Epileptic Animals Research Articles

Long-Term Effects of Anterior Thalamic Nucleus Deep Brain Stimulation on Spatial Learning in the Pilocarpine Model of Temporal Lobe Epilepsy

Cognitive impairment is a significant comorbidity of temporal lobe epilepsy that is associated with extensive hippocampal cell loss. Deep brain stimulation (DBS) of the anterior thalamic nucleus (ANT) has been used for the treatment of refractory partial seizures. In the pilocarpine model of epilepsy, ANT DBS applied during status epilepticus (SE) reduces hippocampal inflammation and apoptosis. When given to chronic epileptic animals it reduces hippocampal excitability and seizure frequency. Here, we tested whether ANT DBS delivered during SE and the silent phase of the pilocarpine model would reduce cognitive impairment when animals became chronically epileptic. SE was induced by a systemic pilocarpine injection (320 mg/kg). Immediately after SE onset, rats were assigned to receive DBS during the first six hours of SE (n = 8; DBSa group) or during SE + the silent period (i.e., 6 h/day until the animals developed the first spontaneous recurrent seizure; n = 10; DBSs group). Four months following SE, animals underwent water maze testing and histological evaluation. Nonstimulated chronic epileptic animals (n = 13; PCTL group) and age-matched naïve rats (n = 11, CTL group) were used as controls. Results were analyzed by repeated-measures analyses of variance (RM_ANOVA) and one-way ANOVAs, followed by Newman-Keuls post hoc tests. Although all groups learned the spatial task, epileptic animals with or without DBS spent significantly less time in the platform quadrant, denoting a spatial memory deficit (p < 0.02). Despite these negative behavioral results, we found that animals given DBS had a significantly higher number of cells in the CA1 region and dentate gyrus. Mossy fiber sprouting was similar among all epileptic groups. Despite lesser hippocampal neuronal loss, ANT DBS delivered either during SE or during SE and the silent phase of the pilocarpine model did not mitigate memory deficits in chronic epileptic rats.

МОЛЕКУЛЯРНЫЕ МЕХАНИЗМЫ ДИФФЕРЕНЦИРОВКИ НЕЙРОНАЛЬНЫХ СТВОЛОВЫХ КЛЕТОК КРЫС ЛИНИИ КРУШИНСКОГО-МОЛОДКИНОЙ

It is well known that epileptogenesis is accompanied with the aberrations in the hippocampal neurogenesis in human as well as in epileptic animals. The Krushinsky--Molodkina (KM) rats genetically prone to audiogenic seizure were recruited in the experiments. Neuronal stem cells (NSC) isolated from the hippocampus of KM rats at the 14--17 days of postnatal development, which corresponds the final stage for the development of the hippocampal neurogenic niche. NSC isolated from the hippocampus of Wistar rats were used as a control. The cells were incubated for 10 days in a medium supplemented with retinoic acid to stimulate neural differentiation. Previously we demonstrated that proliferation level of NSC KM was significantly lower in comparison with the NSC of Wistar rats, but neuronal differentiation rate of NSC KM was significantly higher. This research unravels the underlying molecular mechanisms. Analysis of Akt/GSK3b pathway demonstrated decreased expression of Akt that correlated with attenuation of phosphorylation level of GSK3b at Ser9 and decreased level of b-catenin. It is known that transcriptional factor CREB is one of the main substrates for Akt and on the other hand CREB tightly participates in the regulation of the cell proliferation. Obtained data revealed decreased activity of CREB in NSC KM in comparison to NSC Wistar that could mediate inhibition of cell cycle and potentiation the differentiation. Thus our results demonstrated that glutamatergic differentiation of NSC KM is mediated by AKT/GSK3b/b-catenin/CREB signaling pathway.

