Regular ArticleSeizure reduction through interneuron-mediated entrainment using low frequency optical stimulation
Introduction
Epilepsy is a chronic disorder of the central nervous system characterized by recurrent, unprovoked seizures. Mesial temporal lobe epilepsy (MTLE) involving the hippocampus is a common type of epilepsy, often medically refractory and necessitating surgical resection if an epileptic focus can be identified (Wiebe et al., 2001). A less invasive alternative to surgery shown to reduce disease burden is deep brain electrical stimulation (DBS) (Fisher et al., 2010, Jobst et al., 2010, Morrell, 2011), although the optimal location and stimulus paradigm is highly debated and the mechanisms of seizure suppression remain poorly understood (Sunderam et al., 2010). Nonetheless, recent studies have shown low frequency electrical stimulation to be effective at suppressing epileptiform activity in animal models (Rashid et al., 2012) and reducing seizure frequency in patients (Koubeissi et al., 2013).
Despite the advantages offered by a low frequency electrical treatment strategy, the inherent lack of cell type-specificity in using an electrical stimulus to modulate neuronal activity may limit overall efficacy and create undesired side effects. The confounding effects from non-specific cell activation also make it difficult to determine the mechanisms responsible for the therapeutic effect. There is much debate over whether stimulation of the neurons at the seizure focus (Bragin et al., 2002), afferent connections (Yang et al., 2006), or even local glial cells (Tawfik et al., 2010) are important for reducing seizure activity. The emerging field of optogenetics can help address these questions by providing tools that allow for cell-specific activation (Boyden et al., 2005) or inhibition (Zhang et al., 2007, Chow et al., 2010) in a reversible manner with millisecond time resolution. These optogenetic constructs have recently been shown to suppress neuronal hyperactivity and seizure in various models epilepsy (Paz et al., 2013, Wykes et al., 2012, Krook-Magnuson et al., 2013) but can also be applied to elucidate the mechanisms seizure suppression.
In the present study, we used optogenetic techniques to investigate the role of interneurons in low-frequency stimulation paradigms that have been shown to suppress epileptiform activity in the hippocampus (Rashid et al., 2012, Koubeissi et al., 2013). Two different optogenetic transgenic mouse models expressing ChR2 were used to study the effects of selectively activating hippocampal neurons or only GABA interneurons to assess the role these cell types play in suppressing seizure activity. The pro-epileptogenic compound 4-aminopyridine (4-AP) was used as a model of epilepsy (Perreault and Avoli, 1989) in both in vitro (Zhang et al., 2014) and acute in vivo (Gonzalez-Reyes et al., 2013) preparations. This compound has been shown to trigger epileptiform activity in vivo after local (intra-hippocampal) (Gonzalez-Reyes et al., 2013) and systemic (Lévesque et al., 2013) administration. While this work was primarily aimed to gain insight into the mechanisms of seizure reduction using LFS, the efficacy of this approach in vivo adds to the growing body of evidence that translation of optogenetic techniques can provide more targeted therapy for future clinical applications.
Section snippets
Animals
Thy1-ChR2-YFP (B6.Cg-Tg(Thy1-COP4/EYFP)9Gfng/J Stock Number:007615) and VGAT-ChR2-YFP (B6.Cg-Tg(Slc32a1-COP4*H134R/EYFP)8Gfng/J Stock Number: 014548) transgenic mice (both of C57BL/6 background) were bred from founder animals obtained from the Jackson Laboratory. We used these two transgenic animals since the promoters Thy-1 (Arenkiel et al., 2007) and VGAT (Zhao et al., 2011) control the expression of the ChR2 opsin only in neurons and in GABAergic cells, respectively. Animals were housed no
Optical LFS can suppress epileptiform activity in the hippocampus in vivo
LFS applied electrically to the ventral hippocampal commissure in rats can suppress 90% of the seizures by activating both hippocampi (Rashid et al., 2012). To determine if direct oLFS in the hippocampus could reproduce a similar seizure suppression effect, we first applied oLFS in Thy1-ChR2-YFP mice (Arenkiel et al., 2007) where both excitatory and inhibitory neurons would be optically activated, as with electrical stimuli. Since the efficacy of an optical stimulus applied to suppress seizure
Discussion
In this study, we used a novel optogenetic method of low-frequency stimulation in the hippocampus to investigate the role of hippocampal interneurons in LFS-mediated network suppression in epilepsy. Neuronal activation with the excitatory construct channelrhodopsin-2 was used as an analog to an electrical stimulus, but with the advantage of cell type-specificity through genetic targeting. We showed that oLFS could effectively reduce epileptiform activity in the hippocampus. Selective activation
Acknowledgments
This work was funded by the National Institutes of Health (NINDS) Grant# 5R01NS040785-04 and the Case Western Reserve University MSTP fellowship (NIH T32 GM007250).
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