Kainic Acid Model Research Articles

Dibutyryl Cyclic Adenosine Monophosphate Rescues the Neurons From Degeneration in Stab Wound and Excitotoxic Injury Models.

Dibutyryl cyclic adenosine monophosphate (dBcAMP), a cell-permeable synthetic analog of cAMP, has been shown to induce astrogliosis in culture. However, the exact mechanism underlying how dBcAMP exerts its function in situ is not clear. The objective of this study was to examine the effects of dBcAMP on astrogliosis and survival of neurons in stab wound and kainic acid models of brain injury. Stab wound was done in cerebral cortex of BALB/c male mice. Kainic acid lesion was induced in hippocampus by injecting 1μl kainic acid into the lateral ventricle. Animals in both models of injury were divided into L+dBcAMP and L+PBS groups and treated with dBcAMP or PBS for 3, 5, and 7 days respectively. The brain sections were stained for Cresyl violet and Fluro jade-B to assess the degenerating neurons. Immunostaining for GFAP and Iba-1 was done for assessing the astrogliosis and microglial response respectively. Expression of GFAP and BDNF levels in the tissue were estimated by Western blotting and ELISA respectively. The results showed a gradual increase in the number of both astrocytes and microglia in both injuries with a significant increase in dBcAMP-treated groups. The number of degenerating neurons significantly decreased in dBcAMP treated groups. In addition, it was found that dBcAMP stimulated the expression of GFAP and BDNF in both stab wound and kainic acid injuries. Treatment with BDNF receptor inhibitor AZ-23, showed an increase in the degenerating neurons suggesting the role of BDNF in neuroprotection. This study indicates that dBcAMP protects neurons from degeneration by enhancing the production of BDNF and may be considered for use as therapeutic agent for treatment of brain injuries.

Altered mitochondrial acetylation profiles in a kainic acid model of temporal lobe epilepsy

Impaired bioenergetics and oxidative damage in the mitochondria are implicated in the etiology of temporal lobe epilepsy, and hyperacetylation of mitochondrial proteins has recently emerged as a critical negative regulator of mitochondrial functions. However, the roles of mitochondrial acetylation and activity of the primary mitochondrial deacetylase, SIRT3, have not been explored in acquired epilepsy. We investigated changes in mitochondrial acetylation and SIRT3 activity in the development of chronic epilepsy in the kainic acid rat model of TLE. Hippocampal measurements were made at 48 h, 1 week and 12 weeks corresponding to the acute, latent and chronic stages of epileptogenesis. Assessment of hippocampal bioenergetics demonstrated a ≥ 27% decrease in the ATP/ADP ratio at all phases of epileptogenesis (p < 0.05), whereas cellular NAD+ levels were decreased by ≥ 41% in the acute and latent time points (p < 0.05), but not in chronically epileptic rats. In spontaneously epileptic rats, we found decreased protein expression of SIRT3 and a 60% increase in global mitochondrial acetylation, as well as enhanced acetylation of the known SIRT3 substrates MnSOD, Ndufa9 of Complex I and IDH2 (all p < 0.05), suggesting SIRT3 dysfunction in chronic epilepsy. Mass spectrometry-based acetylomics investigation of hippocampal mitochondria demonstrated a 79% increase in unique acetylated proteins from rats in the chronic phase vs. controls. Pathway analysis identified numerous mitochondrial bioenergetic pathways affected by mitochondrial acetylation. These results suggest SIRT3 dysfunction and aberrant protein acetylation may contribute to mitochondrial dysfunction in chronic epilepsy.

Targeting of microRNA-21-5p protects against seizure damage in a kainic acid-induced status epilepticus model via PTEN-mTOR

