The standardized extract of Centella asiatica L. Urb attenuates the convulsant effect induced by lithium/pilocarpine without affecting biochemical and haematological parameters in rats.

BACKGROUND: Status epilepticus (SE) is a type of epileptic activity characterized by a failure of the inhibitory mechanisms that limit seizures, which are mainly regulated by the GABAergic system. This imbalance increases glutamatergic neurotransmission and consequently produces epileptic activity. It is also associated with oxidative stress due to an imbalance between reactive oxygen species (ROS) and antioxidant defences. Unfortunately, long-term treatment with anti-epileptic drugs (AEDs) may produce hepatotoxicity, nephrotoxicity, and haematological alterations. In this way, some secondary metabolites of plants have been used to ameliorate the deterioration of nervous system disorders through their antioxidant properties, in addition to their anticonvulsant effects. An example is Centella asiatica, a plant noted to have a reputed neuroprotective effect related to its antioxidant activity. However, similar to conventional drugs, natural molecules may produce side effects when consumed in high doses, which could occur with Centella asiatica. Therefore, we aimed to evaluate the effect of a standardized extract of Centella asiatica L. Urb with tested anticonvulsant activity on biochemical and haematological parameters in rats subjected to lithium/pilocarpine-induced seizures. METHODS: Twenty-eight adult male Wistar rats were randomly divided into four groups (n = 7 each): vehicle (purified water), Centella asiatica (200 and 400 mg/kg), and carbamazepine (CBZ) (300 mg/kg) as a pharmacological control of anticonvulsant activity. Treatments were administered orally every 24 h for 35 consecutive days. On Day 36, SE was induced using the lithium/pilocarpine model (3 mEq/kg, i.p. and 30 mg/kg s.c., respectively), and the behavioural and biochemical effects were evaluated. RESULTS: Centella asiatica 400 mg/kg increased the latency to the first generalized seizure and SE onset and significantly reduced the time to the first generalized seizure compared to values in the vehicle group. Biochemical parameters, i.e., haematic cytometry, blood chemistry, and liver function tests, showed no significant differences among the different treatments. CONCLUSION: The dose of Centella asiatica that produces anticonvulsant activity in the lithium/pilocarpine model devoid of hepatotoxicity, nephrotoxicity, and alterations in haematological parameters suggests that the standardized extract of this plant could be of utility in the development of new safe therapies for the treatment of convulsions associated with epilepsy.

Effects of Stevia rebaudiana Bertoni extracts in the rat model of epilepsy induced by pentylenetetrazol: Sirt-1, at the crossroads between inflammation and apoptosis.

The current study investigated the effects of stevia extracts on a PTZ-induced epileptic rat model and its potential mechanism. Thirty male Sprague-Dawley rats were equally subdivided into 3 groups; (1) normal control (NC) group, (2) PTZ-group: received PTZ (50 mg/kg, i.p. every other day) for 2 weeks, and (3) PTZ+ Stevia group: received PTZ and stevia (200 mg/kg orally daily) for 4 weeks (2 weeks before the start of PTZ treatment and 2 weeks with PTZ administration). The first jerk latency and the seizure score were assessed in rats. Also, brain tissue samples were collected by the end of the experiment, and oxidative stress markers (catalase, MDA, and total antioxidant capacity (TAC)) were measured by biochemical analysis in hippocampal brain homogenates. Also, in the hippocampus, the expression of IL6 and Bcl-2 at the mRNA level and expression of Sirt-1, P53, caspase-3, GFAP, and NF-kB in CA3 hippocampal region by immunohistochemistry was investigated. PTZ substantially increased the seizure score and decreased the seizure latency. Also, PTZ significantly increased MDA, GFAP, IL-6, NF-kB, caspase-3, and p53 and significantly reduced Sirt-1, TAC, and Bcl-2 in hippocampal tissues compared to the control group (p < 0.01). However, Stevia Rebaudiana Bertoni (Stevia R.) significantly attenuated the PTZ-induced seizures, improved oxidative stress markers, downregulated GFAP, IL-6, NF-kB, caspase-3, and p53, and upregulated Sirt-1 and Bcl-2 in the CA3 hippocampal region (p < 0.01). In conclusion, Stevia R. exhibits neuroprotective and antiepileptic actions in PTZ-induced epilepsy due to its antioxidant, anti-apoptotic, and anti-inflammatory effects. Additionally, the Sirt-1 pathway might be involved in the antiepileptic and neuroprotective effects of stevia in PTZ-kindled epileptic rat model.