Anticonvulsant effect of Bacopa monnieri extracts on Catecholamine metabolism during PTZ-induced Epilepsy in different brain regions of Albino Rat

In the modern era, Epilepsy is one of the most prevalent serious neurological disorders, affecting 0.5 to 1.0% of the world’s population. Epileptic seizures are paroxysmal clinical events arising from neuronal hyper-excitability and hypersynchrony of the cerebral cortex, either locally (partial seizures) or diffusely in both hemispheres (generalized seizures). The agitated neuronal activity that occurs during a seizure is caused by a sudden imbalance between the inhibitory and excitatory signals in the brain with glutamate, ?-aminobutyric acid (GABA), noradrenaline, serotonin, and dopamine. Since long term usage of antiepileptic drugs cause high incidence of pharmacoresistence and untoward side effects, attention has been paid in recent years to screen bioactive compounds from natural medicinal plants for treatment of several neurological disorders including Epilepsy. Keeping in view of relative importance of natural medicinal plants, the present study is mainly focused to characterize the anti-convulsant effect of Bacopa monnieri (BM), an Indian herb which is being extensively used in Ayurvedic treatments related to neurological complications. The present study was designed to assess the neurotoxicity of PTZ on the metabolism of Biogenic amines (Nor-epinephrine, Epinephrine, Dopamine, Serotonin) in different brain regions (Cerebral cortex, Cerebellum, Pons medulla and Hippocampus) of rat and explore the possible antiepileptic effect of different extracts (Ethanol, n-Hexane, Chloroform, Ethyl acetate, n-Butanol and Aqueous extracts) of BM in comparison with Diazepam (DZ) (Reference control). PTZ-Induced epilepsy caused significant decrease in all the catecholamine levels in all brain regions. On par with Diazepam pretreatment, different extracts of BM except aqueous extract (AE) significantly increased the monoamine contents in all the regions of brain. Mono amine oxidase (MAO) activity levels showed elevation in different brain regions of rat during PTZ-induced epilepsy which was reversed in epileptic animals pretreated with different extracts except AE and DZ. These findings suggest that BM causes perceptible changes in the biogenic amine metabolism as one of the major facets of its anticonvulsant activity. Hence, BM extract can be effectively used in the treatment of Epilepsy and other related neurological disorders.

Effect of Co-administration of Bumetanide and Phenobarbital on Seizure Attacks in Temporal Lobe Epilepsy.

Introduction:The resistance of temporal lobe epilepsy to classic drugs is thought to be due to disruption in the excitation/inhibition of this pathway. Two chloride transporters, NKCC1 and KCC2, are expressed differently for the excitatory state of Gamma-Amino Butyric Acid (GABA). The present study explored the effect of bumetanide as a selective NKCC1 inhibitor either alone or in combination with the phenobarbital in the pilocarpine model of epilepsy.Methods:An animal model of Status Epilepticus (SE) was induced with pilocarpine in Wistar male rats followed by phenobarbital and or bumetanide or saline administration for 45 days after the induction of SE by Intraperitoneal (IP) injection. The rats were monitored, their behavior was recorded, and after 24 hours they were sacrificed to study the expression of NKCC1 and KCC2 using real time PCR.Results:The data showed that the effects of a combination of bumetanide with phenobarbital on frequency rate and duration of seizure attack were more than those of the phenobarbital alone. In addition, in the bumetanide and combined treatment groups, NKCC1 expression decreased significantly, compared with untreated epileptic animals. A delayed decrement in NKCC1/KCC2 expression ratio after bumetanide application was also observed.Conclusion:The combination of bumetanide with phenobarbital increases the inhibition of SE and maximizes the potential of GABA signaling pathway, and can be considered as an effective therapeutic strategy in patients with epilepsy.

Epigenetic Histone Deacetylation Inhibition Prevents the Development and Persistence of Temporal Lobe Epilepsy.

