Rat Model Of Post-traumatic Epilepsy Research Articles

A comparison of the antiepileptogenic efficacy of two rationally chosen multitargeted drug combinations in a rat model of posttraumatic epilepsy

Post-traumatic epilepsy (PTE) is a recurrent and often drug-refractory seizure disorder caused by traumatic brain injury (TBI). No single drug treatment prevents PTE, but preventive drug combinations that may prophylax against PTE have not been studied. Based on a systematic evaluation of rationally chosen drug combinations in the intrahippocampal kainate (IHK) mouse model of acquired epilepsy, we identified two multi-targeted drug cocktails that exert strong antiepileptogenic effects. The first, a combination of levetiracetam (LEV) and topiramate, only partially prevented spontaneous recurrent seizures in the model. We therefore added atorvastatin (ATV) to the therapeutic cocktail (TC) to increase efficacy, forming “TC-001”. The second cocktail – a combination of LEV, ATV, and ceftriaxone, termed “TC-002” – completely prevented epilepsy in the mouse IHK model. In the present proof-of-concept study, we tested whether the two drug cocktails prevent epilepsy in a rat PTE model in which recurrent electrographic seizures develop after severe rostral parasagittal fluid percussion injury (FPI). Following FPI, rats were either treated over 3–4 weeks with vehicle or drug cocktails, starting either 1 or 4–6 h after the injury. Using mouse doses of TC-001 and TC-002, no significant antiepileptogenic effect was obtained in the rat PTE model. However, when using allometric scaling of drug doses to consider the differences in body surface area between mice and rats, PTE was prevented by TC-002. Furthermore, the latter drug cocktail partially prevented the loss of perilesional cortical parvalbumin-positive GABAergic interneurons. Plasma and brain drug analysis showed that these effects of TC-002 occurred at clinically relevant levels of the individual TC-002 drug components. In silico analysis of drug-drug brain protein interactions by the STITCH database indicated that TC-002 impacts a larger functional network of epilepsy-relevant brain proteins than each drug alone, providing a potential network pharmacology explanation for the observed antiepileptogenic and neuroprotective effects observed with this combination.

Celecoxib treatment alleviates cerebral injury in a rat model of post-traumatic epilepsy

Background An important factor contributing to the development and occurrence of post-traumatic epilepsy (PTE) is neuroinflammation and oxidative stress. The effects of celecoxib include inhibiting inflammatory reactions and antioxidant stress and reducing seizures, making it a potential epilepsy treatment solution. Objective To observe the effect of celecoxib on early epilepsy in post-traumatic epilepsy rats. Methods: Twenty-four adult healthy male Sprague-Dawley rats were randomly assigned to three groups: sham-operated, PTE, and celecoxib. A rat model of PTE was established by injecting ferrous chloride into the right frontal cortex. Afterward, the behavior of rats was observed and recorded. 3.0T superconducting magnetic resonance imaging (MRI) was used to describe the changes in ADC values of the brain. HE and Nissl staining were also used to detect the damage to frontal lobe neurons. Furthermore, the expression of COX-2 protein in the right frontal lobe was detected by Western blot. Moreover, the contents of IL-1 and TNF-α in the right frontal lobe were detected by enzyme-linked immunosorbent assay. Results Compared with the PTE group, the degree of seizures in rats treated with celecoxib declined dramatically (P < 0.05). Celecoxib-treated rats had significant decreases in tissue structural damage and cell death in the brain. The results of the MRI showed that celecoxib reduced the peripheral edema zone and ADC value of the cortex around the damaged area of the right frontal lobe in the celecoxib-treatment group, which was significantly decreased compared with the PTE group (P < 0.05). Furthermore, celecoxib decreased the expression of COX-2, IL-1β, and TNF-α in brain tissue (P < 0.05). Conclusions In PTE rats, celecoxib significantly reduced brain damage and effectively reduced seizures. As a result of celecoxib’s ability to inhibit inflammation, it can reduce the edema caused by injury in rat brain tissue.

