Pamela Lachance-Touchette

Genetically encoded impairment of neuronal KCC2 cotransporter function in human idiopathic generalized epilepsy

The KCC2 cotransporter establishes the low neuronal Cl− levels required for GABAA and glycine (Gly) receptor‐mediated inhibition, and KCC2 deficiency in model organisms results in network hyperexcitability. However, no mutations in KCC2 have been documented in human disease. Here, we report two non‐synonymous functional variants in human KCC2, R952H and R1049C, exhibiting clear statistical association with idiopathic generalized epilepsy (IGE). These variants reside in conserved residues in the KCC2 cytoplasmic C‐terminus, exhibit significantly impaired Cl−‐extrusion capacities resulting in less hyperpolarized Gly equilibrium potentials (EGly), and impair KCC2 stimulatory phosphorylation at serine 940, a key regulatory site. These data describe a novel KCC2 variant significantly associated with a human disease and suggest genetically encoded impairment of KCC2 functional regulation may be a risk factor for the development of human IGE.

Screening of GABRB3 in French-Canadian families with idiopathic generalized epilepsy

Mutations in the GABRB3 have been recently associated with childhood absence epilepsy (CAE) in families from Honduras and Mexico. In this study, we aimed to determine the frequency of mutation in this gene in our cohort of families with CAE and other related idiopathic generalized epilepsy (IGE) syndromes. We screened the open reading frame of GABRB3 in 183 French‐Canadian individuals with IGE, including 88 with CAE. A total of nine single nucleotide polymorphisms (SNPs) have been identified, five of which are novel. The previously described P11S missense mutation was found in three affected and one unaffected individuals from a French‐Canadian family. However, the P11S variant was also found in one of our 190 control individuals of French‐Canadian origin, suggesting that this variant is rather a rare polymorphism in this population. Further screening of other IGE cohorts from various ethnic origins would help to confirm the association between this rare functional variant and epilepsy.

Single-cell genetic expression of mutant GABAA receptors causing Human genetic epilepsy alters dendritic spine and GABAergic bouton formation in a mutation-specific manner

Mutations in genes encoding for GABAA receptor subunits is a well-established cause of genetic generalized epilepsy. GABA neurotransmission is implicated in several developmental processes including neurite outgrowth and synapse formation. Alteration in excitatory/inhibitory synaptic activities plays a critical role in epilepsy, thus here we investigated whether mutations in α1 subunit of GABAA receptor may affect dendritic spine and GABAergic bouton formation. In particular, we examined the effects of three mutations of the GABRA1 gene (D219N, A322D and K353delins18X) that were found in a cohort of French Canadian families with genetic generalized epilepsy. We used a novel single-cell genetic approach, by preparing cortical organotypic cultures from GABRA1flox/flox mice and simultaneously inactivating endogenous GABRA1 and transfecting mutant α1 subunits in single glutamatergic pyramidal cells and basket GABAergic interneurons by biolistic transfection. We found that GABRA1−/− GABAergic cells showed reduced innervation field, which was rescued by co-expressing α1-A322D and α1-WT but not α1-D219N. We further found that the expression of the most severe GABRA1 missense mutation (α1-A322D) induced a striking increase of spine density in pyramidal cells along with an increase in the number of mushroom-like spines. In addition, α1-A322D expression in GABAergic cells slightly increased perisomatic bouton density, whereas other mutations did not alter bouton formation. All together, these results suggest that the effects of different GABAAR mutations on GABAergic bouton and dendritic spine formation are specific to the mutation and cannot be always explained by a simple loss-of-function gene model. The use of single cell genetic manipulation in organotypic cultures may provide a better understanding of the specific and distinct neural circuit alterations caused by different GABAA receptor subunit mutations and will help define the pathophysiology of genetic generalized epilepsy syndromes.

Mutation Burden of Rare Variants in Schizophrenia Candidate Genes

Background Schizophrenia (SCZ) is a very heterogeneous disease that affects approximately 1% of the general population. Recently, the genetic complexity thought to underlie this condition was further supported by three independent studies that identified an increased number of damaging de novo mutations DNM in different SCZ probands. While these three reports support the implication of DNM in the pathogenesis of SCZ, the absence of overlap in the genes identified suggests that the number of genes involved in SCZ is likely to be very large; a notion that has been supported by the moderate success of Genome-Wide Association Studies (GWAS). Methods To further examine the genetic heterogeneity of this disease, we resequenced 62 genes that were found to have a DNM in SCZ patients, and 40 genes that encode for proteins known to interact with the products of the genes with DNM, in a cohort of 235 SCZ cases and 233 controls. Results We found an enrichment of private nonsense mutations amongst schizophrenia patients. Using a kernel association method, we were able to assess for association for different sets. Although our power of detection was limited, we observed an increased mutation burden in the genes that have DNM.