ObjectiveStudies have shown that microRNAs play a role in the development of epilepsy by regulating downstream target messenger (m)RNA. The present study aims to determine the changes associated with microRNA-21-5p (miR-21-5p) during epileptogenesis in a kainic acid rat model, and to assess whether the PTEN-mTOR pathway is a target of miR-21-5p. MethodReverse transcription polymerase chain reaction (RT-PCR) was used to examine the quantitative expressions of miR-21-5p and PTEN, and Western blotting was used to test the activity of mTOR in the acute, latent, and chronic stages of epileptogenesis. The antagomir of miR-21-5p was injected into the intracerebroventricular space using a microsyringe. Neuronal death and epilepsy discharge were assessed by Nissl staining and electroencephalography (EEG), respectively. The Morris water maze (MWM) was used to assess the cognitive impairment in rats after status epilepticus (SE). ResultsBoth miR-21-5p and mTOR were upregulated and PTEN was downregulated in rats during acute, latent, and chronic stages of epileptogenesis when compared with those of the control. After using antagomir miR-21-5p in vivo, miR-21-5p and mTOR decreased and the expression of PTEN increased compared with that in the SE model. The silencing of miR-21-5p diminished the number of abnormal spikes on EEG and decreased the number of neuron deletions on Nissl staining. The cognitive and memory impairment caused by epilepsy could also be improved after miR-21-5p knockdown in vivo. ConclusionThe results of the present study demonstrate that PTEN-mTOR is the target of miR-21-5p in a kainic acid model of epilepsy. The knockout of miR-21-5p decreases the neuronal damage in stages of epileptogenesis. The miR-21-5p/PTEN/mTOR axis may be a potential target for preventing and treating seizures and epileptic damage.

Pharmacological characterization of the cannabinoid receptor 2 agonist, β-caryophyllene on seizure models in mice

PurposeActivation of CB1 receptors, produces anticonvulsant effect accompanied by memory disturbance both in animal seizure tests and in patients with epilepsy. Few reports considered the role of CB2 receptor on seizure susceptibility and cognitive functions. The aim of the present study was to explore the effect of a selective CB2 receptor agonist β-caryophyllene (BCP) in models of seizures and cognition in mice. MethodsDose-dependent effects of BCP was studied in maximal electroshock seizure (MES) test, subcutaneous pentylenetetrazole (scPTZ) test and Morris water maze test. Phenytoin and diazepam were used as reference drugs in seizure tests. The effect of sub-chronic treatment with BCP for 7 days (50 and 100 mg kg−1) was assessed on status epilepticus (SE) induced by kainic acid (KA) model and oxidative stress through measurement of malondialdehyde (MDA) level in the hippocampus. The acute neurotoxicity was determined by a rotarod test. ResultsThe BCP exerted a protection in the MES test at the lowest dose of 30 mg kg−1 at the 4-h interval tested comparable to that of the referent drug phenytoin. The CB2 agonist was ineffective in the scPTZ test. The BCP displayed no neurotoxicity in the rotarod test. The BCP decreased the seizure scores in the KA-induced SE, which effect correlated with a diminished lipid peroxidation. The CB2 agonist exerted a dose-dependent decrease of latency to cross the target area during the three days of testing in the Morris water maze test. ConclusionOur results suggest that the CB2 receptor agonists might be clinically useful as an adjunct treatment against seizure spread and status epilepticus and concomitant oxidative stress, neurotoxicity and cognitive impairments.

Peripherally derived T regulatory and γδ T cells have opposing roles in the pathogenesis of intractable pediatric epilepsy.

The pathophysiology of drug-resistant pediatric epilepsy is unknown. Flow cytometric analysis of inflammatory leukocytes in resected brain tissues from 29 pediatric patients with genetic (focal cortical dysplasia) or acquired (encephalomalacia) epilepsy demonstrated significant brain infiltration of blood-borne inflammatory myeloid cells and memory CD4+ and CD8+ T cells. Significantly, proinflammatory (IL-17- and GM-CSF-producing) γδ T cells were concentrated in epileptogenic lesions, and their numbers positively correlated with disease severity. Conversely, numbers of regulatory T (T reg) cells inversely correlated with disease severity. Correspondingly, using the kainic acid model of status epilepticus, we show ameliorated seizure activity in both γδ T cell- and IL-17RA-deficient mice and in recipients of T reg cells, whereas T reg cell depletion heightened seizure severity. Moreover, both IL-17 and GM-CSF induced neuronal hyperexcitability in brain slice cultures. These studies support a major pathological role for peripherally derived innate and adaptive proinflammatory immune responses in the pathogenesis of intractable epilepsy and suggest testing of immunomodulatory therapies.

