PTZ-kindled Model Research Articles

The mechanism of mitochondrial autophagy regulating Clathrin-mediated endocytosis in epilepsy.

The objective of this study is to determine whether inhibition of mitophagy affects seizures through Clathrin-mediated endocytosis (CME). Pentylenetetrazol (PTZ) was intraperitoneally injected daily to establish a chronic PTZ-kindled seizure. The Western blot (WB) was used to compare the differences in Parkin protein expression between the epilepsy group and the control group. Immunofluorescence was used to detect the expression of MitoTracker and LysoTracker. Transferrin-Alexa488 (Tf-A488) was injected into the hippocampus of mice. We evaluated the effect of 3-methyladenine (3-MA) on epilepsy behavior through observation in PTZ-kindled models. The methylated derivative of adenine, known as 3-MA, has been extensively utilized in the field of autophagy research. The transferrin protein is internalized from the extracellular environment into the intracellular space via the CME pathway. Tf-A488 uses a fluorescent marker to track CME. Western blot showed that the expression of Parkin was significantly increased in the PTZ-kindled model (p < 0.05), while 3-MA could reduce the expression (p < 0.05). The fluorescence uptake of MitoTracker and LysoTracker was increased in the primary cultured neurons induced by magnesium-free extracellular fluid (p < 0.05); the fluorescence uptake of Tf-A488 was significantly decreased in the 3-MA group compared with the control group (p < 0.05). Following hippocampal injection of Tf-A488, both the epilepsy group and the 3-MA group exhibited decreased fluorescence uptake, with a more pronounced effect observed in the 3-MA group. Inhibition of mitophagy by 3-MA from day 3 to day 9 progressively exacerbated seizure severity and shortened latency. It is speculated that the aggravation of seizures by 3-MA may be related to the failure to remove damaged mitochondria in time and effectively after inhibiting mitochondrial autophagy, affecting the vesicle endocytosis function of CME and increasing the susceptibility to epilepsy. Abnormal mitophagy was observed in a chronic pentylenetetrazol-induced seizure model and a Mg2+-free-induced spontaneous recurrent epileptiform discharge model. A fluorescent transferrin marker was utilized to track clathrin-mediated endocytosis. Using an autophagy inhibitor (3-methyladenine) on primary cultured neurons, we discovered that inhibition of autophagy led to a reduction in fluorescent transferrin uptake, while impairing clathrin-mediated endocytosis function mediated by mitophagy. Finally, we examined the effects of 3-methyladenine in an animal model of seizures showing that it exacerbated seizure severity. Ultimately, this study provides insights into potential mechanisms through which mitophagy regulates clathrin-mediated endocytosis in epilepsy.

Can endoplasmic reticulum stress observed in the PTZ-kindling model seizures be prevented with TUDCA and 4-PBA?

Epilepsy is a chronic neurological disease with recurrent seizures. Increasing evidence suggests that endoplasmic reticulum (ER) stress may play a role in the pathogenesis of epilepsy. We aimed to investigate the effects of Tauroursodeoxycholic acid (TUDCA) and 4-phenyl-butyric acid (4-PBA), which are known to suppress ER stress, on developed seizures in terms of markers of ER stress, oxidative stress, and apoptosis. The pentylenetetrazole (PTZ) kindling model was induced in Wistar albino rats (n = 48) by administering 35 mg/kg PTZ intraperitoneally (I.P.) every other day for 1 month. TUDCA and 4-PBA were administered via I.P. at a dose of 500 mg/kg dose. ER stress, apoptosis, and oxidative stress were determined in the hippocampus tissues of animals in all groups. Immunohistochemistry, qRT-PCR, ELISA, and Western Blot analyzes were performed to determine the efficacy of treatments. Expressions of ATF4, ATF6, p-JNK1/2, Cleaved-Kaspase3, and Caspase12 significantly increased in PTZ-kindled seizures compared to the control group. Increased NOX2 and MDA activity in the seizures were measured. In addition, stereology analyzes showed an increased neuronal loss in the PTZ-kindled group. qRT-PCR examination showed relative mRNA levels of CHOP. Accordingly, TUDCA and 4-PBA treatment suppressed the expressions of ATF4, ATF6, Cleaved-Caspase3, Kaspase12, NOX2, MDA, and CHOP in TUDCA + PTZ and 4-PBA + PTZ groups. ER stress-induced oxidative stress and apoptosis by reducing neuronal loss and degeneration were also preserved in these groups. Our data show molecularly that TUDCA and 4-PBA treatment can suppress the ER stress process in epileptic seizures.

