Model Of Mesial Temporal Lobe Epilepsy Research Articles

Human pallial MGE-type GABAergic interneuron cell therapy for chronic focal epilepsy.

Mesial temporal lobe epilepsy (MTLE) is the most common focal epilepsy. One-third of patients have drug-refractory seizures and are left with suboptimal therapeutic options such as brain tissue-destructive surgery. Here, we report the development and characterization of a cell therapy alternative for drug-resistant MTLE, which is derived from a human embryonic stem cell line and comprises cryopreserved, post-mitotic, medial ganglionic eminence (MGE) pallial-type GABAergic interneurons. Single-dose intrahippocampal delivery of the interneurons in a mouse model of chronic MTLE resulted in consistent mesiotemporal seizure suppression, with most animals becoming seizure-free and surviving longer. The grafted interneurons dispersed locally, functionally integrated, persisted long term, and significantly reduced dentate granule cell dispersion, a pathological hallmark of MTLE. These disease-modifying effects were dose-dependent, with a broad therapeutic range. No adverse effects were observed. These findings support an ongoing phase 1/2 clinical trial (NCT05135091) for drug-resistant MTLE.

E2730, an uncompetitive γ-aminobutyric acid transporter-1 inhibitor, suppresses epileptic seizures in a rat model of chronic mesial temporal lobe epilepsy.

More than one third of mesial temporal lobe epilepsy (MTLE) patients are resistant to current antiseizure medications (ASMs), and half experience mild-to-moderate adverse effects of ASMs. There is therefore a strong need to develop and test novel ASMs. The objective of this work is to evaluate the pharmacokinetics and neurological toxicity of E2730, a novel uncompetitive inhibitor of γ-aminobutyric acid transporter-1, and to test its seizure suppression effects in a rat model of chronic MTLE. We first examined plasma levels and adverse neurological effects of E2730 in healthy Wistar rats. Adult male rats were implanted with osmotic pumps delivering either 10, 20, or 100 mg/kg/day of E2730 subcutaneously for 1 week. Blood sampling and behavioral assessments were performed at several timepoints. We next examined whether E2730 suppressed seizures in rats with chronic MTLE. These rats were exposed to kainic acid-induced status epilepticus, and 9 weeks later, when chronic epilepsy was established, were assigned to receive one of the three doses of E2730 or vehicle for 1 week in a randomized crossover design. Continuous video-electroencephalographic monitoring was acquired during the treatment period to evaluate epileptic seizures. Plasma levels following continuous infusion of E2730 showed a clear dose-related increase in concentration. The drug was well tolerated at all doses, and any sedation or neuromotor impairment was mild and transient, resolving within 48 h of treatment initiation. Remarkably, E2730 treatment in chronically epileptic rats led to seizure suppression in a dose-dependent manner, with 65% of rats becoming seizure-free at the highest dose tested. Mean seizure class did not differ between the treatment groups. This study shows that continuous subcutaneous infusion of E2730 over 7 days results in a marked, dose-dependent suppression of spontaneous recurrent seizures, with minimal adverse neurological effects, in a rat model of chronic MTLE. E2730 shows strong promise as an effective new ASM to be translated into clinical trials.

Optogenetic activation of septal inhibitory cells abates focal seizures.

Emerging evidence suggests that the medial septum can control seizures occurring in focal epileptic disorders, thus representing a therapeutic target. Therefore, we investigated whether continuous optogenetic activation of inhibitory parvalbumin (PV)-positive interneurons in the medial septum can reduce the occurrence of spontaneous seizures in the pilocarpine model of mesial temporal lobe epilepsy (MTLE). Light pulses (450 nm, 25 mW, 20-ms pulse duration) were delivered at 0.5 Hz (5 min ON, 10 min OFF) with a laser diode fiber light source between day 8 and day 12 after status epilepticus (SE) in PV-ChR2 mice (n = 8). Seizure rates were significantly lower during time periods of optogenetic stimulation (days 8-12) compared with before implementation of optogenetics (days 4-7) (P < 0.05). Moreover, between day 13 and day 21 after SE seizure rates were still significantly lower compared with before optogenetic stimulation (i.e., between day 4 and day 7) (P < 0.05). No seizures were recorded between day 10 and day 12 in all animals, and no seizures occurred up to 3 days after the end of optogenetic stimulation (days 13-15). Our findings indicate that activation of PV interneurons in the medial septum abates seizures in the pilocarpine model of MTLE. Moreover, the persisting anti-ictogenic effects suggest that stimulation of the medial septum could alter the progression of MTLE.NEW & NOTEWORTHY The medial septum could represent a therapeutic target to treat patients with focal epilepsy. In this study, we show that optogenetic activation of inhibitory parvalbumin-positive interneurons in the medial septum can block spontaneous seizures and prevents their reoccurrence for ∼5 days after the end of stimulation. Our findings suggest that the anti-ictogenic effects induced by stimulation of the medial septum could also alter the progression of mesial temporal lobe epilepsy.