Neurobiological Correlates of Alpha-Tocopherol Antiepileptogenic Effects and MicroRNA Expression Modulation in a Rat Model of Kainate-Induced Seizures

Seizure-triggered maladaptive neural plasticity and neuroinflammation occur during the latent period as a key underlying event in epilepsy chronicization. Previously, we showed that α-tocopherol (α-T) reduces hippocampal neuroglial activation and neurodegeneration in the rat model of kainic acid (KA)-induced status epilepticus (SE). These findings allowed us to postulate an antiepileptogenic potential for α-T in hippocampal excitotoxicity, in line with clinical evidence showing that α-T improves seizure control in drug-resistant patients. To explore neurobiological correlates of the α-T antiepileptogenic role, rats were injected with such vitamin during the latent period starting right after KA-induced SE, and the effects on circuitry excitability, neuroinflammation, neuronal death, and microRNA (miRNA) expression were investigated in the hippocampus. Results show that in α-T-treated epileptic rats, (1) the number of population spikes elicited by pyramidal neurons, as well as the latency to the onset of epileptiform-like network activity recover to control levels; (2) neuronal death is almost prevented; (3) down-regulation of claudin, a blood–brain barrier protein, is fully reversed; (4) neuroinflammation processes are quenched (as indicated by the decrease of TNF-α, IL-1β, GFAP, IBA-1, and increase of IL-6); (5) miR-146a, miR-124, and miR-126 expression is coherently modulated in hippocampus and serum by α-T. These findings support the potential of a timely intervention with α-T in clinical management of SE to reduce epileptogenesis, thus preventing chronic epilepsy development. In addition, we suggest that the analysis of miRNA levels in serum could provide clinicians with a tool to evaluate disease evolution and the efficacy of α-T therapy in SE.

Cilostazol attenuates kainic acid-induced hippocampal cell death.

Cilostazol is a selective inhibitor of type 3 phosphodiesterase (PDE3) and has been widely used as an antiplatelet agent. Cilostazol mediates this activity through effects on the cyclic adenosine monophosphate (cAMP) signaling cascade. Recently, it has attracted attention as a neuroprotective agent. However, little is known about cilostazol's effect on excitotoxicity induced neuronal cell death. Therefore, this study evaluated the neuroprotective effect of cilostazol treatment against hippocampal neuronal damage in a mouse model of kainic acid (KA)-induced neuronal loss. Cilostazol pretreatment reduced KA-induced seizure scores and hippocampal neuron death. In addition, cilostazol pretreatment increased cAMP response element-binding protein (CREB) phosphorylation and decreased neuroinflammation. These observations suggest that cilostazol may have beneficial therapeutic effects on seizure activity and other neurological diseases associated with excitotoxicity.

HEK-293 secretome attenuates kainic acid neurotoxicity through insulin like growth factor-phosphatidylinositol-3-kinases pathway and by temporal regulation of antioxidant defense machineries

A major impediment in the success of cell therapy for neurodegenerative diseases is the poor survival of grafted cells in the in vivo milieu, predominantly due to accumulated reactive oxygen species, thus prompting the search for suitable alternatives. Accumulating evidence suggests that the therapeutic potential of transplanted cells is partially attributed to the secretome released by them into the extracellular milieu. Studies that investigated the neuroprotective potential of the secretome attributes to the mere presence of growth factors without addressing other underlying cellular/molecular changes that occur upon post-secretome intervention like re-establishing the host cell’s free radical scavenging machineries. In the present study, we investigated the neuroprotective effects of human embryonic kidney (HEK-293) cell line derived secretome (HEK-S) in an in vitro model of kainic acid (KA) induced neurodegeneration and explored the possible neuroprotective mechanism(s) of HEK-S. Murine hippocampal cells were exposed to toxic doses of KA (200μM) for 6hours (H) or 24H to induce excitotoxicity. Kainic acid exposed hippocampal cells were then treated with HEK-S either simultaneously or 6h post-KA exposure. Our results revealed that HEK-S confers significant neuroprotection in early/later stages of neurodegeneration through insulin like growth factor (IGF) – phosphatidylinositol-3-kinases (PI3K) pathway, efficiently restoring the host’s free radical scavenging mechanisms at molecular-cellular-biochemical levels and also by modulating kainate receptor subunit expressions in host neurons.

Serotonin depletion increases seizure susceptibility and worsens neuropathological outcomes in kainate model of epilepsy.

