Elsevier

Experimental Neurology

Volume 269, July 2015, Pages 120-132
Experimental Neurology

Regular Article
Seizure reduction through interneuron-mediated entrainment using low frequency optical stimulation

https://doi.org/10.1016/j.expneurol.2015.04.001Get rights and content

Highlights

  • Low frequency optical stimulation significantly suppresses epileptiform activity in vitro and in vivo.

  • Selective optical activation of interneurons at low frequency suppresses epileptiform activity in the hippocampus.

  • Activation of GABA interneurons causes entrainment of hippocampal CA3 pyramidal cells by a GABAA mediated mechanism.

Abstract

Low frequency electrical stimulation (LFS) can reduce neural excitability and suppress seizures in animals and patients with epilepsy. However the therapeutic outcome could benefit from the determination of the cell types involved in seizure suppression. We used optogenetic techniques to investigate the role of interneurons in LFS (1 Hz) in the epileptogenic hippocampus. Optical low frequency stimulation (oLFS) was first used to activate the cation channel channelrhodopsin-2 (ChR2) in the Thy1-ChR2 transgenic mouse that expresses ChR2 in both excitatory and inhibitory neurons. We found that oLFS could effectively reduce epileptiform activity in the hippocampus through the activation of GAD-expressing hippocampal interneurons. This was confirmed using the VGAT-ChR2 transgenic mouse, allowing for selective optical activation of only GABA interneurons. Activating hippocampal interneurons through oLFS was found to cause entrainment of neural activity similar to electrical stimulation, but through a GABAA-mediated mechanism. These results confirm the robustness of the LFS paradigm and indicate that GABA interneurons play an unexpected role of shaping inter-ictal activity to decrease neural excitability in the hippocampus.

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).

References (54)

  • I. Vida et al.

    Shunting inhibition improves robustness of gamma oscillations in hippocampal interneuron networks by homogenizing firing rates

    Neuron

    (2006)
  • L.-X. Yang et al.

    Unilateral low-frequency stimulation of central piriform cortex delays seizure development induced by amygdaloid kindling in rats

    Neuroscience

    (2006)
  • S.-H. Zhang et al.

    Low-frequency stimulation of the hippocampal CA3 subfield is anti-epileptogenic and anti-ictogenic in rat amygdaloid kindling model of epilepsy

    Neurosci. Lett.

    (2009)
  • B.E. Alger et al.

    GABA-mediated biphasic inhibitory responses in hippocampus

    Nature

    (1979)
  • P. Andersen et al.

    Two different responses of hippocampal pyramidal cells to application of gamma-amino butyric acid

    J. Physiol.

    (1980)
  • A.N. Barclay et al.

    Localization of the Thy-1 antigen in rat brain and spinal cord by immunofluorescence

    J. Neurochem.

    (1978)
  • Y. Ben-Ari et al.

    Giant synaptic potentials in immature rat CA3 hippocampal neurones

    J. Physiol.

    (1989)
  • Y. Ben-Ari et al.

    GABA: a pioneer transmitter that excites immature neurons and generates primitive oscillations

    Physiol. Rev.

    (2007)
  • E.S. Boyden et al.

    Millisecond-timescale, genetically targeted optical control of neural activity

    Nat. Neurosci.

    (2005)
  • A. Bragin et al.

    Rate of interictal events and spontaneous seizures in epileptic rats after electrical stimulation of hippocampus and its afferents. [Internet]

    Epilepsia

    (2002)
  • B.S. Chang et al.

    Epilepsy

    N. Engl. J. Med.

    (2003)
  • J. Chavas et al.

    Coexistence of excitatory and inhibitory GABA synapses in the cerebellar interneuron network

    J. Neurosci.

    (2003)
  • B.Y. Chow et al.

    High-performance genetically targetable optical neural silencing by light-driven proton pumps

    Nature

    (2010)
  • S.R. Cobb et al.

    Synchronization of neuronal activity in hippocampus by individual GABAergic interneurons

    Nature

    (1995)
  • R. Fisher et al.

    Electrical stimulation of the anterior nucleus of thalamus for treatment of refractory epilepsy

    Epilepsia

    (2010)
  • T.J. Foutz et al.

    Theoretical principles underlying optical stimulation of a channelrhodopsin-2 positive pyramidal neuron

    J. Neurophysiol.

    (2012)
  • B.C. Jobst et al.

    Brain stimulation for the treatment of epilepsy

    Epilepsia

    (2010)
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