Multi-electrode array study of neuronal cultures expressing nicotinic β2-V287L subunits, linked to autosomal dominant nocturnal frontal lobe epilepsy. An in vitro model of spontaneous epilepsy.

Francesca Gullo,Irene Manfredi,Marzia Lecchi,Giorgio Casari,Enzo Wanke,Andrea Becchetti

doi:10.3389/fncir.2014.00087

Abstract

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is a partial sleep-related epilepsy which can be caused by mutant neuronal nicotinic acetylcholine receptors (nAChR). We applied multi-electrode array (MEA) recording methods to study the spontaneous firing activity of neocortical cultures obtained from mice expressing or not (WT) an ADNFLE-linked nAChR subunit (β2-V287L). More than 100,000 up-states were recorded during experiments sampling from several thousand neurons. Data were analyzed by using a fast sliding-window procedure which computes histograms of the up-state durations. Differently from the WT, cultures expressing β2-V287L displayed long (10–32 s) synaptic-induced up-state firing events. The occurrence of such long up-states was prevented by both negative (gabazine, penicillin G) and positive (benzodiazepines) modulators of GABAA receptors. Carbamazepine (CBZ), a drug of choice in ADNFLE patients, also inhibited the long up-states at micromolar concentrations. In cultures expressing β2-V287L, no significant effect was observed on the action potential waveform either in the absence or in the presence of pharmacological treatment. Our results show that some aspects of the spontaneous hyperexcitability displayed by a murine model of a human channelopathy can be reproduced in neuronal cultures. In particular, our cultures represent an in vitro chronic model of spontaneous epileptiform activity, i.e., not requiring pre-treatment with convulsants. This opens the way to the study in vitro of the role of β2-V287L on synaptic formation. Moreover, our neocortical cultures on MEA platforms allow to determine the effects of prolonged pharmacological treatment on spontaneous network hyperexcitability (which is impossible in the short-living brain slices). Methods such as the one we illustrate in the present paper should also considerably facilitate the preliminary screening of antiepileptic drugs (AEDs), thereby reducing the number of in vivo experiments.

Highlights

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is a mendelian partial epilepsy which belongs to the subgroup of familial focal epilepsies with autosomal dominant transmission pattern, which includes the familial temporal lobe epilepsy and familial focal epilepsy with variable foci (Engel, 2001)
These results suggest that heterogeneity of seizure-like events is a typical characteristic of networks expressing β2-V287L
THE EFFECT OF MODULATING GABAERGIC TRANSMISSION Because previous evidence in animal models of ADNFLE has pointed to alterations in the neocortical GABAergic transmission, we studied the effect of several GABAAR ligands on our cultures

Summary

Introduction

Autosomal dominant nocturnal frontal lobe epilepsy (ADNFLE) is a mendelian partial epilepsy which belongs to the subgroup of familial focal epilepsies with autosomal dominant transmission pattern, which includes the familial temporal lobe epilepsy and familial focal epilepsy with variable foci (Engel, 2001). ADNFLE is a good model of the more common sporadic cases of nonlesional NFLE, because the clinical and electroencephalographic features are similar (Picard and Brodtkorb, 2008). About 10–12% of the ADNFLE families bear mutations on genes coding for nicotinic acetylcholine receptors (nAChR) subunits (Steinlein et al, 1995; De Fusco et al, 2000; Phillips et al, 2001; Aridon et al, 2006).

Methods

Results

Conclusion