In single channel studies Zn primarily inhibits GABA A recep

In single channel studies, Zn2+ primarily inhibits GABA-A receptors through the reduction of channel opening probability (Smart, 1992, Smart et al., 1994). In slice recordings, Zn2+ significantly reduces phasic mIPSC event amplitude and kinetics, as well as desensitization kinetics (Barberis et al., 2000, Ruiz et al., 2004). However, we demonstrated that GX-enhanced phasic mIPSCs were unaltered by Zn2+. It could be that GX potentiation overcomes Zn2+ depression of synaptic receptors so that the overall effects of Zn2+ on GX-potentiated phasic currents were undetectable. However, in tonic current potentiation, Zn2+ blocked GX-potentiated tonic response concentration-dependently in δ-rich DGGCs, probably due to its greater sensitivity towards δ-containing receptors. Unexpectedly, Zn2+ prolonged the fast decay time constant τ1 of mIPSCs in the presence of GX. One possibility might be due to the biphasic effects of Zn2+ on GABA-B receptors that tonically enhance GABA-A receptor currents (Turgeon and Albin, 1992, Khatri et al., 2018). In addition, receptor combinations with different subunits respond to Zn2+ and neurosteroids differently. Finally, Zn2+ may have different effects on fast-closing GABA-A receptor subtypes (Draguhn et al., 1990, Smart et al., 1991). Zn2+ has been shown to act on multiple neurotransmission systems in the brain. In addition to its inhibitory effects on GABA-A receptor function (Smart et al., 1991, Smart et al., 1994, Barberis et al., 2000, Carver et al., 2016), Zn2+ also antagonizes excitatory glutamate NMDA receptor kn 47 and decreases the activation and surface expression of NR2A-contaning NMDA receptors in hippocampal neurons (Westbrook and Mayer, 1987, Vogt et al., 2000, Zhu et al., 2012). Additionally, Zn2+ enhances the activity of AMPA receptors (Timofeeva and Nadler, 2006) and increases glycinergic neurotransmission. Therefore, it is possible that the excitability-facilitating and proconvulsant role of Zn2+ against GX-induced protective effects in the kindling seizure model may be resulted from the overall outcome of disrupted neurotransmission by Zn2+. Nevertheless, we have previously shown a critical role of extrasynaptic δGABA-A receptor in seizure susceptibility as mice lacking δ -subunit have significantly reduced tonic currents and greater seizure susceptibility (Carver et al., 2014). Thus, Zn2+ selective antagonistic interactions with GX at the extrasynaptic δGABA-A receptors in the hippocampus may contribute to the inhibition of GX-induced antiseizure activity by Zn2+. Zn2+ exhibits antagonistic activity at α4βγ- and α1βδ-containing GABA-A receptors, albeit at lower sensitivities than α4βδ subtypes (Brown et al., 2002). Specific deletion of δ-subunits in α1βδ-containing DG interneurons leads to reduced GABAergic tonic inhibition and elevated firing rate of interneurons that decrease DGGC excitability and in vivo seizure susceptibility (Lee and Maguire, 2013). Therefore, Zn2+ may also contribute to the disinhibition of α1βδ-containing interneurons. In the seizure model of hippocampal kindling, the composition and expression of GABA-A receptors in the hippocampus have been largely altered (Nishimura et al., 2005). In hippocampus DG, the mRNA levels of δ-subunit were significantly reduced with an increase in α2 -subunits seven days after kindling completion. In CA3 pyramidal neurons, α2 and β3-subunits were significantly upregulated after hippocampal kindling. While in CA1 pyramidal neurons, no significant changes of GABA-A receptor subunits were observed. As a result, the sensitivity of each cell types to Zn2+ modulation may be different from animals without kindling, which may also involve in the Zn2+ antagonism of GX-induced seizure protection. Nevertheless, we observed the net outcome of Zn2+ in hippocampal kindling to be proconvulsant. Zn2+ increases the permeability of blood-brain barrier and proconvulsant activity in rats (Yorulmaz et al., 2013). Severe blood-brain barrier damage is also found in pentylenetetrazol-induced epileptic seizures. The Zn2+ level in the brain may fluctuate due to the contribution of peripheral Zn2+ that penetrates the damaged blood-brain barrier. Zn2+ concentrations are particularly high in some brain regions including hippocampus and amygdala, which are also the most vulnerable regions for the focal point of seizures (Frederickson et al., 2005). The effective concentrations of Zn2+ in the inhibition of GX-potentiated tonic currents are within the levels that may occur in synaptic clefts following the Zn2+ release from the vesicles of presynaptic neurons during neuronal activity. Therefore, Zn2+ may disrupt the neurosteroid-augmented basal inhibitory tone and hinder the balance of hippocampal neural circuits. In physiological condition, free zinc cations released from glutamatergic synapses in the hippocampus may block the inhibition of extrasynaptic activity of neurosteroids, hindering the antiseizure effects of neurosteroids. Therefore, combination therapies of neurosteroids with Zn2+ chelators may be potential avenues for the treatment of seizure-related disorders. We have attempted to examine the effects of Zn2+ chelators on the seizure activity of hippocampal kindling animals. However, a previous study shows that TPEN exerts notable toxic effects in in vivo animal studies, which was the reason these studies were not conducted further (Elsas et al., 2009). In Foresti et al., 2008, they used another Zn2+ chelator and found that pretreatment of Zn2+ chelator DEDTC (700 mg/kg) diminishes the duration of behavioral seizures and electrical afterdischarges, and EEG spikes, without altering seizure severity progression in a rat model of amygdala kindling, showing the antiseizure effects of the Zn2+ chelator (Foresti et al., 2008). Although their experiments show an antiseizure role of the zinc chelator, they cannot exclude the potential toxic effects of DEDTC. In their studies, they selected a dose that did not cause significant behavioral changes but still may be high enough to cause toxic effects in cellular and molecular levels since animals experienced severe ocular and nasal bleeding and reduced locomotor activity with the treatment of DEDTC. Future studies of zinc chelators on the seizure activity are feasible when nontoxic Zn2+ chelators become available.