Serotonin is implicated in the regulation of seizures, but whether or not it can potentiate the effects of epileptogenic factors is not fully established. Using the kainic acid model of epilepsy in rats, we tested the effects of serotonin depletion on (1) susceptibility to acute seizures, (2) development of spontaneous recurrent seizures and (3) behavioral and neuroanatomical sequelae of kainic acid treatment. Serotonin was depleted by pretreating rats with p-chlorophenylalanine. In different groups, kainic acid was injected at 3 different doses: 6.5mg/kg, 9.0mg/kg or 12.5mg/kg. A single dose of 6.5mg/kg of kainic acid reliably induced status epilepticus in p-chlorophenylalanine-pretreated rats, but not in saline-pretreated rats. The neuroexcitatory effects of kainic acid in the p-chlorophenylalanine-pretreated rats, but not in saline-pretreated rats, were associated with the presence of tonic-clonic convulsions and high lethality. Compared to controls, a greater portion of serotonin-depleted rats showed spontaneous recurrent seizures after kainic acid injections. Loss of hippocampal neurons and spatial memory deficits associated with kainic acid treatment were exacerbated by prior depletion of serotonin. The present findings are of particular importance because they suggest that low serotonin activity may represent one of the major risk factors for epilepsy and, thus, offer potentially relevant targets for prevention of epileptogenesis.

Effects of low-frequency electrical stimulation of the anterior piriform cortex on kainate-induced seizures in rats.

Recent evidence in animals and humans suggests that low-frequency stimulation (LFS) has significant antiepileptic properties. The anterior piriform cortex (APC) is a highly susceptible seizure-trigger zone and may be critical for the initiation and propagation of seizures originating from cortical and limbic foci. We used the kainic acid (KA) seizure model in rats to assess the therapeutic effect of LFS of the APC on seizures. Adult male Sprague-Dawley rats were implanted with electrodes in the left APC and recording electrodes bilaterally in the hippocampal CA3 regions. Rats were monitored continuously with video-EEG after the emergence of spontaneous recurrent seizures that followed induction of status epilepticus by intraperitoneal KA. After two weeks of baseline recordings to determine seizure frequency, LFS of the APC was applied 60-min On 15-min Off, for two weeks with 1Hz biphasic square waves, 0.2ms pulse width, at 200μA. Another 2-week period of video-EEG monitoring was done after the cessation of LFS to study the carry-over effect. Changes in seizure frequency, severity, and duration between baseline, during LFS, and post-LFS were analyzed using the Poisson regression model. Overall seizure frequency decreased during the post-LFS period to 5% of that at baseline (p=0.003). Severe seizures (stages 4 and 5 on the Racine scale) decreased to 0% of the baseline during the post-LFS period. Two weeks of LFS of the APC reduced spontaneous seizure frequency and severity in the KA model with the effect outlasting the stimulation. Our findings suggest that the APC can be an important therapeutic target for stimulation in epilepsy.

Calpastatin Overexpression Preserves Cognitive Function Following Seizures, While Maintaining Post-Injury Neurogenesis.

In the adult mammalian brain, new neurons continue to be produced throughout life in two main regions in the brain, the subgranular zone (SGZ) in the hippocampus and the subventricular zone in the walls of the lateral ventricles. Neural stem cells (NSCs) proliferate in these niches, and migrate as neuroblasts, to further differentiate in locations where new neurons are needed, either in normal or pathological conditions. However, the endogenous attempt of brain repair is not very efficient. Calpains are proteases known to be involved in neuronal damage and in cell proliferation, migration and differentiation of several cell types, though their effects on neurogenesis are not well known. Previous work by our group has shown that the absence of calpastatin (CAST), the endogenous inhibitor of calpains, impairs early stages of neurogenesis. Since the hippocampus is highly associated with learning and memory, we aimed to evaluate whether calpain inhibition would help improve cognitive recovery after lesion and efficiency of post-injury neurogenesis in this region. For that purpose, we used the kainic acid (KA) model of seizure-induced hippocampal lesion and mice overexpressing CAST. Selected cognitive tests were performed on the 3rd and 8th week after KA-induced lesion, and cell proliferation, migration and differentiation in the dentate gyrus (DG) of the hippocampus of adult mice were analyzed using specific markers. Cognitive recovery was evaluated by testing the animals for recognition, spatial and associative learning and memory. Cognitive function was preserved by CAST overexpression following seizures, while modulation of post-injury neurogenesis was similar to wild type (WT) mice. Calpain inhibition could still be potentially able to prevent the impairment in the formation of new neurons, given that the levels of calpain activity could be reduced under a certain threshold and other harmful effects from the pathological environment could also be controlled.