Dietary Zinc Differentially Regulates the Effects of the GPR39 Receptor Agonist, TC-G 1008, in the Maximal Electroshock Seizure Test and Pentylenetetrazole-Kindling Model of Epilepsy.

The G-protein coupled receptor 39 (GPR39) is gaining increasing attention as a target for future drugs, yet there are gaps in the understanding of its pharmacology. Zinc is an endogenous agonist or an allosteric modulator, while TC-G 1008 is a synthetic, small molecule agonist. Zinc is also a positive allosteric modulator for the activity of TC-G 1008 at GPR39. Activation of GPR39 by TC-G 1008 facilitated the development of epileptogenesis in the pentylenetetrazole (PTZ)-induced kindling model of epilepsy. Congruently, TC-G 1008 decreased the seizure threshold in the maximal electroshock seizure threshold (MEST) test. Here, we investigated the effects of TC-G 1008 under the condition of zinc deficiency. Mice were fed a zinc-adequate diet (ZnA, 50 mg Zn/kg) or a zinc-deficient diet (ZnD, 3 mg Zn/kg) for 4 weeks. Following 4 weeks of dietary zinc restriction, TC-G 1008 was administered as a single dose and the MEST test was performed. Additional groups of mice began the PTZ-kindling model during which TC-G 1008 was administered repeatedly and the diet was continued. TC-G 1008 administered acutely decreased the seizure threshold in the MEST test in mice fed the ZnD diet but not in mice fed the ZnA diet. TC-G 1008 administered chronically increased the maximal seizure severity and the percentage of fully kindled mice in those fed the ZnA diet, but not in mice fed the ZnD diet. Our data showed that the amount of zinc in a diet is a factor contributing to the effects of TC-G 1008 in vivo.

Downregulatory effect of miR-342-3p on epileptogenesis in the PTZ-kindling model.

Epileptogenesis is a process that results in neurons firing abnormally, causing seizures. Increasing evidence has shown that miRNAs expressed in the epileptic hippocampus are involved in epileptogenesis. We demonstrated the expression changes of miRNAs that may be effective in epileptogenesis in silico analysis in the kindling model created with Pentylenetetrazole (PTZ). Thus, we aimed to identify the target genes responsible for epileptogenesis. Fifteen male Wistar-albino rats (200-230g) were randomly divided into two groups control (n = 6) and PTZ (n = 9). The control group received 0.5ml saline, and the PTZ group (35mg/kg i.p.) intraperitoneally (i.p.) (11 times, every other day) to induce tonic-clonic seizures. Seizures were observed and scored 30min after PTZ injection. After the last dose of PTZ (75mg/kg) administration, the hippocampus tissues of the rats were removed by anesthesia. Analysis of miRNAs was performed with the Affymetrix gene chip miRNA sequence (728 miRNA) and confirmed by the Real-Time Polymerase Chain Reaction (Real-Time PCR) method (29 miRNAs). We evaluated the expression change of the target gene of miRNA, whose expression change was detected using in silico analysis, by q-RT PCR. Eight miRNAs with changes in expression were detected. Of these miRNAs, miR-342-p was downregulated in the PTZ group and was statistically significant (p < 0.005). Ultimately, we determined that the target gene of miR-342-p is a metabotropic glutamate receptor 2 (GRM2) and that GRM2 expression is upregulated. Downregulation of miR-342-3p in the PTZ kindling model may result in the upregulation of GRM2.