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

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

Comparative Antiseizure Analysis of Diverse Natural Coumarin Derivatives in Zebrafish.

Coumarins are a well-known group of plant secondary metabolites with various pharmacological activities, including antiseizure activity. In the search for new antiseizure drugs (ASDs) to treat epilepsy, it is yet unclear which types of coumarins are particularly interesting as a systematic analysis has not been reported. The current study performed behavioral antiseizure activity screening of 18 different coumarin derivatives in the larval zebrafish pentylenetetrazole (PTZ) model using locomotor measurements. Activity was confirmed for seven compounds, which lowered seizure-like behavior as follows: oxypeucedanin 38%, oxypeucedanin hydrate 74%, notopterol 54%, nodakenetin 29%, hyuganin C 35%, daphnoretin 65%, and pimpinellin 60%. These coumarins, together with nodakenin, underwent further antiepileptiform analysis by local field potential recordings from the zebrafish opticum tectum (midbrain). All of them, except for nodakenetin, showed pronounced antiepileptiform activity, decreasing PTZ-induced elevation in power spectral density (PSD) by 83-89% for oxypeucedanin, oxypeucedanin hydrate, and notopterol, 77% for nodakenin, 26% for nodakenetin, 65% for hyuganin C, 88% for daphnoretin, and 81% for pimpinellin. These data demonstrate the potential of diverse coumarin scaffolds for ASD discovery. Finally, the structural differences between active and inactive coumarins were investigated in silico for oxypeucedanin hydrate and byacangelicin for their interaction with GABA-transaminase, a hypothetical target.

The Anticonvulsant Effect of Hydroethanolic Leaf Extract of Calotropis procera (Ait) R. Br. (Apocynaceae).

A number of currently used drugs have been obtained from medicinal plants which are a major source of drugs. These drugs are either used in their pure form or modified to a semisynthetic drug. Drug discovery through natural product research has been fruitful over the years. Traditionally, Calotropis procera is used extensively in the management of epilepsy. This study is conducted to explore the anticonvulsant effect of a hydroethanolic leaf extract of Calotropis procera (CPE) in murine models. This effect was evaluated using picrotoxin-induced convulsions, strychnine-induced convulsions, and isoniazid- and pilocarpine-induced status epilepticus in mice of both sexes. The results showed that CPE (100-300 mg/kg) exhibited an anticonvulsant effect against strychnine-induced clonic seizures by significantly reducing the duration (p = 0.0068) and frequency (p = 0.0016) of convulsions. The extract (100-300 mg/kg) caused a profound dose-dependent delay in the onset of clonic convulsions induced by picrotoxin (p < 0.0001) and tonic convulsions (p < 0.0001) in mice. The duration of convulsions was reduced significantly also for both clonic and tonic (p < 0.0001) seizures as well. CPE (100-300 mg/kg), showed a profound anticonvulsant effect and reduced mortality in the pilocarpine-induced convulsions. ED50 (~0.1007) determined demonstrated that the extract was less potent than diazepam in reducing the duration and onset of convulsions but had comparable efficacies. Flumazenil-a GABAA receptor antagonist-did not reverse the onset or duration of convulsions produced by the extract in the picrotoxin-induced seizure model. In isoniazid-induced seizure, CPE (300 mg kg1, p.o.) significantly (p < 0.001) delayed the onset of seizure in mice and prolonged latency to death in animals. Overall, the hydroethanolic leaf extract of Calotropis procera possesses anticonvulsant properties.

Development of an antiepileptogenesis drug screening platform: Effects of everolimus and phenobarbital.