A machine learning approach to seizure detection in a rat model of post-traumatic epilepsy

Epilepsy is a common neurologic condition frequently investigated using rodent models, with seizures identified by electroencephalography (EEG). Given technological advances, large datasets of EEG are widespread and amenable to machine learning approaches for identification of seizures. While such approaches have been explored for human EEGs, machine learning approaches to identifying seizures in rodent EEG are limited. We utilized a predesigned deep convolutional neural network (DCNN), GoogLeNet, to classify images for seizure identification. Training images were generated through multiplexing spectral content (scalograms), kurtosis, and entropy for two-second EEG segments. Over 2200 h of EEG data were scored for the presence of seizures, with 95.6% of seizures identified by the DCNN and a false positive rate of 34.2% (1.52/h), as compared to visual scoring. Multiplexed images were superior to scalograms alone (scalogram-kurtosis-entropy 0.956 ± 0.010, scalogram 0.890 ± 0.028, t(7) = 3.54, p < 0.01) and a DCNN trained specifically for the individual animal was superior to using DCNNs across animals (intra-animal 0.960 ± 0.0094, inter-animal 0.811 ± 0.015, t(30) = 5.54, p < 0.01). For this dataset the DCNN approach is superior to a previously described algorithm utilizing longer local line lengths, calculated from wavelet-decomposition of EEG, to identify seizures. We demonstrate the novel use of a predesigned DCNN constructed to classify images, utilizing multiplexed images of EEG spectral content, kurtosis, and entropy, to rapidly and objectively identifies seizures in a large dataset of rat EEG with high sensitivity.

Pharmacotherapeutic Effects of Hippophae rhamnoides in Rat Model of Post-traumatic Epilepsy in View of Oxidative Stress, Na+,K+ATPase Activity and Sodium Ion Channel Expression

Pharmacotherapeutic Effects of Hippophae rhamnoides in Rat Model of Post-traumatic Epilepsy in View of Oxidative Stress, Na+,K+ATPase Activity and Sodium Ion Channel Expression

Pyrazolo[4,3-c]pyridine-4-one (PP-4-one) Exhibits Anti-Epileptogenic Effect in Rat Model of Traumatic Epilepsy by Mammalian Target of Rapamycin (mTOR) Signaling Pathway Downregulation

BackgroundPost-traumatic epilepsy (PTE) is a common type of acquired epilepsies secondary to traumatic brain injury (TBI), accounting for approximately 10–25% of patients. The present study evaluated activity of PP-4-one against mTOR signaling activation in a rat model of FeCl2-induced post-traumatic epilepsy.Material/MethodsEpilepsy in rats was induced by injecting 10 μl FeCl2 (concentration 100 mM) at a uniform rate of 1 μl/minute. The iNOS expression was detected using a Leica microscope connected to a digital camera system. Reverse transcription polymerase chain reaction (RT-PCR) was used for determination of NR1 mRNA expression.ResultsThe post-traumatic epilepsy induced neuronal degeneration in the hippocampus and frontal cortex of the rats. Treatment with PP-4-one prevented neuronal degeneration in the hippocampus and frontal cortex in rats with post-traumatic epilepsy. The data revealed markedly higher levels of p-mTOR and p-P70S6K in rat hippocampal tissues after induction of traumatic epilepsy. Treatment of post-traumatic epilepsy rats with PP-4-one significantly suppressed p-mTOR and p-P70S6K expression, and PP-4-one treatment reduced epileptic brain injury in the rats with post-traumatic epilepsy.ConclusionsPP-4-one exhibits an anti-epileptogenic effect in the rat model of PTE by inhibiting behavioral seizures through suppression of iNOS and astrocytic proliferation. Moreover, PP-4-one treatment suppressed NR1 expression and targeted the mTOR pathway in PTE-induced rats. Thus, PP-4-one shows promise as a novel and effective therapeutic agent for treatment of epilepsy induced by PTE.