Epilepsy is a chronic brain disease characterized by repeated unprovoked seizures. Currently, no drug therapy exists for curing epilepsy or disease modification in people at risk. Despite several emerging mechanisms, there have been few studies of epigenetic signaling in epileptogenesis, the process whereby a normal brain becomes progressively epileptic because of precipitating factors. Here, we report a novel role of histone deacetylation as a critical epigenetic mechanism in epileptogenesis. Experiments were conducted using the histone deacetylase (HDAC) inhibitor sodium butyrate in the hippocampus kindling model of temporal lobe epilepsy (TLE), a classic model heavily used to approve drugs for treatment of epilepsy. Daily treatment with butyrate significantly inhibited HDAC activity and retarded the development of limbic epileptogenesis without affecting after-discharge signal. HDAC inhibition markedly impaired the persistence of seizure expression many weeks after epilepsy development. Moreover, subchronic HDAC inhibition for 2 weeks resulted in a striking retardation of epileptogenesis. HDAC inhibition, unexpectedly, also showed erasure of the epileptogenic state in epileptic animals. Finally, butyrate-treated animals exhibited a powerful reduction in mossy fiber sprouting, a morphologic index of epileptogenesis. Together these results underscore that HDAC inhibition prevents the development of TLE, indicating HDAC's critical signaling role in epileptogenesis. These findings, therefore, envisage a unique novel therapy for preventing or curing epilepsy by targeting the epigenetic HDAC pathway.

Effect of eugenol on lithium-pilocarpine model of epilepsy: behavioral, histological, and molecular changes.

Objective(s):Epilepsy establishment gives rise to biochemical and morphological changes in the hippocampus. Oxidative stress, morphological changes, and mossy fiber sprouting (MFS) in the hippocampus underpin the epilepsy establishment. Eugenol is the main component of the essential oil extracted from cloves with the potential to modulate neuronal excitability. Therefore, we investigated the effect of eugenol on convulsive behavior, oxidative stress, and histological changes of the hippocampus in lithium-pilocarpine model of epilepsy.Materials and Methods:Male Wistar rats weighing 220–250 g were divided into 4 groups; Control, Pilocarpine, Eugenol-Pilocarpine, and Eugenol. Oxidative stress markers were assayed by a biochemical method. Nissl and Timm staining were used to show neuronal survival and MFS, respectively. Behavioral convulsions were evaluated using the modified Racine scale.Results:Eugenol decreased seizure stage and duration as well as mortality. Neuronal numbers were preserved by eugenol treatment in epileptic animals, while eugenol alone reduced the number by itself in all hippocampal sub-regions including DG, CA3, and CA1. Furthermore, eugenol alone increased MDA, GPx and SOD markers, while it increased MDA not only in combined treatment with pilocarpine but also in pilocarpine-treated animals. In contrast to MFS enhancement in naïve animals, eugenol partially reversed the MFS enhancement induced by pilocarpine.Conclusion:Eugenol could prevent behavioral convulsions and show neuroprotective effects through increasing neuronal survival probably by decreasing MFS and increasing the GPx antioxidant marker.

Relative Safety of Different Antidepressants for Treatment of Depression in Chronic Epileptic Animals Associated with Depression.

Background and PurposeDepression is one of the major psychiatric comorbidities associated with epilepsy. The inconclusive results of antidepressants (ADs) regarding their safety in regard to convulsions have strongly contributed towards under treatment of depression in people with epilepsy (PWE). Thus, the present study was envisaged to assess the relative safety of four different classes of ADs regarding their convulsive potential in kindled/epileptic animals.MethodsKindling (an animal model to induce chronic epilepsy) was induced in male Swiss albino mice by administration of pentylenetetrazole subconvulsive doses (35 mg/kg, i.p.) at an interval of 48 ± 2 h for 42 days. The epileptic animals were treated with saline; imipramine (20 mg/kg/day i.p.); fluoxetine (20 mg/kg/day i.p.); venlafaxine (10 mg/kg/day i.p.) and mirtazapine (10 mg/kg/day i.p.) for 15 days. Except naive, animals were challenged with pentylenetetrazole subconvulsive dose (35 mg/kg, i.p.) on every 5th day to determine convulsion severity score, latency to first myoclonic jerk, latency to first tonic-clonic convulsions and numbers of tonic-clonic convulsions. Depression was also evaluated every 5th day employing tail suspension test 2 h after pentylenetetrazole subconvulsive dose.ResultsAll ADs have been reported significant antidepressant potential however regarding their safety in regard to convulsions in epileptic animals, variable results are obtained. Chronic administration of venlafaxine and mirtazapine were found to have significant anticonvulsant effect in epileptic animals. The behavioral data was further corroborated by neurochemical findings.ConclusionsThe treatment with venlafaxine and mirtazapine can be considered safe for treatment of depression in epilepsy and may enhance anticonvulsant potential of antiepileptic drugs as an adjuvant therapy. However, pharmacokinetic studies are warranted before translating these findings in PWE.