OBJECTIVE: The kainic acid (KA)-induced status epilepticus (SE) model in rats is a well-defined model of epileptogenesis. This model closely recapitulates many of the clinical and pathological characteristics of human temporal lobe epilepsy (TLE) that arise following SE or another neurological insult. Spontaneous recurrent seizures (SRS) in TLE can present after a latent period following a neurological insult (traumatic brain injury, SE event, viral infection, etc.). Moreover, this model is suitable for preclinical studies to evaluate the long-term process of epileptogenesis and screen putative disease-modifying/antiepileptogenic agents. The burden of human TLE is highly variable, similar to the post-KA SE rat model. In this regard, this model may have broad translational relevance. This report thus details the pharmacological characterization and methodological refinement of a moderate-throughput drug screening program using the post-KA-induced SE model of epileptogenesis in male Sprague Dawley rats to identify potential agents that may prevent or modify the burden of SRS. Specifically, we sought to demonstrate whether our protocol could prevent the development of SRS or lead to a reduced frequency/severity of SRS. METHODS: Rats were administered either everolimus (2-3 mg/kg po) beginning 1, 2, or 24 h after SE onset, or phenobarbital (60 mg/kg ip) beginning 1 h after SE onset. All treatments were administered once/day for 5-7 days. Rats in all studies (n = 12/treatment dose/study) were then monitored intermittently by video-electroencephalography (2 weeks on, 2 weeks off, 2 weeks on epochs) to determine latency to onset of SRS and disease burden. RESULTS: Although no adverse side effects were observed in our studies, no treatment significantly modified disease or prevented the presentation of SRS by 6 weeks after SE onset. SIGNIFICANCE: Neither phenobarbital nor everolimus administered at several time points after SE onset prevented the development of SRS. Nonetheless, we demonstrate a practical and moderate-throughput screen for potential antiepileptogenic agents in a rat model of TLE.

Pharmacological characterisation of anticonvulsant effects elicited by erythrartine.

OBJECTIVES: The erythrinan alkaloids erythravine and 11α-hydroxy-erythravine from Erythrina verna (Vell.) have been extensively investigated for their anxiolytic and anticonvulsant effects. Both are structurally similar to the erythrartine that also exhibit anxiolytic effects, but there is no report on its anticonvulsant potential. Since some anxiolytic drugs can be useful in the management of epileptic seizures, we investigated whether erythrartine could prevent seizures induced by different chemoconvulsants. METHODS: Experiments were performed using different concentrations of erythrartine injected via intracerebroventricular in rats submitted to pilocarpine, kainic acid, pentylenetetrazol or picrotoxin-induced seizures. Moreover, the rotarod test was performed to verify the effects of erythrartine on animal motor coordination. RESULTS: Our data showed for the first time that erythrartine prevented the occurrence of seizures induced by all of the chemoconvulsants tested and did not affect locomotor performance neither produced sedative effect on animals. CONCLUSION: Obtained results validate the ethnopharmacological significance of E. verna and provide new information on erythrartine, another erythrinian alkaloid of biotechnological and medicinal interest.

Valeriana officinalis extract and 7-Nitroindozole ameliorated seizure behaviours, and 7-Nitroindozole reduced blood pressure and ECG parameters in pentylenetetrazole-kindled rats.

Nitric oxide (NO) is known to be a neuromodulator with dual proconvulsive and anticonvul- sive action. Valeriana officinalis (VAL) was previously believed to be antiepileptic, but is today known as a sedative and sleep regulator. Seizures may be associated with abnormal electrocardio- graphic changes and cardiac dysfunction arising from epilepsy may be related with neuronal nitric oxide (nNO). This study was aimed to investigate the effects of the neuronal nitric oxide synthase (nNOS) inhibitor 7-Nitroindazole (7-NI) and VAL on seizure behaviours and electrocar- diographic parameters in the pentylentetrazole (PTZ)-kindled seizure model. Wistar rats were randomised into saline control, PTZ-kindled, 7-NI, VAL and VAL+PTZ, 7-NI+PTZ and VAL+7-NI+PTZ groups. Latency, stage, frequency of seizures, blood pressure (BP), heart rate (HR) and corrected QT (QTc) values were evaluated. Frequency and stage of seizures, BP and HR increased, while seizure latency decreased and QTc was prolonged in the PTZ-kindled group. 7-NI and VAL had no effects on BP and HR variables under normal conditions, but ameliorated the seizure stage and frequency of seizures. 7-NI treatment also resulted in a reduction of the increased BP and prolonged QTc values observed in PTZ-kindled rats. Considering these results, QTc prolongation may be used as a predictor for recurrent seizures. 7-NI and VAL exhibited different effects on seizures and ECG variables. 7-NI shows potential as an anticonvulsant drug agent in epileptic patients with cardiac dysfunctions and those additional studies including in-vivo experiments are essential.