Acute and spontaneous seizure onset zones in the intraperitoneal kainic acid model.

Hippocampal monitoring is often used in the intraperitoneal kainic acid (KA) seizure model for detection and quantification of early ictal activity. Here, we investigated extra-hippocampal seizure onset zones (SOZs) in this model. Eight male Sprague Dawley rats implanted with depth electrodes were continuously recorded during intraperitoneal KA injections until status epilepticus (SE) was induced. Another group of four rats was monitored chronically up to two weeks after emergence of spontaneous recurrent seizures. All rats had hippocampal electrodes. Other sampled brain regions included, among others, the claustrum, piriform cortex, and orbital cortex. Seizures recorded with video-EEG were visually analyzed. In the 58 seizures recorded during KA injections, the SOZ was extrahippocampal in 7 (12%), diffuse in 29 (50%), and hippocampal in 22 (38%). Of the 14 spontaneous seizures recorded, none were solely extrahippocampal, 10 (71%) were diffuse, and 4 (29%) were of hippocampal onset. All extra-hippocampal seizures propagated to the hippocampus within 4 to 50s (mean=14, n=7). No distinctive semiological manifestations correlated with the SOZs. We conclude that seizures can have multifocal SOZs in the KA model. This finding is important to consider when using this model, among other purposes, to screen for new therapies, study pharmacoresistance, or investigate comorbidities of epilepsy.

Wireless telemetry as an ideal tool for continuous intrahippocampal EEG recording in chronic epilepsy mouse models