Pronounced antiseizure activity of the subtype-selective GABAA positive allosteric modulator darigabat in a mouse model of drug-resistant focal epilepsy.

AimDarigabat is an α2/3/5 subunit‐selective positive allosteric modulator of GABAA receptors that has demonstrated broad‐spectrum activity in several preclinical models of epilepsy as well as in a clinical photoepilepsy trial. The objective here was to assess the acute antiseizure effect of darigabat in the mesial temporal lobe epilepsy (MTLE) mouse model of drug‐resistant focal seizures.MethodsThe MTLE model is generated by single unilateral intrahippocampal injection of low dose (1 nmole) kainic acid in adult mice, and subsequent epileptiform activity is recorded following implantation of a bipolar electrode under general anesthesia. After a period of epileptogenesis (~4 weeks), spontaneous and recurrent hippocampal paroxysmal discharges (HPD; focal seizures) are recorded using intracerebral electroencephalography. The number and cumulated duration of HPDs were recorded following administration of vehicle (PO), darigabat (0.3–10 mg kg−1, PO), and positive control diazepam (2 mg kg−1, IP).RESULTSDarigabat dose‐dependently reduced the expression of HPDs, demonstrating comparable efficacy profile to diazepam at doses of 3 and 10 mg kg−1.CONCLUSIONSDarigabat exhibited a robust efficacy profile in the MTLE model, a preclinical model of drug‐resistant focal epilepsy. A Phase II proof‐of‐concept placebo‐controlled, adjunctive‐therapy trial (NCT04244175) is ongoing to evaluate efficacy and safety of darigabat in patients with drug‐resistant focal seizures.

Opposing reduced VPAC1 and enhanced VPAC2 VIP receptors in the hippocampus of the Li2+-pilocarpine rat model of temporal lobe epilepsy

VIP binding sites are upregulated in mesial temporal lobe epilepsy (MTLE) patients, also suffering from severe cognitive deficits. Although altered VIP and VIP receptor levels were described in rodent models of epilepsy, the VIP receptor subtype(s) were never identified. We now investigated how VPAC1 and VPAC2 receptor levels change in the Li2+-pilocarpine rat model of MTLE. Cognitive decline and altered synaptic plasticity as estimated from phosphorylation of AMPA GluA1 subunit on Ser831 and Ser845 and AMPA GluA1/GluA2 ratio was also probed. Animals showing spontaneous recurrent seizures (SRSs) for at least 4 weeks showed impaired learning in the radial arm maze (RAM) and presented decreased VPAC1 and increased VPAC2 receptor levels. In addition, SRSs rats showed increased AMPA GluA1 phosphorylation in Ser831 and Ser845, marked decrease in GluA1 levels and a milder decrease in GluA2 levels. Consequently, the GluA1/GluA2 ratio was also decreased in SRSs rats.Altered VIP receptor levels may differentially prevent or contribute to MTLE pathology, since VPAC1 receptors promote the endogenous control of LTP, mediate endogenous VIP neuroprotection against altered synaptic plasticity following epileptiform activity, and mediate anti-inflammatory actions in microglia, while VPAC2 receptors mediate VIP endogenous neuroprotection against neonatal excitotoxicity and prevent reactive astrogliosis. This discovery imposes a different mindset for considering VIP receptors as therapeutic targets in MTLE, allowing a differential targeting of the cellular events contributing to epileptogenesis.