Thiamine alleviates cognitive impairment and epileptogenesis by relieving brain inflammation in PTZ-induced kindling rat model

ABSTRACT Objective Epileptogenesis, the process by which the brain becomes epileptic, is related to neuroinflammation, hyperexcitability cognitive deficits. Evidence suggests that improving brain inflammation can inhibit the epileptogenesis process and help the emergence of new drugs for the treatment of epilepsy. Therefore, the PTZ kindling model of epilepsy was utilized to assess the neuroprotective role of thiamine in epileptogenesis. Methods Male rats were exposed to PTZ-induced kindling and pretreated with low thiamine (25 mg/kg) or high thiamine (50 mg/kg). Cyclooxygenase (COX-1 and COX-2), interleukin 1-beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and nuclear factor-κB (NF-κB) concentrations in the brain were analyzed using biochemical assays. Cognitive function was evaluated using the passive avoidance test. Results Thiamine ameliorated epileptogenesis and enhanced the rats’ performance in the passive avoidance test. Also, thiamine significantly decreased the level of neuroinflammatory mediators in the brain induced by PTZ. Conclusion These results provide evidence that thiamine alleviates PTZ-induced neuroinflammation and cognitive impairments.

Pentylenetetrazole-induced kindling rat model: miR-182 and miR-27b-3p mediated neuroprotective effect of thymoquinone in the hippocampus

ABSTRACT Objectives: Epilepsy is a neurological disease that pathologically affects brain functions. The epileptic hippocampus has modified microRNA(miRNA) levels. Therefore, we aimed to evaluate the neuroprotective effect of thymoquinone (TQ) in PTZ-induced epilepsy and to demonstrate the overlap between miRNA and mRNA expression profiles. Methods: Male adult Wistar albino rats (200–230 g, n = 20) were divided into three groups as control (n = 6), PTZ (n = 7), and TQ + PTZ (n = 7). The PTZ kindling model was created by injecting PTZ in sub convulsive doses to rats on days 1, 3, 5, 8, 10, 12, 15, 17, 19, 22, and 24 of the study into animals. Clonic and tonic seizures were induced by injecting a convulsive dose of PTZ on day 26 of the study. Rats in the TQ+PTZ group were treated by oral gavage with a 20 mg/kg TQ 2 h before each PTZ injection. The rats in the control group were treated with 0.5 ml saline. Seizure severity was evaluated with the Racine scale. The genes and signaling pathways targeted by miRNAs were determined by bioinformatics analysis. Results: In the rat hippocampus, mature 728 miRNAs were analyzed by microarray and the nine miRNA were verified by quantitative Real-Time PCR. rno-miR-182 and rno-miR-27b-3p were up-regulated in the PTZ group and down-regulated in the TQ + PTZ group. Discussion: In the PTZ kindling epilepsy model, the expression of these two miRNAs was regulated by TQ and exerted a neuroprotective effect by controlling the activities of target genes.

The effect of a novel anticonvulsant chemical Q808 on gut microbiota and hippocampus neurotransmitters in pentylenetetrazole-induced seizures in rats

BackgroundThe gut microbiota can modulate brain function and behavior and is increasingly recognized as an important factor in mediating the risk of epilepsy and the effects of seizure interventions. Drug therapy is one of the factors that influence the composition of the intestinal microbiota. Q808 is an innovative chemical with strong anticonvulsant activity and low neurotoxicity. However, studies evaluating the effect of Q808 on gut microbial communities are lacking. In this study, we aimed to evaluate the anticonvulsant activity of Q808 on a pentylenetetrazol (PTZ)—induced seizure model and analyze and compare the intestinal microbiota composition of non-PTZ vehicle control group, the PTZ-induced seizure model rats with and without Q808, through 16S rDNA sequencing. Neurotransmitter levels in the hippocampus were quantitatively estimated using HPLC–MS.ResultsThe results suggest that Q808 effectively alleviates seizures in chronic PTZ-kindled model rats. Additionally, based on the analyzed abundance of the gut microbiota, dysbacteriosis of model rats was found to be corrected after Q808 treatment at the phylum level. The unique bacterial taxa (e.g., Lactobacillus) that are associated with acetylcholine production, were significantly increased. Several short-chain fatty acids (SCFAs)-producing bacteria, including Roseburia, Alloprevptella, Prevotellaceae_NK3B31_group, Prevotellaceae_UCG-001, and Prevotella_9, were enriched. In the hippocampus, the contents of acetylcholine increased, whereas the levels of 3-methoxytyramine, glutamine, and 5-hydroxyindole acetic acid (5-HIAA) decreased after Q808 treatment.ConclusionsThis study demonstrates that Q808 can be used to remodel the dysbiosis of the gut microbiome and influence neurotransmitter levels in the hippocampus of PTZ-induced seizure model rats. We hope that these novel findings prompt further research on the interaction between gut microbiota and seizures and the mechanism of Q808.