Effect of Curcuma zedoaria hydro-alcoholic extract on learning, memory deficits and oxidative damage of brain tissue following seizures induced by pentylenetetrazole in rat.

BACKGROUND: Previous studies have shown that seizures can cause cognitive disorders. On the other hand, the Curcuma zedoaria (CZ) has beneficial effects on the nervous system. However, there is little information on the possible effects of the CZ extract on seizures. The aim of this study was to investigate the possible effects of CZ extract on cognitive impairment and oxidative stress induced by epilepsy in rats. METHODS: Rats were randomly divided into different groups. In all rats (except the sham group), kindling was performed by intraperitoneal injection of pentylenetetrazol (PTZ) at a dose of 35 mg/kg every 48 h for 14 days. Positive group received 2 mg/kg diazepam + PTZ; treatment groups received 100, 200 or 400 mg/kg CZ extract + PTZ; and one group received 0.5 mg/kg flumazenil and CZ extract + PTZ. Shuttle box and Morris Water Maze tests were used to measure memory and learning. On the last day of treatments PTZ injection was at dose of 60 mg/kg, tonic seizure threshold and mortality rate were recorded in each group. After deep anesthesia, blood was drawn from the rats' hearts and the hippocampus of all rats was removed. RESULTS: Statistical analysis of the data showed that the CZ extract significantly increased the tonic seizure threshold and reduced the pentylenetetrazol-induced mortality and the extract dose of 400 mg/kg was selected as the most effective dose compared to the other doses. It was also found that flumazenil (a GABAA receptor antagonist) reduced the tonic seizure threshold compared to the effective dose of the extract. The results of shuttle box and Morris water maze behavioral tests showed that memory and learning decreased in the negative control group and the CZ extract treatment improved memory and learning in rats. The CZ extract also increased antioxidant capacity, decreased MDA and NO in the brain and serum of pre-treated groups in compared to the negative control group. CONCLUSION: It is concluded that the CZ extract has beneficial effects on learning and memory impairment in PTZ-induced epilepsy model, which has been associated with antioxidant effects in the brain or possibly exerts its effects through the GABAergic system.

Screening of Some Novel 4, 5 Disubstituted 1, 2, 4-Triazole-3-thiones for Anticonvulsant Activity.

OBJECTIVE: In the present study, we synthesized fifteen 4, 5-disubstituted 1, 2, 4-triazol- 3-thione derivatives and evaluated for anticonvulsant activity with neurotoxicity determination. METHODS: The synthesized compounds were characterized using FTIR, 1H-NMR and MS. The molecular docking study was also performed to study the interactions of compounds with LYS329 residue of gamma amino butyric acid aminotransferase (GABA-AT) using Autodock 4.2 software. The anticonvulsant activity was assessed by maximal electroshock (MES) test and subcutaneous pentylenetetrazol (scPTZ) tests. The neurotoxicity was assessed by rotarod ataxia test. RESULTS: In MES test, compounds 5a, 8a and 9a were found active at 100 mg/kg and five compounds were found active at 300 mg/kg dose after 1 hr of administration. After 4 hr of drug administration, only two compounds 8a and 9a exhibited protection at 100 mg/kg. In scPTZ test, three compounds 2a, 6a and 8a were found active at 100 mg/kg and 7a was active at 300 mg/kg after 1 hr of test drug administration. Most of the compounds were found active in MES test with 8a and 9a being the most active among all. In docking study, 2a was found to be best compound based on the binding energy of -6.5 kcal/mol and estimated inhibition constant of 17.2 µM. CONCLUSION: Majority of synthesized compounds were found active in MES test, whereas only few were found to possess anti scPTZ activity. Among all compounds, only 14a caused motor coordination impairment in rotarod ataxia test at 300 mg/kg 1 hr duration.