Event Abstract Back to Event Wireless telemetry as an ideal tool for continuous intrahippocampal EEG recording in chronic epilepsy mouse models Yana Van Den Herrewegen1*, An Buckinx1*, Gino De Smet1, Ilse J. Smolders1 and Dimitri De Bundel1 1 Vrije Universiteit Brussel, Experimental Pharmacology (EFAR), cluster Neurosciences, Belgium Tethered recording systems are routinely used as a method for long-term continuous EEG monitoring in animal models of epilepsy. Nevertheless, tethered systems display some disadvantages when chronically used in mice, in some cases even compromising experiments and behavioral follow-up of the mice. Therefore, wireless telemetry systems are increasingly being used as an alternative method to precisely measure biopotentials in the rodent brain. As EEG recordings are the most powerful read-outs in animal models of epilepsy, we evaluated the use of wireless radio-telemetry in the intrahippocampal post-status epilepticus kainic acid model, an established mouse model for studying temporal lobe epilepsy and epileptogenesis. Via intraperitoneal implantation of a radiofrequency transmitter (ETA-F10, DSI) and electrodes positioned in the hippocampus, we were able to record intrahippocampal signals for up to two months. We could clearly monitor the epileptic activity during the status epilepticus, followed by a short latent period and the subsequent occurrence of the typical spontaneous epileptic discharges. This study discusses the advantages of telemetric EEG recordings and strategies to maximize the signal-to-noise ratio in order to obtain high-quality deep-brain EEG recordings. In conclusion, wireless telemetry systems are a valuable EEG monitoring method in the intrahippocampal kainic acid mouse model and other models of chronic epilepsy in mice. References Bouilleret V, Ridoux V, Depaulis A, et al. Recurrent seizures and hippocampal sclerosis following intrahippocampal kainate injection in adult mice: electroencephalography, histopathology and synaptic reorganization similar to mesial temporal lobe epilepsy. Neuroscience. 1999; Mar;89(3):717-29. Duveau V, Pouyatos B, Bressand K, et al. Differential Effects of Antiepileptic Drugs on Focal Seizures in the Intrahippocampal Kainate Mouse Model of Mesial Temporal Lobe Epilepsy. CNS Neuroscience & Therapeutics: CNS Neurosci Ther. 2016;(22):497-50. doi: 10.1111/cns.12523 Lundt A, Wormuth C, Siwek ME, et al. EEG Radiotelemetry in Small Laboratory Rodents: A Powerful State-of-the Art Approach in Neuropsychiatric, Neurodegenerative, and Epilepsy Research. Neural Plasticity. 2016;2016:8213878. doi:10.1155/2016/8213878. Twele F, Tollner K, Bankstahl M, Loscher W, et al. The effects of carbamazepine in the intrahippocampal kainate model of temporal lobe epilepsy depend on seizure definition and mouse strain. Epilepsia Open, 2016; 1(1):45–60,. doi: 10.1002/epi4.2 Zayachkivsky A, Lehmkuhle MJ, Dudek FE. Long-term Continuous EEG Monitoring in Small Rodent Models of Human Disease Using the Epoch Wireless Transmitter System. Journal of Visualized Experiments : JoVE. 2015;(101):52554. doi:10.3791/52554. Keywords: wireless radio-telemetry, EEG, Epilepsy, Temporal Lobe, mouse models, Seizures Conference: 12th National Congress of the Belgian Society for Neuroscience, Gent, Belgium, 22 May - 22 May, 2017. Presentation Type: Poster Presentation Topic: Novel Methods and Technology Development Citation: Van Den Herrewegen Y, Buckinx A, De Smet G, Smolders IJ and De Bundel D (2019). Wireless telemetry as an ideal tool for continuous intrahippocampal EEG recording in chronic epilepsy mouse models. Front. Neurosci. Conference Abstract: 12th National Congress of the Belgian Society for Neuroscience. doi: 10.3389/conf.fnins.2017.94.00001 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 02 May 2017; Published Online: 25 Jan 2019. * Correspondence: Miss. Yana Van Den Herrewegen, Vrije Universiteit Brussel, Experimental Pharmacology (EFAR), cluster Neurosciences, Brussels, 1090, Belgium, yana.van.den.herrewegen@vub.ac.be Miss. An Buckinx, Vrije Universiteit Brussel, Experimental Pharmacology (EFAR), cluster Neurosciences, Brussels, 1090, Belgium, an.buckinx@vub.be Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Yana Van Den Herrewegen An Buckinx Gino De Smet Ilse J Smolders Dimitri De Bundel Google Yana Van Den Herrewegen An Buckinx Gino De Smet Ilse J Smolders Dimitri De Bundel Google Scholar Yana Van Den Herrewegen An Buckinx Gino De Smet Ilse J Smolders Dimitri De Bundel PubMed Yana Van Den Herrewegen An Buckinx Gino De Smet Ilse J Smolders Dimitri De Bundel Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Scavenging of highly reactive gamma-ketoaldehydes attenuates cognitive dysfunction associated with epileptogenesis

Cognitive dysfunction is a major comorbidity of the epilepsies; however, treatments targeting seizure-associated cognitive dysfunction, particularly deficits in learning and memory are not available. Isoketals and neuroketals, collectively known as gamma-ketoaldehydes are formed via the non-enzymatic, free radical catalyzed oxidation of arachidonic acid and docosahexaenoic acid, respectively. They are attractive candidates for oxidative protein damage and resultant cognitive dysfunction due to their formation within the plasma membrane and their high proclivity to form cytotoxic adducts on protein lysine residues. We tested the hypothesis that gamma-ketoaldehydes mechanistically contribute to seizure-associated memory impairment using a specific gamma-ketoaldehyde scavenger, salicylamine in the kainic acid and pilocarpine rat models of temporal lobe epilepsy. We show that gamma-ketoaldehydes are increased following epileptogenic injury in hippocampus and perirhinal cortex, two brain regions imperative for learning and memory. Treatment with an orally bioavailable, brain permeable scavenger, salicylamine attenuated 1) spatial memory deficits 2) reference memory deficits and 3) neuronal loss and astrogliosis in two mechanistically distinct models of epilepsy without affecting the epileptogenic injury or the development of chronic epilepsy. We have previously demonstrated that reactive oxygen species and the lipid peroxidation biomarkers, F2-isoprostanes are produced following status epilepticus. However, which reactive species specifically mediate oxidative damage to cellular macromolecules remains at large. We provide novel data suggesting that memory impairment occurs via gamma-ketoaldehyde production in two models of epilepsy and that treatment with a gamma-ketoaldehyde scavenger can protect vulnerable neurons. This work suggests a novel target and therapy to treat seizure-induced memory deficits in epilepsy.

