Abstract
The GABAA receptor (GABAAR) α1 subunit A295D epilepsy mutation reduces the surface expression of α1A295Dβ2γ2 GABAARs via ER-associated protein degradation. Suberanilohydroxamic acid (SAHA, also known as Vorinostat) was recently shown to correct the misfolding of α1A295D subunits and thereby enhance the functional surface expression of α1A295Dβ2γ2 GABAARs. Here we investigated whether SAHA can also restore the surface expression of γ2 GABAAR subunits that incorporate epilepsy mutations (N40S, R43Q, P44S, R138G) known to reduce surface expression via ER-associated protein degradation. As a control, we also investigated the γ2K289M epilepsy mutation that impairs gating without reducing surface expression. Effects of mutations were evaluated on inhibitory postsynaptic currents (IPSCs) mediated by the major synaptic α1β2γ2 GABAAR isoform. Recordings were performed in neuron-HEK293 cell artificial synapses to minimise contamination by GABAARs of undefined subunit composition. Transfection with α1β2γ2N40S, α1β2γ2R43Q, α1β2γ2P44S and α1β2γ2R138G subunits produced IPSCs with decay times slower than those of unmutated α1β2γ2 GABAARs due to the low expression of mutant γ2 subunits and the correspondingly high expression of slow-decaying α1β2 GABAARs. SAHA pre-treatment significantly accelerated the decay time constants of IPSCs consistent with the upregulation of mutant γ2 subunit expression. This increase in surface expression was confirmed by immunohistochemistry. SAHA had no effect on either the IPSC kinetics or surface expression levels of α1β2γ2K289M GABAARs, confirming its specificity for ER-retained mutant γ2 subunits. We also found that α1β2γ2K289M GABAARs and SAHA-treated α1β2γ2R43Q, α1β2γ2P44S and α1β2γ2R138G GABAARs all mediated IPSCs that decayed at significantly faster rates than wild type receptors as temperature was increased from 22 to 40°C. This may help explain why these mutations cause febrile seizures (FS). Given that SAHA is approved by therapeutic regulatory agencies for human use, we propose that it may be worth investigating as a treatment for epilepsies caused by the N40S, R43Q, P44S and R138G mutations. Although SAHA has already been proposed as a therapeutic for patients harbouring the α1A295D epilepsy mutation, the present study extends its potential utility to a new subunit and four new mutations.
Highlights
The epilepsies are a related group of neurological disorders characterised by seizures resulting from abnormal, hypersynchronised electrical activity in neurons
In the present study we extended this line of investigation by asking whether SAHA can recover the surface expression of epilepsy mutations to γ2 subunits that result in endoplasmic reticulum (ER)-retention
Properties of inhibitory postsynaptic currents (IPSCs) Generated by GABAA receptor (GABAAR) Incorporating Epilepsy-Causing γ2 Mutant Subunits
Summary
The epilepsies are a related group of neurological disorders characterised by seizures resulting from abnormal, hypersynchronised electrical activity in neurons. Drugs that potentiate GABAARs, such as benzodiazepines, neurosteroids and barbiturates, are often effective in treating epilepsies whereas drugs that inhibit GABAARs, such as bicuculline and picrotoxin, can give rise to seizures (Rogawski and Löscher, 2004; Riss et al, 2008). Together, this is consistent with the widely held view that GABAergic inhibition restrains the tendency of recurrently connected excitatory neural networks to transition, via positive feedback, into synchronous epileptiform activity (Rogawski and Löscher, 2004)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have