Antiepileptic Effects of Protein-Rich Extract from Bombyx batryticatus on Mice and Its Protective Effects against H2O2-Induced Oxidative Damage in PC12 Cells via Regulating PI3K/Akt Signaling Pathways.

Bombyx batryticatus is a known traditional Chinese medicine (TCM) utilized to treat convulsions, epilepsy, cough, asthma, headaches, and purpura in China for thousands of years. This study is aimed at investigating the antiepileptic effects of protein-rich extracts from Bombyx batryticatus (BBPs) on seizure in mice and exploring the protective effects of BBPs against H2O2-induced oxidative stress in PC12 cells and their underlying mechanisms. Maximal electroshock-induced seizure (MES) and pentylenetetrazole- (PTZ-) induced seizure in mice and the histological analysis were carried out to evaluate the antiepileptic effects of BBPs. The cell viability of PC12 cells stimulated by H2O2 was determined by MTT assay. The apoptosis and ROS levels of H2O2-stimulated PC12 cells were determined by flow cytometry analysis. Furthermore, the levels of malondialdehyde (MDA), superoxide dismutase (SOD), lactate dehydrogenase (LDH), and glutathione (GSH) in PC12 cells were assayed by ELISA and expressions of caspase-3, caspase-9, Bax, Bcl-2, PI3K, Akt, and p-Akt were evaluated by Western blotting and quantitative real-time polymerase chain reaction (RT-qPCR) assays. The results revealed that BBPs exerted significant antiepileptic effects on mice. In addition, BBPs increased the cell viability of H2O2-stimulated PC12 cells and reduced apoptotic cells and ROS levels in H2O2-stimulated PC12 cells. By BBPs treatments, the levels of MDA and LDH were reduced and the levels of SOD and GSH-Px were increased in H2O2-stimulated PC12 cells. Moreover, BBPs upregulated the expressions of PI3K, Akt, p-Akt, and Bcl-2, whereas they downregulated the expressions of caspase-9, caspase-3, and Bax in H2O2-stimulated PC12 cells. These findings suggested that BBPs possessed potential antiepileptic effects on MES and PTZ-induced seizure in mice and protective effects on H2O2-induced oxidative stress in PC12 cells by exerting antioxidative and antiapoptotic effects via PI3K/Akt signaling pathways.

Exploratory EEG studies for the assessment of neurological liabilities in conscious dogs in early drug development.

INTRODUCTION: Convulsions in toxicology studies can be the first indication of seizure liability. Drug levels during convulsions are not usually evaluated. This, and exposure variability after oral administration, complicates estimation of safety margins. The electroencephalogram (EEG) enables symptoms to be attributed to seizures and to collect samples during epileptiform activity without clinical convulsion. We evaluated an EEG-study design for optimized detection of neurological symptoms. Additionally, we assessed whether EEG- based anticonvulsive treatment is feasible, to prevent progression to convulsions and if dogs have higher sensitivity towards neurological symptoms than non-human-primates. METHODS: Three compounds that previously were tested in non-human-primates were selected to evaluate the dog EEG-study design. Two substances were administered in escalating intravenous doses; the third was given as single oral dose. Per compound, one male and one female telemetered dog were evaluated; males also had cerebrospinal-fluid-ports. Drug levels, video-EEG and clinical symptoms were evaluated and compared to previous studies. RESULTS: While similar neurological symptoms were induced, intravenous administration reduced experimental time compared to standard toxicology studies. EEG analysis could link animal behavior to seizures but did not allow convulsion prevention. This was due to artefacts and the short latency between onset of epileptiform EEG activity and clinical convulsions. Free plasma concentrations during convulsions were comparable between dogs and non-human-primates. DISCUSSION: The findings suggest that infusion studies provide a possibility to investigate neurological adverse effects in few animals in a short time period. For candidates with a high risk for seizures, such studies can guide dose selection for longer regulatory studies and improve safety margin definition.

Early α-linolenic acid exposure to embryo reduces pentylenetetrazol-induced seizures in zebrafish larva.