Bilateral optogenetic activation of inhibitory cells favors ictogenesis

Mesial temporal lobe epilepsy (MTLE) is the most common type of focal refractory epilepsy and is characterized by recurring seizures that are often refractory to medication. Since parvalbumin-positive (PV) interneurons were recently shown to play significant roles in ictogenesis, we established here how bilateral optogenetic stimulation of these interneurons in the hippocampus CA3 regions modulates seizures, interictal spikes and high-frequency oscillations (HFOs; ripples: 80–200 Hz, fast ripples: 250–500 Hz) in the pilocarpine model of MTLE. Bilateral optogenetic stimulation of CA3 PV-positive interneurons at 8 Hz (lasting 30 s, every 2 min) was implemented in PV-ChR2 mice for 8 consecutive days starting on day 7 (n = 8) or on day 13 (n = 6) after pilocarpine-induced status epilepticus (SE). Seizure occurrence was higher in both day 7 and day 13 groups of PV-ChR2 mice during periods of optogenetic stimulation (“ON”), compared to when stimulation was not performed (“OFF”) (day 7 group = p < 0.01, day 13 group = p < 0.01). In the PV-ChR2 day 13 group, rates of seizures (p < 0.05), of interictal spikes associated with fast ripples (p < 0.01), and of isolated fast ripples (p < 0.01) during optogenetic stimulations were significantly higher than in the PV-ChR2 day 7 group. Our findings reveal that bilateral activation of PV-interneurons in the hippocampus (leading to a presumptive increase in GABA signaling) favors ictogenesis. These effects may also mirror the neuropathological changes that occur over time after SE in this animal model.

Seizure activity triggers tau hyperphosphorylation and amyloidogenic pathways.

Although epilepsies and neurodegenerative disorders show pathophysiological similarities, their direct functional associations are unclear. Here, we tested the hypothesis that experimental seizures can induce tau hyperphosphorylation and amyloidogenic modifications over time, with intersections with neuroinflammation. We used a model of mesial temporal lobe epilepsy (MTLE) where unilateral intrahippocampal injection of kainic acid (KA) in C57BL/6mice elicits epileptogenesis and spontaneous focal seizures. We used a model of generalized status epilepticus (SE) obtained by intraperitoneal KA injection in C57BL/6mice. We performed analyses and cross-comparisons according to a schedule of 72 h, 1 week, and 8 weeks after KA injection. In experimental MTLE, we show AT100, PHF1, and CP13 tau hyperphosphorylation during epileptogenesis (72h-1week) and long-term (8weeks) during spontaneous seizures in the ipsilateral hippocampi, the epileptogenic zone. These pathological modifications extended to the contralateral hippocampus, a seizure propagating zone with no histological lesion or sclerosis. Two kinases, Cdk5 and GSK3β, implicated in the pathological phosphorylation of tau, were activated. In this MTLE model, the induction of the amyloidogenic pathway (APP, C99, BACE1) was prominent and long-lasting in the epileptogenic zone. These Alzheimer's disease (AD)-relevant markers, established during seizure progression and recurrence, reciprocated an enduring glial (GFAP, Iba1) inflammation and the inadequate activation of the endogenous, anti-inflammatory, glucocorticoid receptor system. By contrast, a generalized SE episode provoked a predominantly transient induction of tau hyperphosphorylation and amyloidogenic markers in the hippocampus, along with resolving inflammation. Finally, we identified overlapping profiles of long-term hippocampal tau hyperphosphorylation by comparing MTLE to J20mice, the latter a model relevant to AD. MTLE and a generalized SE prompt persistent and varying tau hyperphosphorylation or amyloidogenic modifications in the hippocampus. In MTLE, an AD-relevant molecular trajectory intertwines with neuroinflammation, spatiotemporally involving epileptogenic and nonlesional seizure propagating zones.

