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Hippocampal GAD67 Transduction Using rAAV8 Regulates Epileptogenesis in EL Mice.

ABSTRACT: Gene therapy has been employed as a therapeutic approach for intractable focal epilepsies. Considering the potential of focal GABAergic neuromodulation in regulating epileptogenesis, the GABA-producing enzyme, ?-aminobutyric acid decarboxylase 67 (GAD67), is highly suitable for epilepsy therapy. The EL/Suz (EL) mouse is a model of multifactorial temporal lobe epilepsy. In the present study, we examined focal gene transduction in epileptic EL mice using recombinant adeno-associated virus serotype 8 (rAAV8) expressing human GAD67 to enhance GABA-mediated neural inhibition. Eight-week-old mice were bilaterally injected with rAAV8-GFP or rAAV8-GAD67 in the hippocampal CA3 region. After four weeks, the GAD67-transduced EL mice, but not the rAAV-GFP-treated EL mice, exhibited a significant reduction in seizure generation. The GAD67-mediated depression became stable after 14 weeks. The excitability of the CA3 region was markedly reduced in the GAD67-transduced EL mice, consistent with the results of the Ca2+ imaging using hippocampal slices. In addition, downregulation of c-Fos expression was observed in GAD67-transduced hippocampi. Our findings showed that rAAV8-GAD67 induced significant changes in the GABAergic system in the EL hippocampus. Thus, rAAV8-mediated GAD67 gene transfer is a promising therapeutic strategy for the treatment of epilepsies.

SUBMITTER: Shimazaki K

PROVIDER: S-EPMC6369250 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Hippocampal GAD67 Transduction Using rAAV8 Regulates Epileptogenesis in EL Mice.

Shimazaki Kuniko K Kobari Takashi T Oguro Keiji K Yokota Hidenori H Kasahara Yuko Y Murashima Yoshiya Y Watanabe Eiju E Kawai Kensuke K Okada Takashi T

Molecular therapy. Methods & clinical development 20190108

Gene therapy has been employed as a therapeutic approach for intractable focal epilepsies. Considering the potential of focal GABAergic neuromodulation in regulating epileptogenesis, the GABA-producing enzyme, γ-aminobutyric acid decarboxylase 67 (GAD67), is highly suitable for epilepsy therapy. The EL/Suz (EL) mouse is a model of multifactorial temporal lobe epilepsy. In the present study, we examined focal gene transduction in epileptic EL mice using recombinant adeno-associated virus serotype ...[more]

PMID: 30788386

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Project description:ObjectiveTissue remodeling has been described in brain circuits that are involved in the generation and propagation of epileptic seizures. Human and animal studies suggest that the anterior piriform cortex (aPC) is crucial for seizure expression in focal epilepsies. Here, we investigate the effect of kainic-acid (KA)-induced seizures on the effective connectivity of the aPC with bilateral hippocampal CA3 regions using cerebro-cerebral evoked potentials (CCEPs).MethodsAdult male Sprague-Dawley rats were implanted with a tripolar electrode in the left aPC for stimulation and recording, and with unipolar recording electrodes in bilateral CA3 regions. Single pulse stimulations were given to the aPC and CCEPs were averaged before KA injections and after the emergence of spontaneous recurrent seizures (SRS). Similar recordings at equivalent time intervals were obtained from animals that received saline injections instead of KA (controls).ResultsIn the experimental group, the percentage change of increased amplitude of the contralateral (but not ipsilateral) CA3 CCEPs between pre-KA injection and after the emergence of SRS was significantly greater than in controls. No significant single-pulse-induced spectral change responses were observed in either epileptic or control rats when comparing pre- and post-stimulus time intervals. Also, we found no correlation between seizure frequency and the extent of amplitude changes in the CCEPs.ConclusionsIn the KA model, epileptogenesis results in plastic changes that manifest as an amplification of evoked potential amplitudes recorded in the contralateral hippocampus in response to single-pulse stimulation of the aPC. These results suggest epileptogenesis-induced facilitation of interhemispheric connectivity between the aPC and the hippocampus. Since the amplitude increase of the contralateral CCEP is a possible in vivo biomarker of epilepsy, any intervention (e.g. neuromodulatory) that can reverse this phenomenon may hold a potential antiepileptic efficacy.

Project description:ObjectiveHomeostatic synaptic plasticity (HSP) serves as a gain control mechanism at central nervous system (CNS) synapses, including those between the dentate gyrus (DG) and CA3. Improper circuit control of DG-CA3 synapses is hypothesized to underlie epileptogenesis. Here, we sought to (1) identify compounds that preferentially modulate DG-CA3 synapses in primary neuronal culture and (2) determine if these compounds would delay or prevent epileptogenesis in vivo.MethodsWe previously developed and validated an in vitro assay to visualize the behavior of DG-CA3 synapses and predict functional changes. We used this "synapse-on-chip" assay (quantification of synapse size, number, and type using immunocytochemical markers) to dissect the mechanisms of HSP at DG-CA3 synapses. Using chemogenetic constructs and pharmacological agents we determined the signaling cascades necessary for gain control at DG-CA3 synapses. Finally, we tested the implicated cascades (using kappa opioid receptor (OR) agonists and antagonists) in two models of epileptogenesis: electrical amygdala kindling in the mouse and chemical (pentylenetetrazole) kindling in the rat.ResultsIn vitro, synapses between DG mossy fibers (MFs) and CA3 neurons are the primary homeostatic responders during sustained periods of activity change. Kappa OR signaling is both necessary and sufficient for the homeostatic elaboration of DG-CA3 synapses, induced by presynaptic DG activity levels. Blocking kappa OR signaling in vivo attenuates the development of seizures in both mouse and rat models of epilepsy.SignificanceThis study elucidates mechanisms by which synapses between DG granule cells and CA3 pyramidal neurons undergo activity-dependent homeostatic compensation, via OR signaling in vitro. Modulation of kappa OR signaling in vivo alters seizure progression, suggesting that breakdown of homeostatic closed-loop control at DG-CA3 synapses contributes to seizures, and that targeting endogenous homeostatic mechanisms at DG-CA3 synapses may prove useful in combating epileptogenesis.

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Hippocampal GAD67 Transduction Using rAAV8 Regulates Epileptogenesis in EL Mice.

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Hippocampal GAD67 Transduction Using rAAV8 Regulates Epileptogenesis in EL Mice.

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