Over the past few years, there has been a tremendous increase in interest of general population toward food-based therapies for management of chronic clinical conditions due to their lesser adverse effects with prolonged use over pharmacotherapies. Foods enriched with omega-3 fatty acids have shown some promising results in case of epilepsy. The present study was envisioned to investigate the effect of early exposure of α-linolenic acid (ALA), an essential omega-3 fatty acid in developing zebrafish (Danio rerio) embryos toward pentylenetetrazol (PTZ)-induced seizure susceptibility. The healthy wild-type zebrafish embryos were incubated in system water or system water containing different ALA concentrations (1-20 µM) till 7 dpf (days post fertilization). Each larva at 7 dpf was placed in 8 mM PTZ solution and seizure event was recorded. ALA incubation at 10 µM and 20 µM concentrations showed a dose-dependent reduction in PTZ-mediated hyperactive responses in larvae indicated by a marked decrease in total distance travelled and speed, as compared to vehicle control. Furthermore, both the treated groups showed increase in the latency to PTZ-induced clonus-like seizures in larvae, as compared to vehicle control. ALA incubated larvae at 10 µM and 20 µM concentrations also showed a significant reduction in c-fos mRNA level. A marked increase in the level of ALA and docosahexaenoic acid was also observed in the larvae incubated at highest effective concentration of ALA. The present study concluded that embryonic exposure of ALA reduced PTZ-induced seizures in zebrafish larva.

Pycnogenol Protects against Pentylenetetrazole-Induced Oxidative Stress and Seizures in Mice.

BACKGROUND: Epilepsy is one of the most common and severe brain disorders in the world, characterized by recurrent spontaneous seizures due to an imbalance between cerebral excitability and inhibition. Oxidative stress is a biochemical state in which reactive oxygen species are generated and associated with various diseases including epilepsy. Pycnogenol, a polyphenol obtained from the pine tree and has antioxidant & anti-inflammatory activity. So, the aim of the study was to evaluate the effect of Pycnogenol on pentylenetetrazole (PTZ)-induced seizures in mice. METHODS: The mice of swiss strain each weighing 18-30g were used. Pycnogenol (50&100mg/kg) was suspended in carboxymethyl cellulose in saline and administered orally. Diazepam (1mg/kg, i.p) was used as a standard drug. The anticonvulsant effects of the drugs were measured using PTZ and cognitive behaviour was also assessed. The biochemical estimation was done by measuring Thiobarbituric acid, Superoxide dismutase, Catalase, and reduced glutathione followed by the histopathological study. RESULT: Pycnogenol 50 & 100mg/kg showed a significant increase in latency to PTZ-induced seizures, decrease in duration and frequency of convulsions compared to control animals; however, the effects were dose-dependent and were more significant at a higher dose. No impairment in cognitive functions like memory and muscle relaxant was observed following pycnogenol 50 & 100 mg/kg. The effect of Pycnogenol on biochemical parameter was found to be significant. It significantly (p<0.01) decreases the level of TBARS and increases the levels of SOD, catalase, and GSH in the brain tissue. The histopathological evaluation showed less neuronal degeneration in the brain due to PTZ-induced seizures in comparison to control group. CONCLUSION: Thus pycnogenol has a protective approach towards convulsion and can be included as an adjuvant therapy with antiepileptic drugs.

Inhibitory effects of chronic administration of vitamin D3 on pentylenetetrazole-induced seizures in mice.

Vitamin D3 as a neuroactive steroid hormone plays an important role in the nervous system. Recent clinical and experimental studies have shown an association between vitamin D-related disorders and epilepsy. Therefore, this study was designed to examine the effects of chronic administration of vitamin D3 on pentylenetetrazole (PTZ)-induced seizure in mice. This interventional study was conducted on 120 mice in 12 groups. Two control groups acutely and chronically received a mixture of almond oil and paraffin; three groups were acutely given vitamin D3 at doses of 2000, 4000 and 6000 IU/kg; three groups chronically received vitamin D3 with similar doses for two weeks and two groups chronically and acutely received a sub-effective dose of vitamin D3 and diazepam. Slow intravenous infusion of PTZ (5 mg/mL) was performed at a constant rate (0.3 mL/min) via an infusion pump to induce clonic and tonic seizures. Acute injection of different concentrations of vitamin D3 (2000, 4000 and 6000 IU/kg i.p.) did not significantly increase a seizure threshold. However, a seizure threshold in the groups chronically treated with 4000, and 6000 IU/kg of vitamin D3 was significantly higher than that in the control group (P < 0.001). Moreover, a combination of the sub-effective dose of vitamin D3 (2000 IU/kg) and diazepam (0.1 mg/kg) significantly increased seizure threshold. Our findings suggest that administration of vitamin D supplement can be considered as a potential add-on treatment in seizure and due to the vitamin D deficiency results from the long-term use of most anti-seizure drugs, this supplementation becomes more important.