Spontaneous recurrent seizures in an intra-amygdala kainate microinjection model of temporal lobe epilepsy are differentially sensitive to antiseizure drugs

The discovery and development of novel antiseizure drugs (ASDs) that are effective in controlling pharmacoresistant spontaneous recurrent seizures (SRSs) continues to represent a significant unmet clinical need. The Epilepsy Therapy Screening Program (ETSP) has undertaken efforts to address this need by adopting animal models that represent the salient features of human pharmacoresistant epilepsy and employing these models for preclinical testing of investigational ASDs. One such model that has garnered increased interest in recent years is the mouse variant of the Intra-Amygdala Kainate (IAK) microinjection model of mesial temporal lobe epilepsy (MTLE). In establishing a version of this model, several methodological variables were evaluated for their effect(s) on pertinent quantitative endpoints. Although administration of a benzodiazepine 40 min after kainate (KA) induced status epilepticus (SE) is commonly used to improve survival, data presented here demonstrates similar outcomes (mortality, hippocampal damage, latency periods, and 90-day SRS natural history) between mice given midazolam and those that were not. Using a version of this model that did not interrupt SE with a benzodiazepine, a 90-day natural history study was performed and survival, latency periods, SRS frequencies and durations, and SRS clustering data were quantified. Finally, an important step towards model adoption is to assess the sensitivities or resistances of SRSs to a panel of approved and clinically used ASDs. Accordingly, the following ASDs were evaluated for their effects on SRSs in these mice: phenytoin (20 mg/kg, b.i.d.), carbamazepine (30 mg/kg, t.i.d.), valproate (240 mg/kg, t.i.d.), diazepam (4 mg/kg, b.i.d.), and phenobarbital (25 and 50 mg/kg, b.i.d.). Valproate, diazepam, and phenobarbital significantly attenuated SRS frequency relative to vehicle controls at doses devoid of observable adverse behavioral effects. Only diazepam significantly increased seizure freedom. Neither phenytoin nor carbamazepine significantly altered SRS frequency or freedom under these experimental conditions. These data demonstrate that SRSs in this IAK model of MTLE are pharmacoresistant to two representative sodium channel-inhibiting ASDs (phenytoin and carbamazepine) and partially sensitive to GABA receptor modulating ASDs (diazepam and phenobarbital) or a mixed-mechanism ASD (valproate). Accordingly, this model is being incorporated into the NINDS-funded ETSP testing platform for treatment resistant epilepsy.

Evolution of interictal spiking during the latent period in a mouse model of mesial temporal lobe epilepsy.

Interictal spikes and high-frequency oscillations (HFOs, ripples: 80–200 ​Hz, fast ripples: 250–500 ​Hz) occur in epileptic patients and in animal models of mesial temporal lobe epilepsy (MTLE). In this study, we explored how type 1 and type 2 interictal spikes as well as ripples and fast ripples evolve during the latent period in the hippocampus of pilocarpine-treated mice. Depth EEG recordings were obtained from the hippocampus CA3 subfield of adult male mice (n ​= ​5, P60–P100) starting one day before pilocarpine-induced status epilepticus up to the first spontaneous seizure, the so-called latent period. We found that rates of type 1 (n ​= ​1 655) and type 2 (n ​= ​2 309) interictal spikes were significantly lower during the late phase of the latent period compared to its early and mid phase (p ​< ​0.001). However, rates of type 1 spikes associated with ripples (n ​= ​266) or fast ripples (n ​= ​106), as well as rates of type 2 interictal spikes associated with ripples (n ​= ​233), were significantly higher during the late phase compared to the early and mid phases (p ​< ​0.05). Our findings reveal that an increase of type 1 interictal spikes co-occurring with ripples or fast ripples and an increase of type 2 interictal spikes co-occurring with ripples mark the end of the latent period. We propose that changes in the occurrence of interictal spike associated with HFOs represent a biomarker of epileptogenicity in this mouse model of MTLE.

Hippocampal low-frequency stimulation prevents seizure generation in a mouse model of mesial temporal lobe epilepsy.

Mesial temporal lobe epilepsy (MTLE) is the most common form of focal, pharmacoresistant epilepsy in adults and is often associated with hippocampal sclerosis. Here, we established the efficacy of optogenetic and electrical low-frequency stimulation (LFS) in interfering with seizure generation in a mouse model of MTLE. Specifically, we applied LFS in the sclerotic hippocampus to study the effects on spontaneous subclinical and evoked generalized seizures. We found that stimulation at 1 Hz for 1 hr resulted in an almost complete suppression of spontaneous seizures in both hippocampi. This seizure-suppressive action during daily stimulation remained stable over several weeks. Furthermore, LFS for 30 min before a pro-convulsive stimulus successfully prevented seizure generalization. Finally, acute slice experiments revealed a reduced efficacy of perforant path transmission onto granule cells upon LFS. Taken together, our results suggest that hippocampal LFS constitutes a promising approach for seizure control in MTLE.