Ferulic Acid Supplementation for Management of Depression in Epilepsy

Neuroinflammation driven altered neurochemical milieu have been reported to play a significant role in pathogenesis of comorbid depression in epilepsy. Most of the antiepileptic drugs (AEDs) such as levetiracetam, taigabine, topiramate have not been reported any significant effect in alleviating neuroinflammation, which may explain their ineffectiveness in ameliorating depression associated with epilepsy. The supplementation of antidepressants (ADs) attracts various pharmacokinetic and pharmacodynamic interactions with AEDs and was considered unsafe in epilepsy. This scenario pushes us to search therapies beyond ADs by critically exploring the disease mechanism. Thus, as suggested by our previous findings, anti-inflammatory phytotherapy (Ferulic acid) appears a promising adjuvant therapy with levetiracetam for effective and safe management of depression associated with epilepsy. Pentylenetetrazole kindling induced epileptic animals were treated with vehicle, levetiracetam (40mg/kg/day i.p.) and levetiracetam in combination with two doses of ferulic acid (40, 80mg/kg)/day/p.o. for 15 days. Every 5th day during the treatment, depression was evaluated and animals were administered pentylenetetrazole to evaluate the effect of different pharmacological interventions on seizure severity. The epileptic animals were reported decreased seizure threshold associated with comorbid depression. The treatment with levetiracetam was found ineffective in ameliorating the associated depression. However ferulic acid supplementation with levetiracetam ameliorated comorbid depression supported with restored circulating corticosterone levels, decreased proinflammatory cytokines (IL-1β, TNF-α) and indoleamine 2,3-dioxygenase activity in mice brain. Thus, suggesting supplementation of anti-inflammatory phytomolecules such as ferulic acid as safe and effective adjuvant therapy for the management of comorbid depression in epilepsy.

Agmatine for combined treatment of epilepsy, depression and cognitive impairment in chronic epileptic animals

Epilepsy is fourth most common neurological disorders associated with depression and cognitive deficits. As per present scenario, none of the antiseizure drugs have been reported successful to have ameliorative effect on epilepsy associated depression and cognitive deficits. Thus, the study was envisioned to assess an ameliorative potential of agmatine on epilepsy and its efficacy and safety for management of associated depression and cognitive deficits. The animals were made epileptic employing pentylenetetrazole (35mg/kg i.p. every 48±2h) kindling model of epilepsy and subsequently were treated with vehicle, valproic acid (300mg/kg/day i.p.) and agmatine (2.5, 5, and 10mg/kg)/day/i.p. for 15days. Except naïve, all the groups were challenged with same pentylenetetrazole dose as employed during kindling on days 5, 10, and 15 to evaluate seizure severity. Two hours after seizure severity test, tail suspension test and passive shock avoidance paradigm was employed to evaluate depression and cognitive behavior respectively. Results suggested that epileptic animals were significantly associated with depression and cognitive impairment. Chronic valproate treatment significantly reduced seizure severity, but was found unable to mitigate depression and cognitive deficits. However, agmatine treatment dose dependently ameliorated seizure severity as well as associated depression and cognitive deficits. On 15th day, animals were euthanized and pertinent neurochemical estimations were carried out in cortical and hippocampal areas of the mice brain. Thus, study concluded that agmatine ameliorated seizure severity, depression and cognitive impairment in epileptic animals, possibly via restoring glutamate-GABA neurotransmission and serotonin synthesis with decreased nitrosative stress.

Association of RASgrf1 methylation with epileptic seizures.