Harmonization of pipeline for detection of HFOs in a rat model of post-traumatic epilepsy in preclinical multicenter study on post-traumatic epileptogenesis

Studies of chronic epilepsy show pathological high frequency oscillations (HFOs) are associated with brain areas capable of generating epileptic seizures. Only a few of these studies have focused on HFOs during the development of epilepsy, but results suggest pathological HFOs could be a biomarker of epileptogenesis. The Epilepsy Bioinformatics Study for Antiepileptogenic Therapy” (EpiBioS4Rx) is a multi-center project designed to identify biomarkers of epileptogenesis after a traumatic brain injury (TBI) and evaluate treatments that could modify or prevent the development of post-traumatic epilepsy. One goal of the EpiBioS4Rx project is to assess whether HFOs could be a biomarker of post-traumatic epileptogenesis. The current study describes the work towards this goal, including the development of common surgical procedures and EEG protocols, an interim analysis of the EEG for HFOs, and identifying issues that need to be addressed for a robust biomarker analysis. At three participating sites – University of Eastern Finland (UEF), Monash University in Melbourne (Melbourne) and University of California, Los Angeles (UCLA) – TBI was induced in adult male Sprague-Dawley rats by lateral fluid-percussion injury. After injury and in sham-operated controls, rats were implanted with screw and microwire electrodes positioned in neocortex and hippocampus to record EEG. A separate group of rats had serial magnetic resonance imaging after injury and then implanted with electrodes at 6 months. Recordings 28 days post-injury were available from UEF and UCLA, but not Melbourne due to technical issues with their EEG files. Analysis of recordings from 4 rats – UEF and UCLA each had one TBI and one sham-operated control – showed EEG contained evidence of HFOs. Computer-automated algorithms detected a total of 1,819 putative HFOs and of these only 40 events (2%) were detected by all three sites. Manual review of all events verified 130 events as HFO and the remainder as false positives. Review of the 40 events detected by all three sites was associated with 88% agreement. This initial report from the EpiBioS4Rx Consortium demonstrates the standardization of EEG electrode placements, recording protocol and long-term EEG monitoring, and differences in detection algorithm HFO results between sites. Additional work on detection strategy, detection algorithm performance, and training in HFO review will be performed to establish a robust, preclinical evaluation of HFOs as a biomarker of post-traumatic epileptogenesis.

Harmonization of pipeline for preclinical multicenter plasma protein and miRNA biomarker discovery in a rat model of post-traumatic epileptogenesis

The Epilepsy Bioinformatics Study for Antiepileptogenic Therapy (EpiBioS4Rx) is an international, multicenter, multidisciplinary study aimed at preventing epileptogenesis (EpiBioS4Rx: https://epibios.loni.usc.edu/). One of the study’s major objectives is the discovery of diagnostic, prognostic, and predictive plasma protein and microRNA (miRNA) biomarkers that are sensitive, specific, and translatable to the human condition. Epilepsy due to structural brain abnormalities, secondary to neurological insults such as traumatic brain injury (TBI), currently represents ∼50% of all epilepsy cases. In the preclinical EpiBioS4Rx study, TBI was induced in adult male Sprague Dawley rats using a standardized protocol for lateral fluid-percussion injury. Whole blood was collected from the tail vein at baseline and 2, 9 and 30 days post-injury and processed for plasma separation. Biomaterial properties, sample preparation and integrity, and choice of analysis platform can significantly impact measured marker levels and, in turn, interpretation with respect to injury and/or other variables. We present here the results of procedural harmonization for the first 320 rats included in the EpiBioS4Rx study study, from three international research centers, and preliminary proteomic and miRNA analyses. We also discuss experimental considerations for establishing rigorous quality controls with the goal of harmonizing operating procedures across study sites, and delivering high-quality specimens for preclinical biomarker discovery in a rat model of post-traumatic epilepsy (PTE).

