilepsy cases and 1485 independent controls (OR 1·46 [95% CI 1·05-2·03]; pNonsyn=0·0081, adjusted pNonsyn=0·016). Comparison of genes encoding GABAA receptors in the independent replication cohort of 583 familial and sporadic genetic generalised epilepsy index cases, based on candidate-gene panel sequencing, with a third independent control cohort of 635 controls confirmed the overall enrichment of rare missense variants for 15 GABAA receptor genes in cases compared with controls (OR 1·46 [95% CI 1·02-2·08]; pNonsyn=0·013, adjusted pNonsyn=0·027). Functional studies for two selected genes (GABRB2 and GABRA5) showed significant loss-of-function effects with reduced current amplitudes in four of seven tested variants compared with wild-type receptors. INTERPRETATION: Functionally relevant variants in genes encoding GABAA receptor subunits constitute a significant risk factor for genetic generalised epilepsy. Examination of the role of specific gene groups and pathways can disentangle the complex genetic architecture of genetic generalised epilepsy. FUNDING: EuroEPINOMICS (European Science Foundation through national funding organisations), Epicure and EpiPGX (Sixth Framework Programme and Seventh Framework Programme of the European Commission), Research Unit FOR2715 (German Research Foundation and Luxembourg National Research Fund).