Multi-omics analysis suggests enhanced epileptogenesis in the Cornu Ammonis 3 of the pilocarpine model of mesial temporal lobe epilepsy.

Mesial temporal lobe epilepsy (MTLE) is a chronic neurological disorder characterized by the occurrence of seizures, and histopathological abnormalities in the mesial temporal lobe structures, mainly hippocampal sclerosis (HS). We used a multi-omics approach to determine the profile of transcript and protein expression in the dorsal and ventral hippocampal dentate gyrus (DG) and Cornu Ammonis 3 (CA3) in an animal model of MTLE induced by pilocarpine. We performed label-free proteomics and RNAseq from laser-microdissected tissue isolated from pilocarpine-induced Wistar rats. We divided the DG and CA3 into dorsal and ventral areas and analyzed them separately. We performed a data integration analysis and evaluated enriched signaling pathways, as well as the integrated networks generated based on the gene ontology processes. Our results indicate differences in the transcriptomic and proteomic profiles among the DG and the CA3 subfields of the hippocampus. Moreover, our data suggest that epileptogenesis is enhanced in the CA3 region when compared to the DG, with most abnormalities in transcript and protein levels occurring in the CA3. Furthermore, our results show that the epileptogenesis in the pilocarpine model involves predominantly abnormal regulation of excitatory neuronal mechanisms mediated by N-methyl D-aspartate (NMDA) receptors, changes in the serotonin signaling, and neuronal activity controlled by calcium/calmodulin-dependent protein kinase (CaMK) regulation and leucine-rich repeat kinase 2 (LRRK2)/WNT signaling pathways.

Histological Correlates of Diffusion-Weighted Magnetic Resonance Microscopy in a Mouse Model of Mesial Temporal Lobe Epilepsy.

Mesial temporal lobe epilepsy (MTLE) is the most common type of focal epilepsy. It is frequently associated with abnormal MRI findings, which are caused by underlying cellular, structural, and chemical changes at the micro-scale. In the current study, it is investigated to which extent these alterations correspond to imaging features detected by high resolution magnetic resonance imaging in the intrahippocampal kainate mouse model of MTLE. Fixed hippocampal and whole-brain sections of mouse brain tissue from nine animals under physiological and chronically epileptic conditions were examined using structural and diffusion-weighted MRI. Microstructural details were investigated based on a direct comparison with immunohistochemical analyses of the same specimen. Within the hippocampal formation, diffusion streamlines could be visualized corresponding to dendrites of CA1 pyramidal cells and granule cells, as well as mossy fibers and Schaffer collaterals. Statistically significant changes in diffusivities, fractional anisotropy, and diffusion orientations could be detected in tissue samples from chronically epileptic animals compared to healthy controls, corresponding to microstructural alterations (degeneration of pyramidal cells, dispersion of the granule cell layer, and sprouting of mossy fibers). The diffusion parameters were significantly correlated with histologically determined cell densities. These findings demonstrate that high-resolution diffusion-weighted MRI can resolve subtle microstructural changes in epileptic hippocampal tissue corresponding to histopathological features in MTLE.

Paradoxical effects of optogenetic stimulation in mesial temporal lobe epilepsy.