DNA methylation, one of the mechanisms of epigenetic regulation, has been suggested to be related with epilepsy. RASgrf1 is a paternally imprinted gene and has a differentially methylated region (DMR) at the promoter that can silence gene expression. We have previously observed the down-regulation of RASgrf1 in the temporal neocortex of epilepsy patients and in the hippocampus of epileptic animals. Here, we further explored the dynamic change (1-day acute period, 10-day latent period and 45-day chronic phase) of DNA methylation and RASgrf1 expression after acute epileptic seizures in kainic acid (KA)-treated mice, and we observed the impact of N-phthalyl-L-tryptophan (RG108), a DNA methyltransferase (DNMT) inhibitor, on an acute epileptic model by polymerase chain reaction (PCR), western blotting, and bisulfite sequencing PCR (BSP). The results directly showed that the methylation of the RASgrf1 promoter gradually increased and reached a maximal level at the latent period, with subsequent suppression of RASgrf1 mRNA and protein expression levels, which reached a minimum level in the chronic phase. RG108 inhibited the increased methylation of the RASgrf1 gene, with significant inhibition occurring at the latent period, and restored RASgrf1 expression levels in the chronic phase. In addition, we demonstrated that RG108 could suppress acute epileptic seizures in KA-treated mice and epileptic discharges in 4-aminopyridine (4-AP)-treated hippocampal slices. These findings demonstrate that RASgrf1 is closely associated with epilepsy via the aberrant methylation of RASgrf1, and regulating the methylation status of relevant genes might be an intriguing topic in future research on epilepsy.

Imaging blood-brain barrier dysfunction as a biomarker for epileptogenesis.

A biomarker that will enable the identification of patients at high-risk for developing post-injury epilepsy is critically required. Microvascular pathology and related blood-brain barrier dysfunction and neuroinflammation were shown to be associated with epileptogenesis after injury. Here we used prospective, longitudinal magnetic resonance imaging to quantitatively follow blood-brain barrier pathology in rats following status epilepticus, late electrocorticography to identify epileptic animals and post-mortem immunohistochemistry to confirm blood-brain barrier dysfunction and neuroinflammation. Finally, to test the pharmacodynamic relevance of the proposed biomarker, two anti-epileptogenic interventions were used; isoflurane anaesthesia and losartan. Our results show that early blood-brain barrier pathology in the piriform network is a sensitive and specific predictor (area under the curve of 0.96, P < 0.0001) for epilepsy, while diffused pathology is associated with a lower risk. Early treatments with either isoflurane anaesthesia or losartan prevented early microvascular damage and late epilepsy. We suggest quantitative assessment of blood-brain barrier pathology as a clinically relevant predictive, diagnostic and pharmaco!dynamics biomarker for acquired epilepsy.

Ginkgo biloba L. attenuates spontaneous recurrent seizures and associated neurological conditions in lithium-pilocarpine rat model of temporal lobe epilepsy through inhibition of mammalian target of rapamycin pathway hyperactivation

Ethnopharmacological relevanceGinkgo biloba L. (Ginkgoaceae) has been widely used in traditional medicine for variety of neurological conditions particularly behavioral and memory impairments. Aim of the studyThe present study was envisaged to explore the effect of a standardized fraction of Ginkgo biloba leaves (GBbf) in rat model of lithium-pilocarpine induced spontaneous recurrent seizures, and associated behavioral impairments and cognitive deficit. Materials and methodsRats showing appearance of spontaneous recurrent seizures following lithium pilocarpine (LiPc)-induced status epilepticus (SE) were treated with different doses of GBbf or vehicle for subsequent 4 weeks. The severity of seizures and aggression in rats were scored following treatment with GBbf. Further, open field, forced swim, novel object recognition and Morris water maze tests were conducted. Histopathological, protein levels and gene expression studies were performed in the isolated brains. ResultsTreatment with GBbf reduced seizure severity score and aggression in epileptic animals. Improved spatial cognitive functions and recognition memory, along with reduction in anxiety-like behavior were also observed in the treated animals. Histopathological examination by Nissl staining showed reduction in neuronal damage in the hippocampal pyramidal layer. The dentate gyrus and Cornu Ammonis 3 regions of the hippocampus showed reduction in mossy fiber sprouting. GBbf treatment attenuated ribosomal S6 and pS6 proteins, and hippocampal mTOR, Rps6 and Rps6kb1 mRNA levels. ConclusionsThe results of present study concluded that GBbf treatment suppressed lithium-pilocarpine induced spontaneous recurrent seizures severity and incidence with improved cognitive functions, reduced anxiety-like behavior and aggression. The effect was found to be due to inhibition of mTOR pathway hyperactivation linked with recurrent seizures.

