Inhibitory synapse deficits caused by familial α1 GABAA receptor mutations in epilepsy

- Epilepsy-causing mutations in GABA receptors impair synaptic inhibition.
- Not all mutant receptors are responsive to standard anticonvulsant drugs.
- Some mutant receptors showed increased temperature sensitivity at synapses.
- Suberanilohydroxamic acid restores misfolded receptors.
- GABA receptor mutations give rise to changes in neuronal morphology.

Epilepsy is a spectrum of neurological disorders with many causal factors. The GABA type-A receptor (GABAAR) is a major genetic target for heritable human epilepsies. Here we examine the functional effects of three epilepsy-causing mutations to the α1 subunit (α1T10'I, α1D192N and α1A295D) on inhibitory postsynaptic currents (IPSCs) mediated by the major synaptic GABAAR isoform, α1β2γ2L. We employed a neuron - HEK293 cell heterosynapse preparation to record IPSCs mediated by mutant-containing GABAARs in isolation from other GABAAR isoforms. IPSCs were recorded in the presence of the anticonvulsant drugs, carbamazepine and midazolam, and at elevated temperatures (22, 37 and 40 °C) to gain insight into mechanisms of febrile seizures. The mutant subunits were also transfected into cultured cortical neurons to investigate changes in synapse formation and neuronal morphology using fluorescence microscopy. We found that IPSCs mediated by α1T10'Iβ2γ2L, α1D192Nβ2γ2L GABAARs decayed faster than those mediated by α1β2γ2L receptors. IPSCs mediated by α1D192Nβ2γ2L and α1A295Dβ2γ2L receptors also exhibited a heightened temperature sensitivity. In addition, the α1T10'Iβ2γ2L GABAARs were refractory to modulation by carbamazepine or midazolam. In agreement with previous studies, we found that α1A295Dβ2γ2L GABAARs were retained intracellularly in HEK293 cells and neurons. However, pre-incubation with 100 nM suberanilohydroxamic acid (SAHA) induced α1A295Dβ2γ2L GABAARs to mediate IPSCs that were indistinguishable in magnitude and waveform from those mediated by α1β2γ2L receptors. Finally, mutation-specific changes to synaptic bouton size, synapse number and neurite branching were also observed. These results provide new insights into the mechanisms of epileptogenesis of α1 epilepsy mutations and suggest possible leads for improving treatments for patients harbouring these mutations.