To establish the effects induced by long-term, unilateral stimulation of parvalbumin (PV)-positive interneurons on seizures, interictal spikes, and high-frequency oscillations (80-500Hz) occurring after pilocarpine-induced status epilepticus (SE)-a proven model of mesial temporal lobe epilepsy (MTLE)-in transgenic mice expressing or not expressing ChR2. PV-ChR2 (n = 6) and PV-Cre (n = 6) mice were treated with pilocarpine to induce SE. Three hours after SE onset, unilateral optogenetic stimulation (450nm, 25mW, 20-millisecond pulses delivered at 8Hz for 30 seconds every 2 minutes) of CA3 PV-positive interneurons was implemented for 14 continuous days in both groups. Rates of seizures (p < 0.01), interictal spikes (p < 0.001), and interictal spikes with fast ripples (250-500Hz) (p < 0.001) were lower in PV-ChR2 than in PV-Cre mice. Ripples (80-200Hz) occurring outside of interictal spikes had higher rates in the PV-ChR2 group (p < 0.01), whereas isolated fast ripples had lower rates (p < 0.01). However, seizure probability was higher during optogenetic stimulation in PV-ChR2 compared to PV-Cre animals (p < 0.05). Our findings show that the unilateral activation of CA3 PV-positive interneurons exerts anti-ictogenic effects associated with decreased rates of interictal spikes and fast ripples in this MTLE model. However, PV-positive interneuron stimulation can paradoxically trigger seizures in epileptic animals, supporting the notion that γ-aminobutyric acid type A signaling can also initiate ictogenesis. ANN NEUROL 2019;86:714-728.

Reactive Disruption of the Hippocampal Neurogenic Niche After Induction of Seizures by Injection of Kainic Acid in the Amygdala.

Adult neurogenesis persists in the adult hippocampus due to the presence of multipotent neural stem cells (NSCs). Hippocampal neurogenesis is involved in a range of cognitive functions and is tightly regulated by neuronal activity. NSCs respond promptly to physiological and pathological stimuli altering their neurogenic and gliogenic potential. In a mouse model of mesial temporal lobe epilepsy (MTLE), seizures triggered by the intrahippocampal injection of the glutamate receptor agonist kainic acid (KA) induce NSCs to convert into reactive NSCs (React-NSCs) which stop producing new neurons and ultimately generate reactive astrocytes thus contributing to the development of hippocampal sclerosis and abolishing neurogenesis. We herein show how seizures triggered by the injection of KA in the amygdala, an alternative model of MTLE which allows parallel experimental manipulation in the dentate gyrus, also trigger the induction of React-NSCs and provoke the disruption of the neurogenic niche resulting in impaired neurogenesis. These results highlight the sensitivity of NSCs to the surrounding neuronal circuit activity and demonstrate that the induction of React-NSCs and the disruption of the neurogenic niche are not due to the direct effect of KA in the hippocampus. These results also suggest that neurogenesis might be lost in the hippocampus of patients with MTLE. Indeed we provide results from human MTLE samples absence of cell proliferation, of neural stem cell-like cells and of neurogenesis.

Time-dependent evolution of seizures in a model of mesial temporal lobe epilepsy

Low-voltage fast (LVF) and hypersynchronous (HYP) - onset seizures occur in the EEG obtained with depth electrodes from mesial temporal lobe epilepsy (MTLE) patients and animal models. In epileptic rats analyzed up to approximately two weeks after pilocarpine-induced status epilepticus (SE), these patterns are associated with specific high-frequency oscillation (HFO) content: ripples (80–200Hz) or fast-ripples (250–500Hz) predominate in LVF or HYP seizures, respectively. To establish whether these features change over the course of the disease, we recorded the EEG from the hippocampal CA3 subfield, subiculum, entorhinal cortex and dentate gyrus in two groups of pilocarpine-treated rats: the “early stage group” (n=8) was analyzed from day 3 to 20 post-SE while the “late stage group” (n=7) was studied from day 27 to 53 post-SE. We found that: (i) HYP and LVF seizures prevail in the early and late stage group, respectively; (ii) HYP seizures mainly originate from CA3 in the early stage group only; (iii) LVF seizures in both early and late stage group originate from a diffuse network; (iv) LVF and HYP seizures in the early stage group are mainly associated with ripples and fast ripples, respectively; but (v) fast ripples predominate in the late stage group, regardless of seizure onset pattern. Finally, extensive neuronal loss occurred in the hippocampus of the late stage group. Our results reveal that significant changes in ictogenesis and HFO occurrence, which are associated with the manifestation of severe hippocampal damage, occur over time in this MTLE model.

Circadian clustering of spontaneous epileptic seizures emerges after pilocarpine-induced status epilepticus.