Physical Exercise Restores the Generation of Newborn Neurons in an Animal Model of Chronic Epilepsy.

Neurogenesis impairment is associated with the chronic phase of the epilepsy in humans and also observed in animal models. Recent studies with animal models have shown that physical exercise is capable of improving neurogenesis in adult subjects, alleviating cognitive impairment and depression. Here, we show that there is a reduction in the generation of newborn granule cells in the dentate gyrus of adult rats subjected to a chronic model of epilepsy during the postnatal period of brain development. We also show that the physical exercise was capable to restore the number of newborn granule cells in this animals to the level observed in the control group. Notably, a larger number of newborn granule cells exhibiting morphological characteristics indicative of correct targeting into the hippocampal circuitry and the absence of basal dendrite projections was also observed in the epileptic animals subjected to physical exercise compared to the epileptic animals. The results described here could represent a positive interference of the physical exercise on the neurogenesis process in subjects with chronic epilepsy. The results may also help to reinterpret the benefits of the physical exercise in alleviating symptoms of depression and cognitive dysfunction.

Antiepileptic action of c-Jun N-terminal kinase (JNK) inhibition in an animal model of temporal lobe epilepsy

Several phosphorylation signaling pathways have been implicated in the pathogenesis of epilepsy arising from both genetic causes and acquired insults to the brain. Identification of dysfunctional signaling pathways in epilepsy may provide novel targets for antiepileptic therapies. We previously described a deficit in phosphorylation signaling mediated by p38 mitogen-activated protein kinase (p38 MAPK) that occurs in an animal model of temporal lobe epilepsy, and that produces neuronal hyperexcitability measured in vitro. We asked whether in vivo pharmacological manipulation of p38 MAPK activity would influence seizure frequency in chronically epileptic animals. Administration of a p38 MAPK inhibitor, SB203580, markedly worsened spontaneous seizure frequency, consistent with prior in vitro results. However, anisomycin, a non-specific p38 MAPK activator, significantly increased seizure frequency. We hypothesized that this unexpected result was due to activation of a related MAPK, c-Jun N-terminal kinase (JNK). Administration of JNK inhibitor SP600125 significantly decreased seizure frequency in a dose-dependent manner without causing overt behavioral abnormalities. Biochemical analysis showed increased JNK expression and activity in untreated epileptic animals. These results show for the first time that JNK is hyperactivated in an animal model of epilepsy, and that phosphorylation signaling mediated by JNK may represent a novel antiepileptic target.

Neuron subset-specific Pten deletion induces abnormal skeletal activity in mice

Individuals with a history of epilepsy are at higher risk for bone fractures compared to the general population. Although clinical studies support an association between low bone mineral density (BMD) and anti-seizure medications, little is known on whether a history of seizures is linked to altered bone health. Therefore, in this study we tested the hypothesis that bone mass, morphology, and bone mineralization are altered by seizures in genetically epileptic animals and in animals subjected to an episode of status epilepticus. In this study, we used NS-Pten conditional knockout mice (a well-studied genetic model of epilepsy). We used microCT analysis to measure BMD, morphology, and mineralization in NS-Pten+/+ (wildtype) and NS-Pten−/− (knockout) mice at 4 and 8weeks, as well as adult Kv4.2+/+ and Kv4.2−/− mice. We measured BMD, bone morphology, and mineralization in adult NS-Pten+/+ mice that received status epilepticus through kainic acid (20mg/kg intraperitoneal). Further, we measured locomotion for NS-Pten+/+ and NS-Pten−/− mice at 4 and 6weeks. We found that NS-Pten−/− mice exhibited low BMD in the tibial metaphysis and midshaft compared to non-epileptic mice. Morphologically, NS-Pten−/− mice exhibited decreased trabecular volume fraction, and endocortical expansion in both the metaphyeal and diaphyseal compartments. In the midshaft, NS-Pten−/− mice exhibited reduced tissue mineral density, indicating impaired mineralization in addition to morphological deficits. NS-Pten−/− mice exhibited hyperactivity in open field testing, suggesting low bone mass in NS-Pten−/− mice was not attributable to hypoactivity. Differences in BMD were not observed following kainate-induced seizures or in the Kv4.2−/− model of seizure susceptibility. Our findings suggest that deletion of Pten in the brain results in impaired bone mass and mineralization, which may contribute to weaker bones and thereby a higher fracture risk.