Assessing seizure liability using multi-electrode arrays (MEA).

The purpose of these studies was to develop ex vivo tissue-based and in vitro cell-based assays using multi-electrode array (MEA) technology to predict seizure liability at the early stage of preclinical studies. Embryonic rat hippocampal neurons and adult rat hippocampal slices were used in these studies. Spontaneous activity in cultured neurons and evoked field potentials in hippocampal brain slices were recorded using MEA technology. Six seizurogenic compounds bicuculline, pentylenetetrazole, picrotoxin, gabazine, 4-Aminopyridine and BMS-A increased field potential area and peak number in brain slices and spontaneous spike activity in hippocampal neurons. Physostigmine, another seizurogenic compound, had no effect on brain slices at lower concentrations (0.1, 1, and 10 µM), and mildly increased field potential area at 100 µM. However, physostigmine induced multiple peaks in evoked field potential starting at 10 µM. Physostigmine showed greater potency in the cultured neuron assay, and increased spike rates in the nanomolar range. Two seizurogenic compounds, BMS-B and BMS-C increased the spontaneous activity in hippocampal neurons, but did not increase area and peak number of field potentials in brain slices. These findings suggest that MEA technology and rat hippocampal brain slices or rat embryonic hippocampal neurons, may be useful as early, predictive in vitro assays for seizure liability.

High-throughput brain activity mapping and machine learning as a foundation for systems neuropharmacology.

Technologies for mapping the spatial and temporal patterns of neural activity have advanced our understanding of brain function in both health and disease. An important application of these technologies is the discovery of next-generation neurotherapeutics for neurological and psychiatric disorders. Here, we describe an in vivo drug screening strategy that combines high-throughput technology to generate large-scale brain activity maps (BAMs) with machine learning for predictive analysis. This platform enables evaluation of compounds' mechanisms of action and potential therapeutic uses based on information-rich BAMs derived from drug-treated zebrafish larvae. From a screen of clinically used drugs, we found intrinsically coherent drug clusters that are associated with known therapeutic categories. Using BAM-based clusters as a functional classifier, we identify anti-seizure-like drug leads from non-clinical compounds and validate their therapeutic effects in the pentylenetetrazole zebrafish seizure model. Collectively, this study provides a framework to advance the field of systems neuropharmacology.

Corpus callosum low-frequency stimulation suppresses seizures in an acute rat model of focal cortical seizures.

OBJECTIVE: Low-frequency fiber-tract stimulation has been shown to be effective in treating mesial temporal lobe epilepsies through activation of the hippocampal commissure in rodents and human patients. The corpus callosum is a major pathway connecting the two hemispheres of the brain; however, few experiments have documented corpus callosum stimulation. The objective is to determine the efficacy of corpus callosum stimulation at low frequencies to suppress cortical seizures. METHODS: 4-Aminopyridine was injected in the primary motor cortex of 24 rats under anesthesia. Recording electrodes were placed in the contralateral motor cortex and hippocampus. Three pairs of stimulating electrodes were inserted into the corpus callosum along its longitudinal axis. Local field potentials were recorded 1 hour before, during, and after stimulation to determine the effect of stimulation on seizure duration. Stimulation was delivered from each pair of electrodes independently in separate experiments. Furthermore, electrical stimulation was applied to the region of the corpus callosum with the highest degree of innervation of the seizure focus to compare the efficacy of different stimulation frequencies (1-30 Hz) on seizure suppression. RESULTS: Corpus callosum stimulation was effective at suppressing seizures at 10 Hz by 76% (P < 0.05, n = 5) and at 20 Hz by 95% (P < 0.0001, n = 14). Stimulation at frequencies of 1 and 30 Hz did not have a significant effect on reducing the total time spent seizing (P > 0.9999, n = 5). Furthermore, stimulation was only effective at suppressing seizures when the pair of electrodes was placed within the section of corpus callosum containing fibers innervating the seizure focus. Secondarily generalized seizures in the hippocampus were eliminated when seizures in the cortical focus were suppressed. SIGNIFICANCE: Low-frequency fiber-tract stimulation of the corpus callosum suppresses both cortical and cortically induced hippocampal seizures in an acute model of focal cortical seizures. The stimulation paradigm is selective, as it is only effective when targeted to specific regions of the corpus callosum that project maximally to cortical regions generating the seizure activity. Selective placement of stimulation electrodes along the corpus callosum could be used as a patient-specific treatment for cortical epilepsies.