Seizures in mesial temporal lobe epilepsy (MTLE) associated with hippocampal sclerosis are thought to develop with various latency intervals after an initial transient brain insult. To study seizure dynamics after an initial transient precipitating insult in a systematic fashion, we utilized continuous video-electroencephalography (EEG) monitoring after the induction of status epilepticus (SE) in a mouse MTLE model. Continuous 24/7 video/telemetric hippocampal EEG recordings in the systemic pilocarpine MTLE mouse model. After SE, we observed emerging seizures interfering with the circadian EEG rhythms. The physiologic circadian EEG pattern of mice was transiently suppressed for 2.9 (mean) ± (SEM) 0.5 days after SE. This period was accompanied predominately by nonconvulsive seizure activity, followed by convulsive seizures at later stages. After the circadian rhythm was restored, spontaneous generalized seizures occurred mainly in a clustered manner in a narrow time window between 4 and 7 p.m. (light cycle 7 a.m./7 p.m.). Moreover, we demonstrate that depth-electrode implantation surgery transiently disturbs the physiologic EEG circadian cycle; variation of the time point of SE induction after electrode insertion surgery revealed a substantial impact on the epilepsy phenotype, which was more severe when SE occurred after postsurgical reappearance of EEG circadian cycling. These data have several experimental and pathophysiologic implications. The impact of depth-electrode surgery on the phenotype has to be tightly controlled. In mice monitored after pilocarpine-induced SE, the "epileptogenesis" period is characterized by the dynamics of epileptiform activity toward behavioral recurrent seizure patterns. The striking clustering of spontaneous seizures at the transition from sleep to activity stages of mice has to be taken into account for future studies on the model. Improving our understanding of the molecular mechanisms that determine the circadian dynamics of seizure threshold remains an intriguing task for the future.

Interleukin-1β Plays a Pivotal Role via the PI3K/Akt/mTOR Signaling Pathway in the Chronicity of Mesial Temporal Lobe Epilepsy

Objective: Mesial temporal lobe epilepsy (MTLE) is the most common type of refractory epilepsy. It is often associated with hippocampal sclerosis, which is histopathologically characterized by selective neuron loss, mossy fiber sprouting, and synapse reconstruction, and is the primary cause of refractory epilepsy. Its mechanism has not been fully elucidated. Substantial evidence now supports that inflammatory pathways are activated in epilepsy foci. We have confirmed that the interleukin-1β (IL-1β) level is involved in the epileptogenesis of MTLE, and we further investigated how it works in its chronicity in this study. Methods: The MTLE model was induced by pilocarpine, and Western blot and co-immunoprecipitation were used to detect proteins related to the PI3K/Akt/mammalian target of rapamycin (mTOR) signaling pathway in the hippocampi of MTLE rats and MTLE children. Meanwhile, primary hippocampal neurons were cultured and transfected by lentivirus, and the same methods were used to test the related protein expression; fluorescent dye FM4-64 was used to measure synaptic vesicle endocytosis (SVE) of neurons. Results: We revealed that mTOR is continuously activated in the rat MTLE model and children with MTLE, and it correlated with the IL-1β level. We further proved that IL-1β activates neurons via the PI3K/Akt/mTOR signaling pathway, accompanied by the upregulation of MAP2 and the enhancement of SVE in hippocampal neurons. Conclusion: Our findings suggest that IL-1β can activate mTOR, followed by activated neurons, which is critical in the pathogenesis of MTLE chronicity. These findings contribute to the understanding of the pathogenesis of MTLE, and targeting inflammation modulators in MTLE may provide new pathways for therapy of refractory epilepsy.

Network evolution in mesial temporal lobe epilepsy revealed by diffusion tensor imaging.