Massively augmented hippocampal dentate granule cell activation accompanies epilepsy development

In a mouse model of temporal lobe epilepsy, multicellular calcium imaging revealed that disease emergence was accompanied by massive amplification in the normally sparse, afferent stimulation-induced activation of hippocampal dentate granule cells. Patch recordings demonstrated reductions in local inhibitory function within the dentate gyrus at time points where sparse activation was compromised. Mimicking changes in inhibitory synaptic function and transmembrane chloride regulation was sufficient to elicit the dentate gyrus circuit collapse evident during epilepsy development. Pharmacological blockade of outward chloride transport had no effect during epilepsy development, and significantly increased granule cell activation in both control and chronically epileptic animals. This apparent occlusion effect implicates reduction in chloride extrusion as a mechanism contributing to granule cell hyperactivation specifically during early epilepsy development. Glutamine plays a significant role in local synthesis of GABA in synapses. In epileptic mice, sparse granule cell activation could be restored by glutamine application, implicating compromised GABA synthesis. Glutamine had no effect on granule cell activation earlier, during epilepsy development. We conclude that compromised feedforward inhibition within the local circuit generates the massive dentate gyrus circuit hyperactivation evident in animals during and following epilepsy development. However, the mechanisms underlying this disinhibition diverge significantly as epilepsy progresses.

Role of proinflammatory cytokines in the inhibition of hippocampal neurogenesis in mesial temporal lobe epilepsy

BackgroundIn mesial temporal lobe epilepsy, reduced hippocampal neurogenesis correlates with severe impairment in declarative learning and memory. The proinflammatory cytokines interleukin 1β and high-mobility group box 1 (HMGB1) and their receptors have key roles in generating and perpetuating seizures. This study aimed to assess their potential role in inhibiting neurogenesis in patients with mesial temporal lobe epilepsy and whether any effect would be pharmacologically reversible as a possible mechanism to restore learning and memory deficits. MethodsWe generated three-dimensional (3D) stem-cell cultures (Hi-Spot, Capsant Neurotechnologies Ltd, UK), preserving the inflammatory microenvironment and key features of the in-vivo stem-cell niche, from sclerotic tissue obtained from patients undergoing surgery for epilepsy (n=23). Concentrations of interleukin 1β and HMGB1 and their receptors in sclerotic tissue were determined with PCR, western blot, ELISA, and immunohistochemistry and compared with normal cortical tissue. Neurogenesis in the presence or absence of interleukin 1β or HMGB1 antagonists was examined by quantifying the number of newly born neurons. FindingsWe expanded the use of air–liquid interphase technology previously used to generate Hi-Spots from the cortex of rats to generate 3D Hi-Spots from the adult human hippocampus. Using ELISA, we demonstrated that interleukin 1β concentrations were significantly elevated in sclerotic hippocampal Hi-Spots compared with control (mean 24·4 pg/mL [SD 2·3] vs 3·6 [1·4], p<0·01). Interleukin 1 receptor mRNA was four times higher than control with dense protein coexpression by nestine-positive neural stem cells. Blockade of interleukin 1 receptor increased the number of newly born neurons in sclerotic Hi-Spots (mean 14·23 cells per mm2 [SD 1·13] vs 4·8 [0·7], p<0·01) and significantly decreased HMGB1 activity (5·5 ng/mL [0·25] vs 7·7 [0·29], p<0·01). Treatment with the HMGB1 antagonist BoxA increased the percentage of newly born neurons and completely reversed the antineurogenic effects of interleukin 1β, indicating possible crosstalk between interleukin 1β and HMGB1 in modulating hippocampal neurogenesis. InterpretationWe provide a novel in-vitro molecular model that relies on the interleukin 1β–interleukin 1 receptor–HMGB1 axis. Whereas previous studies have elegantly shown that blockade of this axis renders epileptic animals free of seizures, our observations support the development of therapeutic interventions that counteract the microenvironment to promote hippocampal neurogenesis. This possibility could lead to pharmacological strategies to treat learning and memory deficits in mesial temporal lobe epilepsy. FundingEpilepsy Research UK.