Algorithm for automatic detection of spontaneous seizures in rats with post-traumatic epilepsy

BackgroundLabor intensive electroencephalogram (EEG) analysis is a major bottleneck to identifying anti-epileptogenic treatments in experimental models of post-traumatic epilepsy. We aimed to develop an algorithm for automated seizure detection in experimental post-traumatic epilepsy. New methodContinuous (24/7) 1-month-long video-EEG monitoring with three epidural screw electrodes was started 154 d after lateral fluid-percussion induced traumatic brain injury (TBI; n = 97) or sham-injury (n = 29) in adult male Sprague–Dawley rats. First, an experienced researcher screened a total of 90,720 h of digitized recordings on a computer screen to annotate the occurrence of spontaneous seizures. The same files were then analyzed using an algorithm in Spike2 (ver.9), which searching for temporally linked power peaks (14–42 Hz) in all three EEG channels, and then positive events were marked as a probable seizures. Finally, an experienced researcher confirmed all seizure candidates visually on the computer screen. ResultsVisual analysis identified 197 seizures in 29 rats. Automatic detection identified 4346 seizure candidates in 109 rats, of which 202 in the same 29 rats were true positives, resulting in a false positive rate of 0.046/h or 1.10/d. The algorithm demonstrated 5% specificity and 100% sensitivity. The algorithm analyzed 1-month 3-channel EEG in 7 cohorts in 2 h, whereas analysis by an experienced technician took ∼500 h. Comparison with Existing MethodsThe algorithm had 100% sensitivity. It performed slightly better and was substantially faster than investigator-performed visual analysis. ConclusionsWe present a novel seizure detection algorithm for automated detection of seizures in a rat model of post-traumatic epilepsy.

Autoregressive spectral analysis of cortical electroencephalographic signals in a rat model of post-traumatic epilepsy

Objectives:Post-traumatic epilepsy (PTE) is a common consequence of traumatic brain injury (TBI) and significant predictor of poor prognosis in TBI patients. To develop clinical interventions for PTE risk reduction, there is a need to elucidate the epileptogenic mechanisms induced by brain injury.Methods:The iron-induced rat model of epilepsy used here mimics many aspects of human PTE. Intracortical injection of iron results in local neuronal damage and the establishment of an epileptic focus, leading to chronic spontaneous electroencephalographic (EEG) signals and motor seizures, with progressively increasing frequency over many months. Identifying unique aspects of PTE seizure semiology for prognosis and treatment may be aided by novel methods of EEG analysis. Here, autoregressive (AR) methods were compared to the conventional fast Fourier transform (FFT) for processing EEG signals in iron-induced epilepsy.Results:Power spectra obtained using AR showed higher frequency resolution over a given epoch than the spectra obtained using FFT. Moreover, changes in total AR spectral power and frequency distribution over brief successive periods provided convenient indexes for long-term monitoring of seizures.Discussion:Autoregression analysis may prove complementary to FFT for EEG analysis in PTE patients.

Optimized methods for epilepsy therapy development using an etiologically realistic model of focal epilepsy in the rat

Conventionally developed antiseizure drugs fail to control epileptic seizures in about 30% of patients, and no treatment prevents epilepsy. New etiologically realistic, syndrome-specific epilepsy models are expected to identify better treatments by capturing currently unknown ictogenic and epileptogenic mechanisms that operate in the corresponding patient populations. Additionally, the use of electrocorticography permits better monitoring of epileptogenesis and the full spectrum of acquired seizures, including focal nonconvulsive seizures that are typically difficult to treat in humans. Thus, the combined use of etiologically realistic models and electrocorticography may improve our understanding of the genesis and progression of epilepsy, and facilitate discovery and translation of novel treatments. However, this approach is labor intensive and must be optimized. To this end, we used an etiologically realistic rat model of posttraumatic epilepsy, in which the initiating fluid percussion injury closely replicates contusive closed-head injury in humans, and has been adapted to maximize epileptogenesis and focal non-convulsive seizures. We obtained week-long 5-electrode electrocorticography 1month post-injury, and used a Monte-Carlo-based non-parametric bootstrap strategy to test the impact of electrode montage design, duration-based seizure definitions, group size and duration of recordings on the assessment of posttraumatic epilepsy, and on statistical power to detect antiseizure and antiepileptogenic treatment effects. We found that use of seizure definition based on clinical criteria rather than event duration, and of recording montages closely sampling the activity of epileptic foci, maximize the power to detect treatment effects. Detection of treatment effects was marginally improved by prolonged recording, and 24h recording epochs were sufficient to provide 80% power to detect clinically interesting seizure control or prevention of seizures with small groups of animals. We conclude that appropriate electrode montage and clinically relevant seizure definition permit convenient deployment of fluid percussion injury and electrocorticography for epilepsy therapy development.