The Role of Quercetin in Gene Expression of GluR1 Subunit of AMPA Receptors, and NR2A and NR2B Subunits of NMDA Receptors in Kainic Acid Model of Seizure in Mice

Background: Flavonoids are recently being recognized for their important biological effects. Quercetin is a flavonoid that has several pharmacological properties including anti-inflammatory, neuroprotective, and anticonvulsant effects. Objectives: This study was designed to elucidate the role of quercetin in the gene expression of ionotropic glutamate receptor subunits in kainic acid (KA)-induced seizure in mice. Methods: This experimental study was conducted on 56 male BALB/c mice. Quercetin (50 and 100 mg/kg, intraperitoneally) was administered 7 days before administration of KA. The hippocampi of the animals were removed and used for molecular analysis 2 hours and 7 days after KA administration. Results: Pretreatment of mice with quercetin (100 mg/kg) significantly enhanced the gene expression of GluR1 subunit of AMPA and NR2A and NR2B subunits of NMDA only 7 days after KA administration compared to the control and KA groups in the mice hippocampus (P < 0.001, P < 0.001, and P < 0.05, respectively). Conclusions: Increment of gene expression of subunits of AMPA and NMDA receptors by quercetin might explain its protective effect on synaptic plasticity and memory.

Resting state functional network disruptions in a kainic acid model of temporal lobe epilepsy.

We studied the graph topological properties of brain networks derived from resting-state functional magnetic resonance imaging in a kainic acid induced model of temporal lobe epilepsy (TLE) in rats. Functional connectivity was determined by temporal correlation of the resting-state Blood Oxygen Level Dependent (BOLD) signals between two brain regions during 1.5% and 2% isoflurane, and analyzed as networks in epileptic and control rats. Graph theoretical analysis revealed a significant increase in functional connectivity between brain areas in epileptic than control rats, and the connected brain areas could be categorized as a limbic network and a default mode network (DMN). The limbic network includes the hippocampus, amygdala, piriform cortex, nucleus accumbens, and mediodorsal thalamus, whereas DMN involves the medial prefrontal cortex, anterior and posterior cingulate cortex, auditory and temporal association cortex, and posterior parietal cortex. The TLE model manifested a higher clustering coefficient, increased global and local efficiency, and increased small-worldness as compared to controls, despite having a similar characteristic path length. These results suggest extensive disruptions in the functional brain networks, which may be the basis of altered cognitive, emotional and psychiatric symptoms in TLE.

Exploratory Metabolomics Profiling in the Kainic Acid Rat Model Reveals Depletion of 25-Hydroxyvitamin D3 during Epileptogenesis.

Currently, no reliable markers are available to evaluate the epileptogenic potential of a brain injury. The electroencephalogram is the standard method of diagnosis of epilepsy; however, it is not used to predict the risk of developing epilepsy. Biomarkers that indicate an individual’s risk to develop epilepsy, especially those measurable in the periphery are urgently needed. Temporal lobe epilepsy (TLE), the most common form of acquired epilepsy, is characterized by spontaneous recurrent seizures following brain injury and a seizure-free “latent” period. Elucidation of mechanisms at play during epilepsy development (epileptogenesis) in animal models of TLE could enable the identification of predictive biomarkers. Our pilot study using liquid chromatography-mass spectrometry metabolomics analysis revealed changes (p-value ≤ 0.05, ≥1.5-fold change) in lipid, purine, and sterol metabolism in rat plasma and hippocampus during epileptogenesis and chronic epilepsy in the kainic acid model of TLE. Notably, disease development was associated with dysregulation of vitamin D3 metabolism at all stages and plasma 25-hydroxyvitamin D3 depletion in the acute and latent phase of injury-induced epileptogenesis. These data suggest that plasma VD3 metabolites reflect the severity of an epileptogenic insult and that a panel of plasma VD3 metabolites may be able to serve as a marker of epileptogenesis.