The objective of the present study is to identify novel, time-indexed imaging biomarkers of epileptogenesis in mesial temporal lobe epilepsy (MTLE). We used high-resolution brain diffusion tensor imaging (DTI) of the translationally relevant methionine sulfoximine (MSO) brain infusion model of MTLE. MSO inhibits astroglial glutamine synthetase, which is deficient in the epileptogenic hippocampal formation of patients with MTLE. MSO-infused (epileptogenic) rats were compared with phosphate-buffered saline (PBS)-infused (nonepileptogenic) rats at early (3-4days) and late (6-9weeks) time points during epileptogenesis. The epileptogenic rats exhibited significant changes in DTI-measured fractional anisotropy (FA) in numerous brain regions versus nonepileptogenic rats. Changes included decreases and increases in FA in regions such as the entorhinal-hippocampal area, amygdala, corpus callosum, thalamus, striatum, accumbens, and neocortex. The FA changes evolved over time as animals transitioned from early to late epileptogenesis. For example, some areas with significant decreases in FA early in epileptogenesis changed to significant increases late in epileptogenesis. Finally, the FA changes significantly correlated with the seizure load. Our results suggest (1) that high-resolution DTI can be used for early identification and tracking of the epileptogenic process in MTLE, and (2) that the process identified by DTI is present in multiple brain areas, even though infusion of MSO is restricted to the unilateral entorhinal-hippocampal region.

The effect of IL-1β on synaptophysin expression and electrophysiology of hippocampal neurons through the PI3K/Akt/mTOR signaling pathway in a rat model of mesial temporal lobe epilepsy

Background: The inflammation induced by interleukin-1β (IL-1β) is a critical factor in the pathogenesis of mesial temporal lobe epilepsy (MTLE). Synaptophysin (SYN) and other changes, including neuron electrophysiology, participate in the pathophysiological processes of MTLE. Phosphatidylinositol 3-kinase (PI3K)/Akt/ mammalian target of rapamycin (mTOR) signaling pathway may play a critical role in regulating SYN expression and electrophysiology of hippocampal neurons.Methods: We used lithium-pilocarpine-treated rats as model of human MTLE, detecting epileptic seizures with digital video-EEG, and evaluating the proteins related to the PI3K/Akt/mTOR signaling pathway by western blot (WB). Then, we cultured primary neuron and established a neuronal epilepsy model using Mg2+-free media. Immunocytochemistry and WB were used to investigate SYN expression, and whole-cell current clamp recording techniques were used to detect the electrophysiological properties of cultured neurons.Results: We have demonstrated that IL-1β can activate the PI3K/Akt/mTOR signaling pathway in primary hippocampal neurons, and we speculate that IL-1β may affect SYN expression and neuron electrophysiology through PI3K/Akt/mTOR signaling pathway.Conclusion: We confirmed that IL-1β stimulated SYN expression and epileptiform discharges, and that blocking the PI3K/Akt/mTOR pathway alleviated these phenomena. Therefore, activation of the PI3K/Akt/mTOR signaling pathway by IL-1β contributes to the pathogenesis of MTLE, and modulating this pathway is a promising strategy of study for therapies to prevent or reverse the cellular and molecular mechanisms of epileptogenesis in MTLE.

Concept cells

Mesial temporal lobe epilepsy (MTLE) is the most common type of focal refractory epilepsy and is characterized by recurring seizures that are often refractory to medication. Since parvalbumin-positive (PV) interneurons were recently shown to play significant roles in ictogenesis, we established here how bilateral optogenetic stimulation of these interneurons in the hippocampus CA3 regions modulates seizures, interictal spikes and high-frequency oscillations (HFOs; ripples: 80–200 Hz, fast ripples: 250–500 Hz) in the pilocarpine model of MTLE. Bilateral optogenetic stimulation of CA3 PV-positive interneurons at 8 Hz (lasting 30 s, every 2 min) was implemented in PV-ChR2 mice for 8 consecutive days starting on day 7 (n = 8) or on day 13 (n = 6) after pilocarpine-induced status epilepticus (SE). Seizure occurrence was higher in both day 7 and day 13 groups of PV-ChR2 mice during periods of optogenetic stimulation (“ON”), compared to when stimulation was not performed (“OFF”) (day 7 group = p < 0.01, day 13 group = p < 0.01). In the PV-ChR2 day 13 group, rates of seizures (p < 0.05), of interictal spikes associated with fast ripples (p < 0.01), and of isolated fast ripples (p < 0.01) during optogenetic stimulations were significantly higher than in the PV-ChR2 day 7 group. Our findings reveal that bilateral activation of PV-interneurons in the hippocampus (leading to a presumptive increase in GABA signaling) favors ictogenesis. These effects may also mirror the neuropathological changes that occur over time after SE in this animal model.