Paroxysmal Discharges in Tissue Slices From Pediatric Epilepsy Surgery Patients: Critical Role of GABAB Receptors in the Generation of Ictal Activity

Abstract

In the present study, we characterized the effects of bath application of the proconvulsant drug 4-aminopyridine (4-AP) alone or in combination with GABA A and/or GABA B receptor antagonists, in cortical dysplasia (CD type I and CD type IIa/b), tuberous sclerosis complex (TSC), and non-CD cortical tissue samples from pediatric epilepsy surgery patients. Whole-cell patch clamp recordings in current and voltage clamp modes were obtained from cortical pyramidal neurons (CPNs), interneurons, and balloon/giant cells. In pyramidal neurons, bath application of 4-AP produced an increase in spontaneous synaptic activity as well as rhythmic membrane oscillations. In current clamp mode, these oscillations were generally depolarizing or biphasic and were accompanied by increased membrane conductance. In interneurons, membrane oscillations were consistently depolarizing and accompanied by bursts of action potentials. In a subset of balloon/giant cells from CD type IIb and TSC cases, respectively, 4-AP induced very low-amplitude, slow membrane oscillations that echoed the rhythmic oscillations from pyramidal neurons and interneurons. Bicuculline reduced the amplitude of membrane oscillations induced by 4-AP, indicating that they were mediated principally by GABA A receptors. 4-AP alone or in combination with bicuculline increased cortical excitability but did not induce seizure-like discharges. Ictal activity was observed in pyramidal neurons and interneurons from CD and TSC cases only when phaclofen, a GABA B receptor antagonist, was added to the 4-AP and bicuculline solution. These results emphasize the critical and permissive role of GABA B receptors in the transition to an ictal state in pediatric CD tissue and highlight the importance of these receptors as a potential therapeutic target in pediatric epilepsy.

Characterizing the diversity of L2/3 human neocortical neurons in epilepsy

“…3H) and lower input resistance (MWU: p= 0.0323, MD= 50.17, 95%CI= 2.587 to 76.10) (Fig. 3I) indicating fAHP neurons are most likely not part of the LTS neuron subtype as these results contradict previous literature on regular LTS interneuron input resistance and voltage sag (Beierlein et al, 2003, Li et al, 2009, Tremblay et al, 2016, Levinson et al, 2020) 6,47–49 . The decrease in voltage sag of fAHP neurons may indicate deficits in HCN1 channels which play a role in Ih current, a contributor to enhanced frequency-dependent gain and possibly decreased excitability (Brennan et al, 2016, Herrmann et al, 2015, Kalmbach et al, 2018, Moradi Chameh et al, 2021) 7,15,50,51 .…”

“…3Aiii). Interestingly, L2/3 cortical pyramidal neurons in rats have been shown to have the fAHP followed by a fast ADP similar to Figure 3Ai that can trigger bursting which is a common phenotype seen in human epileptic brain tissue from cortex and hippocampus samples (Avoli et al, 1987, Higgs et al, 2009, Chen et al, 2011, Levinson et al, 2020) 6,32,43,46 .…”

“…L2/3 human pyramidal neurons are a diverse group of cells with various morphologies, electrophysiologic properties, and transcriptomic diversity making them difficult to characterize on electrophysiologic properties alone (Foehrig et al, 1991, DeFelipe et al, 1992, Deitcher et al,2017, Kalmbach et al, 2018, Berg et al, 2020) 5,8,9,15,41 . In addition, the influence of epileptic seizures on neuron firing properties is not well understood (Levinson et al, 2020, Moradi Chameh et al, 2021) 6,7 .Therefore, in order to confirm that we were successfully targeting pyramidal cells in our epileptic tissue slices, a subset of neurons were filled with biocytin and stained with streptavidin conjugated to Alexa Fluor 593 or 488 and three-dimensional (3D) neuron reconstructions were performed. Figure 1 illustrates the diversity of L2/3 putative pyramidal neuron action potential (APs) shapes and firing patterns that were induced by depolarizing current steps (600ms).…”

“…The etiology of epilepsy is immense and has been studied in vitro for the past few decades (Abdijadid et al, 2015, Avoli et al, 1991a, 1991b, 1994, Calcagnotto et al, 2005, Chang et al, 2019, Chen et al 2011, Cossart et al, 2001, Heinemann et al, 2014, Iffland and Crino, 2017, Jozwiak et al, 2006, Lasarge et al, 2014, Le Duiguo et al, 2018, Levinson et al, 2020, Lipton et al, 2014, Nakagawa et al, 2017, Lipton et al, 2014, O’Dell et al, 2012, Hodge et al, 2019, Saxena et al, 2015, Schwartzkroin et al, 1994, Schwartkroin et al, 1983, Schwarz et al, 2019, Ting et al, 2018a, 2018b, Toth et al, 2018, Yasin et al, 2013) 3,6,21,24–26,28,30,36,38,46,54–69 . With each etiology leading to differences in neuronal migration, differentiation, and synaptic development, a wide array of neuronal intrinsic properties and network level properties emerge.…”

GABAA Receptor-Mediated Epileptogenicity in Focal Cortical Dysplasia (FCD) Depends on Age at Epilepsy Onset

“…Increased spontaneous GABAergic activity and pacemaker GABA synaptic activity has been shown in samples with pathological high-frequency oscillations (HFOs) obtained from patients with FCD (Cepeda et al, 2014(Cepeda et al, , 2019. It has been recently shown that epileptiform synchronization, mediated by 4-aminopyridine (4AP)-containing medium, leading to in vitro ictal activity in the human FCD tissues may be facilitated by the decreased ability of GABA B receptors to control GABA release from interneuron terminals (D'Antuono et al, 2004;Levinson et al, 2020). Thus, it may be possible that such an increase in spontaneous GABA A receptor activity could be due to the altered function of presynaptic GABA B receptors.…”