Plasma cysteine/cystine redox couple disruption in animal models of temporal lobe epilepsy

Currently the field of epilepsy lacks peripheral blood-based biomarkers that could predict the onset or progression of chronic seizures following an epileptogenic injury. Thiol/disulfide ratios have been shown to provide a sensitive means of assessing the systemic redox potential in tissue and plasma. In this study, we utilized a rapid, simple and reliable method for simultaneous determination of several thiol-containing amino acids in plasma using HPLC with electrochemical detection in kainic acid (KA) and pilocarpine rat models of epilepsy. In contrast to GSH and GSSG levels, the levels of cysteine (Cys) were decreased by 42% and 62% and cystine (Cyss) were increased by 46% and 23% in the plasma of KA- and pilocarpine-injected rats, respectively after 48h. In chronically epileptic rats, plasma cysteine was decreased by 40.4% and 37.7%, and plasma GSSG increased by 33.8% and 35.0% following KA and pilocarpine, respectively. Treatment of rats with a catalytic antioxidant, 60min after KA or pilocarpine significant attenuated the decrease of plasma Cys/Cyss ratios at the 48h time point in both models. These observations suggest that the decreased cysteine and ratio of Cys/Cyss in plasma could potentially serve as redox biomarkers in temporal lobe epilepsy.

Effects of genistein on cognitive dysfunction and hippocampal synaptic plasticity impairment in an ovariectomized rat kainic acid model of seizure

The major objective of this study was to investigate the probable effects of genistein (one of the most important soy phytoestrogens-SPEs) on seizure-induced cognitive dysfunction, hippocampal early long-term potentiation (E-LTP) impairment and morphological damage to CA1 neurons in ovariectomized (OVX) rats.Three weeks after ovariectomy, cannulae were implanted over the left lateral ventricle. After a 7-day recovery period, animals were injected by genistein (0.5 or 5mg/kg) or vehicle during four consecutive days, each 24h. One h after the last treatment, kainic acid (KA) or vehicle was perfused into the left lateral ventricle to induce generalized tonic-clonic seizures. Finally, 7 days later, spatial learning and memory of animals were examined using the Morris water maze (MWM) task, hippocampal E-LTP was assessed using in-vivo field potential recordings and the morphology of hippocampal CA1 area was examined using Fluoro-Jade C staining.KA-induced generalized seizures resulted in spatial learning and memory impairment, E-LTP deficit and CA1 cell injury. Seizure-induced abnormalities improved partially only by the lower dose of genistein (0.5mg/kg). However, genistein at the higher dose (5mg/kg) did not have any beneficial effects. Also, genistein did not affect seizure activity.It is concluded that genistein may have partially preventive effects against seizure-induced cognitive impairment in OVX rats. Also, it seems that such effects of genistein are correlated with its beneficial effects on hippocampal synaptic plasticity and morphology.

Regulation of astrocyte glutamate transporter-1 (GLT1) and aquaporin-4 (AQP4) expression in a model of epilepsy

Astrocytes regulate extracellular glutamate and water homeostasis through the astrocyte-specific membrane proteins glutamate transporter-1 (GLT1) and aquaporin-4 (AQP4), respectively. The role of astrocytes and the regulation of GLT1 and AQP4 in epilepsy are not fully understood. In this study, we investigated the expression of GLT1 and AQP4 in the intrahippocampal kainic acid (IHKA) model of temporal lobe epilepsy (TLE). We used real-time polymerase chain reaction (RT-PCR), Western blot, and immunohistochemical analysis at 1, 4, 7, and 30days after kainic acid-induced status epilepticus (SE) to determine hippocampal glial fibrillary acidic protein (GFAP, a marker for reactive astrocytes), GLT1, and AQP4 expression changes during the development of epilepsy (epileptogenesis). Following IHKA, all mice had SE and progressive increases in GFAP immunoreactivity and GFAP protein expression out to 30days post-SE. A significant initial increase in dorsal hippocampal GLT1 immunoreactivity and protein levels were observed 1day post SE and followed by a marked downregulation at 4 and 7days post SE with a return to near control levels by 30days post SE. AQP4 dorsal hippocampal protein expression was significantly downregulated at 1day post SE and was followed by a gradual return to baseline levels with a significant increase in ipsilateral protein levels by 30days post SE. Transient increases in GFAP and AQP4 mRNA were also observed. Our findings suggest that specific molecular changes in astrocyte glutamate transporters and water channels occur during epileptogenesis in this model, and suggest the novel therapeutic strategy of restoring glutamate and water homeostasis.