Novel frontiers in epilepsy treatments: preventing epileptogenesis by targeting inflammation

Currently available epilepsy drugs only affect the symptoms (seizures), and there is a need for innovative treatments that target the underlying disease. Increasing evidence points to inflammation as a potentially important mechanism in epileptogenesis. In the last decade, a new generation of etiologically realistic syndrome-specific experimental models have been developed, which are expected to capture the epileptogenic mechanisms operating in corresponding patient populations, and to exhibit similar treatment responsiveness. Recently, an intervention known to have broad-ranging anti-inflammatory effects (selective brain cooling) has been found to prevent the development of spontaneously occurring seizures in an etiologically realistic rat model of post-traumatic epilepsy. Several drugs used clinically for other indications also have the potential for inhibiting inflammation, and should be investigated for antiepileptogenic activity in these models. If results of such studies are positive, these compounds could rapidly enter Phase III trials in patients at high risk of developing epilepsy.

Hippocampal Gene Expression Profiling in a Rat Model of Posttraumatic Epilepsy Reveals Temporal Upregulation of Lipid Metabolism-Related Genes

Traumatic brain injury occasionally causes posttraumatic epilepsy. To elucidate the molecular events responsible for posttraumatic epilepsy, we established a rodent model that involved the injection of microliter quantities of FeCl3 solution into the amygdalar nuclear complex. We previously compared hippocampal gene expression profiles in the traumatic epilepsy model and normal rats at 5 days after brain injury (acute phase) to determine the role of inflammation. In this study, we focused on later stages of epileptogenesis. We compared gene expression profiles at 5, 15 (sub-chronic phase), and 30 days (chronic phase) after brain injury to identify temporal changes in molecular networks involved in epileptogenesis. A total of 81 genes were significantly (at least twofold) up- or downregulated over the course of disease progression. We found that genes related to lipid metabolism, namely, Apoa1, Gh, Mc4r, Oprk1, and Pdk4, were temporarily upregulated in the sub-chronic phase. Changes in lipid metabolism regulation might be related to seizure propagation during epileptogenesis. This temporal description of hippocampal gene expression profiles throughout epileptogenesis provides clues to potential markers of disease phases and new therapeutic targets.

Carbenoxolone pretreatment and treatment of posttraumatic epilepsy.

Gap junction blocking agents can inhibit spontaneous discharge frequency in cells. We established a rat model of posttraumatic epilepsy induced using ferric ions. Rats were intraperitoneally injected with carbenoxolone, 20 mg/kg, prior to and 30 minutes after model establishment, once a day for 14 consecutive days. Immunohistochemistry showed glial cell proliferation around a cortical focus and significantly increased connexin expression in posttraumatic epilepsy. However, carbenoxolone pretreatment or treatment significantly reduced connexin expression in the cortex, inhibited glial fibrillary acidic protein expression and ameliorated seizure degree in rats. These findings indicate that large amounts of glial cell proliferation and abnormal gap junction generation play a role in posttraumatic epilepsy, and that carbenoxolone may prevent and treat this disease.

Presynaptic Inhibitory Terminals Are Functionally Abnormal in a Rat Model of Posttraumatic Epilepsy

Partially isolated "undercut" neocortex with intact pial circulation is a well-established model of posttraumatic epileptogenesis. Results of previous experiments showed a decreased frequency of miniature inhibitory postsynaptic currents (mIPSCs) in layer V pyramidal (Pyr) neurons of undercuts. We further examined possible functional abnormalities in GABAergic inhibition in rat epileptogenic neocortical slices in vitro by recording whole cell monosynaptic IPSCs in layer V Pyr cells and fast-spiking (FS) GABAergic interneurons using a paired pulse paradigm. Compared with controls, IPSCs in Pyr neurons of injured slices showed increased threshold and decreased peak amplitude at threshold, decreased input/output slopes, increased failure rates, and a shift from paired pulse depression toward paired pulse facilitation (increased paired pulse ratio or PPR). Increasing [Ca(2+)](o) from 2 to 4 mM partially reversed these abnormalities in Pyr cells of the epileptogenic tissue. IPSCs onto FS cells also had an increased PPR and failures. Blockade of GABA(B) receptors did not affect the paired results. These findings suggest that there are functional alterations in GABAergic presynaptic terminals onto both Pyr and FS cells in this model of posttraumatic epileptogenesis.