Dentate gyrus autonomous ictal activity in the status epilepticus rat model of epilepsy

The dentate gyrus (DG) as part of the hippocampal formation is believed to serve as a gatekeeper with strong inhibitory properties against uncontrolled propagation of neuronal activity from the entorhinal cortex and neocortical structures. In temporal lobe epilepsy, the DG becomes hyperexcitable and loses its gate function, enabling propagation of ictal activity into downstream structures such as CA3 and CA1 areas. Furthermore, the DG, apart from facilitating propagation, may also be able to autonomously generate ictal activity, but this point has remained open so far. To tackle this question, we used intrinsic optical imaging in combination with electrophysiological recordings in brain slice preparations from rats in which status epilepticus had been induced electrically several weeks prior to measurements. Upon omission of Mg++ from the artificial cerebrospinal fluid, in 15 out of 33 slices (45.4%) from 9 out of 13 epileptic animals (69.2%), spontaneous and autonomous ictal activity, mostly seizure-like events (SLE), was observed in the DG. This activity manifested independently from SLE generated in adjacent cortices and never occurred in slices from control animals. SLE generated in the DG differed from those with origin in the entorhinal or temporal cortex by longer latency to the first event after Mg++ omission (p<0.001), a higher SLE frequency (p<0.05), higher amplitude (p<0.001) and a longer SLE duration (p<0.05). We conclude that in epilepsy, the DG, in addition to facilitated gating of activity from upstream structures, can serve as an independent generator of ictal activity.

Do oral contraceptives increase epileptic seizures?

ABSTRACTIntroduction: Hormonal contraceptives are used by over 100 million people worldwide. Recently, there has been an emerging interest in studying the potential impact of oral contraceptives (OCs) on certain neurological conditions. It has been suspected for some time that hormonal birth control increases seizure activity in women with epilepsy, but there is little supportive data.Areas covered: Literature from PubMed and online sources was analyzed with respect to hormonal contraception and epilepsy or seizures. New evidence indicates that OCs can cause an increase in seizures in women with epilepsy. The epilepsy birth control registry, which surveyed women with epilepsy, found that those using hormonal contraceptives self-reported 4.5 times more seizures than those that did not use such contraceptives. A preclinical study confirmed these outcomes wherein epileptic animals given ethinyl estradiol, the primary component of OCs, had more frequent seizures that are more likely to be resistant.Expert commentary: OC pills may increase seizures in women with epilepsy and such refractory seizures are more likely to cause neuronal damage in the brain. Thus, women of child bearing age with epilepsy should consider using non-hormonal forms of birth control to avoid risks from OC pills. Additional research into the mechanisms and prospective clinical investigation are needed.

MBD3 expression and DNA binding patterns are altered in a rat model of temporal lobe epilepsy.

The aim of the present study was to examine involvement of MBD3 (methyl-CpG-binding domain protein 3), a protein involved in reading DNA methylation patterns, in epileptogenesis and epilepsy. We used a well-characterized rat model of temporal lobe epilepsy that is triggered by status epilepticus, evoked by electrical stimulation of the amygdala. Stimulated and sham-operated animals were sacrificed 14 days after stimulation. We found that MBD3 transcript was present in neurons, oligodendrocytes, and astrocytes in both control and epileptic animals. We detected the nuclear localization of MBD3 protein in neurons, mature oligodendrocytes, and a subpopulation of astrocytes but not in microglia. Amygdala stimulation significantly increased the level of MBD3 immunofluorescence. Immunoprecipitation followed by mass spectrometry and Western blot revealed that MBD3 in the adult brain assembles the NuRD complex, which also contains MTA2, HDAC2, and GATAD2B. Using chromatin immunoprecipitation combined with deep sequencing, we observed differences in the occupancy of DNA regions by MBD3 protein between control and stimulated animals. This was not followed by subsequent changes in the mRNA expression levels of selected MBD3 targets. Our data demonstrate for the first time alterations in the MBD3 expression and DNA occupancy in the experimental model of epilepsy.