Effects of GABAA Receptor α3 Subunit Epilepsy Mutations on Inhibitory Synaptic Signaling

Abstract

Missense mutations T166M, Q242L, T336M, and Y474C in the GABAA receptor (GABAAR) α3 subunit gene are associated with epileptic seizures, dysmorphic features, intellectual disability, and developmental delay. When incorporated into GABAARs expressed in oocytes, all mutations are known to reduce GABA-evoked whole-cell currents. However, their impact on the properties of inhibitory synaptic currents (IPSCs) is unknown, largely because it is difficult to establish, much less control, the stoichiometry of GABAAR expressed in native neuronal synapses. To circumvent this problem, we employed a HEK293 cell-neuron co-culture expression system that permits the recording of IPSCs mediated by a pure population of GABAARs with a defined stoichiometry. We first demonstrated that IPSCs mediated by α3-containing GABAARs (α3β3γ2) decay significantly slower than those mediated by α1-containing isoforms (α1β2γ2 or α1β3γ2). GABAAR α3 mutations did not affect IPSC peak amplitudes or 10–90% rise times, but three of the mutations affected IPSC decay. T336M significantly accelerated the IPSC decay rate whereas T166M and Y474C had the opposite effect. The acceleration of IPSC decay kinetics caused by the T366M mutation was returned to wild-type-like values by the anti-epileptic medication, midazolam. Quantification experiments in HEK293 cells revealed a significant reduction in cell-surface expression for all mutants, in agreement with previous oocyte data. Taken together, our results show that impaired surface expression and altered IPSC decay rates could both be significant factors underlying the pathologies associated with these mutations.

Structural and functional properties of spinal dorsal horn neurons after peripheral nerve injury change overtime via astrocyte activation

“…On the other hand, IPSC amplitude was dramatically suppressed, and the suppression was greater in the late post-PNI period. The decay time of IPSC has been shown to be highly dependent on GABA receptor isoforms, 37 and chronic pain is accompanied by altered expression of some GABA receptor subunits, 38 and our results support these reports. One of the main reasons why our results differ from previous electrophysiological reports of inhibitory synaptic transmission after PNI is the difference between in vivo and in vitro.…”

GABRG2 Variants Associated with Febrile Seizures

“…Furthermore, key phenotypic differences may even be seen between individuals with the same genetic alteration [ 71 ]. In a recent study on GABRB3 variants, it was reported that there was a significant genotype–phenotype correlation in individuals affected by GABRB3 -related disorders, as they were clustered in three regions of the β3 subunit that were key to its function (GABA binding region, coupling region, and TMD) [ 72 ], clearly linking GABR variant pathogenicity and receptor structure–function [ 29 , 73 , 74 , 75 , 76 , 77 ].…”

Joint, multifaceted genomic analysis enables diagnosis of diverse, ultra-rare monogenic presentations

“…Two of these genes, CACNA1C and GABRA3, harbored high impact de novo and hemizygous missense variants respectively, corresponding to known patient diagnoses. 55,56 The genome of another, undiagnosed, now deceased patient from this cluster with no prior candidate variants contained a synonymous de novo variant predicted to alter splicing in another gene in the same functional pathway, HCN4 (Figure 4d). All three patients exhibited seizures at a young age, speech delays, severe hypotonia, spasticity and visual impairment.…”