α-Tocopheryl- l-ascorbate-2-O-phosphate diester, a hydroxyl radical scavenger, prevents the occurrence of epileptic foci in a rat model of post-traumatic epilepsy

Intracortical injection of iron ions has been used to model post-traumatic epilepsy. The results obtained using these models suggest that oxidation of neural membranes by active oxygen free radicals may be involved in the etiology of post-traumatic epilepsy. This is a study of the effects of α-tocopheryl- l -ascorbate-2-O-phosphate diester potassium salt (EPC-K1), known as a hydroxyl radical scavenger, on the peroxidation of neural membranes by FeCl 3 in vitro and on the occurrence of epileptic discharges in the FeCl 3 injected post-traumatic epilepsy model rats. EPC-K1 dose-dependently inhibited the production of thiobarbituric acid reactive substances (TBARS) and protein carbonyl (P-Carb), both indices of biogenic macromolecular peroxidation. In vivo studies, sporadic spike discharges and/or epileptiform activities were observed in electrocorticograms (ECoG) of male Sprague–Dawley rat 15–90 min after 500 nmol of FeCl 3 was injected into the motor cortex. On the other hand, when 200 mg/kg of EPC-K1 was injected intraperitoneally 60 min prior to the injection of FeCl 3, the occurrence of epileptic discharges was prevented or delayed. When EPC-K1 (2.5–5 nmol) was injected along with the ferric ions, the occurrence of epileptiform activities was also prevented or delayed. EPC-K1 prevented the induction of early convulsion, the major risk factor of post-traumatic epilepsy. Rats in the Fe+EPC group were injected with 500 nmol of FeCl 3 into the left motor cortex and were given an EPC-K1-diet (CE-2 chow contained 0.2% of EPC-K1, and daily EPC-K1 intake was about 80 mg/kg/day). In the Fe+EPC group rats, the percent induction of epileptic discharges in ECoGs was significantly lower than that in the Fe+CE group rats, which were fed CE-2 after FeCl 3 injection. In the homotropic contra lateral cortex, TBARS and P-Carb content did not show any changes. However, the relative TBARS content in the focal area significantly increased in the Fe+CE and Fe+EPC group rats 3 h after the injection. It became normal 3 days after in the Fe+EPC group. The relative P-Carb content in the focal area significantly increased in the Fe+CE and Fe+EPC group rats 3 h after the injection. However, it became normal after 3 days. In the present study, EPC-K1, which consists of vitamins E and C connected by a phosphate, protected the oxidation of neural membranes and prevented the occurrence of ferric ion-induced epileptic discharges by its radical scavenger activity. These data suggest that EPC-K1 may be clinically useful in not only preventing the focus formation of post-traumatic epilepsy, but also in treating and attenuating the progression of free radical-induced degenerative disorders.

Decreased cortical levels of astrocytic glutamate transport protein GLT-1 in a rat model of posttraumatic epilepsy

The extracellular homeostasis of glutamate in the brain is maintained by the efficient uptake into astroglial cells. The high extracellular glutamate levels seen during seizures are therefore probably a result of both an increased synaptic release and a deranged glutamate uptake. In this study we used immuno-blotting technique to measure the cortical levels of the astrocytic glutamate transport protein (GLT-1) and of the glutamate and aspartate transporting protein (GLAST) in an epilepsy model induced by ferrous chloride injection in the cortex of rats. The levels of GLT-1 were lower in epileptic rats than in controls, day 1 and 5 after induction, but not at 3 months. Glial fibrillary protein (GFAP) levels increased with time in the epileptic model, whereas GLAST and β-tubulin III remained unchanged compared to controls. The results suggest that the transient decrease of GLT-1 could play a role in epileptogenesis, while recurrent seizure activity may be maintained by other mechanisms.