Deep brain stimulation probing performance is enhanced by pairing stimulus with epileptic seizure.

The unpredictability of spontaneous and recurrent seizures significantly impairs the quality of life of patients with epilepsy. Probing neural network excitability with deep brain electrical stimulation (DBS) has shown promising results predicting pathological shifts in brain states. This work presents a proof-of-principal that active electroencephalographic (EEG) probing, as a seizure predictive tool, is enhanced by pairing DBS and the electrographic seizure itself. The ictogenic model used consisted of inducing seizures by continuous intravenous infusion of pentylenetetrazol (PTZ - 2.5 mg/ml/min) while a probing DBS was delivered to the thalamus (TH) or amygdaloid complex to detect changes prior to seizure onset. Cortical electrophysiological recordings were performed before, during, and after PTZ infusion. Thalamic DBS probing, but not amygdaloid, was able to predict seizure onset without any observable proconvulsant effects. However, previously pairing amygdaloid DBS and epileptic polyspike discharges (day-1) elicited distinct preictal cortically recorded evoked response (CRER) (day-2) when compared with control groups that received the same amount of electrical pulses at different moments of the ictogenic progress at day-1. In conclusion, our results have demonstrated that the pairing strategy potentiated the detection of an altered brain state prior to the seizure onset. The EEG probing enhancement method opens many possibilities for both diagnosis and treatment of epilepsy.

A Caenorhabditis elegans assay of seizure-like activity optimised for identifying antiepileptic drugs and their mechanisms of action.

BACKGROUND: Epilepsy affects around 1% of people, but existing antiepileptic drugs (AEDs) only offer symptomatic relief and are ineffective in approximately 30% of patients. Hence, new AEDs are sorely needed. However, a major bottleneck is the low-throughput nature of early-stage AED screens in conventional rodent models. This process could potentially be expedited by using simpler invertebrate systems, such as the nematode Caenorhabditis elegans. NEW METHOD: Head-bobbing convulsions were previously reported to be inducible by pentylenetetrazol (PTZ) in C. elegans with loss-of-function mutations in unc-49, which encodes a GABAA receptor. Given that epilepsy-linked mutations in human GABAA receptors are well documented, this could represent a clinically-relevant system for early-stage AED screens. However, the original agar plate-based assay is unsuited to large-scale screening and has not been validated for identifying AEDs. Therefore, we established an alternative streamlined, higher-throughput approach whereby mutants were treated with PTZ and AEDs via liquid-based incubation. RESULTS: Convulsions induced within minutes of PTZ exposure in unc-49 mutants were strongly inhibited by the established AED ethosuximide. This protective activity was independent of ethosuximide's suggested target, the T-type calcium channel, as a null mutation in the worm cca-1 ortholog did not affect ethosuximide's anticonvulsant action. COMPARISON WITH EXISTING METHOD: Our streamlined assay is AED-validated, feasible for higher throughput compound screens, and can facilitate insights into AED mechanisms of action. CONCLUSIONS: Based on an epilepsy-associated genetic background, this C. elegans unc-49 model of seizure-like activity presents an ethical, higher throughput alternative to conventional rodent seizure models for initial AED screens.