Anticonvulsant and antioxidant effect of hydroalcoholic extract of Valeriana wallichii rhizomes in acute and chronic models of epilepsy in albino rats

Oxidative stress plays an important role in the aetiology of seizure-induced neuronal death. The present study was aimed to evaluate the anticonvulsant and antioxidant activity of hydroalcoholic extract of Valeriana wallichii (HAEVW) rhizomes in albino rats. Flavonoids possess a potent anticonvulsant effect through modulation of GABA/BDZ receptor and neuroprotective effect by regulating oxidative stress in PTZ-induced convulsion. Total phenolics and flavonoids content of HAEVW rhizomes were measured and the anticonvulsant activity was evaluated using PTZ and MES induced convulsion models. The in-vivo antioxidant activity was evaluated using the PTZ-kindled model by estimating brain malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), glutathione (GSH), and nitric oxide (NO). It was observed that the flavonoids content of the extract was 80% of the total phenolics content and HAEVW (200 and 400 mg/kg) showed a significant (P <0.01) delay in onset, decrease in duration of tonic convulsion and hind limb tonic extension (HLTE) in PTZ and MES induced convulsion respectively. In the PTZ-kindling model, HAEVW rhizomes (400 mg/kg) restored the MDA and all antioxidant parameters to normal except NO. It was concluded that flavonoids of rhizomes of V. wallichii may be responsible for the anticonvulsant and antioxidant activity of the extract.

Inflammation-Associated HMGB1, α7nAChR, Kir6.2 and Beclin1 Immunoreactivity in PTZ-Kindled Model Seizures in Rats

Inflammation-Associated HMGB1, α7nAChR, Kir6.2 and Beclin1 Immunoreactivity in PTZ-Kindled Model Seizures in Rats

Metabolic Alterations Predispose to Seizure Development in High-Fat Diet-Treated Mice: the Role of Metformin.

The link between epilepsy and type 2 diabetes (T2DM) and/or metabolic syndrome (MetS) has been poorly investigated. Therefore, we tested whether a high-fat diet (HFD), inducing insulin-resistant diabetes and obesity in mice, would increase susceptibility to develop generalized seizures induced by pentylentetrazole (PTZ) kindling. Furthermore, molecular mechanisms linked to glucose brain transport and the effects of the T2DM antidiabetic drug metformin were also studied along with neuropsychiatric comorbidities. To this aim, two sets of experiments were performed in CD1 mice, in which we firstly evaluated the HFD effects on some metabolic and behavioral parameters in order to have a baseline reference for kindling experiments assessed in the second section of our protocol. We detected that HFD predisposes towards seizure development in the PTZ-kindling model and this was linked to a reduction in glucose transporter-1 (GLUT-1) expression as observed in GLUT-1 deficiency syndrome in humans but accompanied by a compensatory increase in expression of GLUT-3. While we confirmed that HFD induced neuropsychiatric alterations in the treated mice, it did not change the development of kindling comorbidities. Furthermore, we propose that the beneficial effects of metformin we observed towards seizure development are related to a normalization of both GLUT-1 and GLUT-3 expression levels. Overall, our results support the hypothesis that an altered glycometabolic profile could play a pro-epileptic role in human patients. We therefore recommend that MetS or T2DM should be constantly monitored and possibly avoided in patients with epilepsy, since they could further aggravate this latter condition.

Effects of P-gpMAbNano-structured material nanoparticles on epilepsy and expression of SRY-related HMG Box 21 in epilepsy

SRY-related HMG box (SOX)21, one of the most highly expressed transcription inhibitors in the central nervous system (CNS), is involved in neurogenesis-related transcription and proliferation, which are associated with certain neurological disorders. However, it is the role of SOX21 in the pathogenesis of epilepsy remains unclear. In this study, our aim was to examine the expression and function of SOX21 in patients with temporal lobe epilepsy (TLE), as well as pentylenetetrazol (PTZ)-kindled rats, and to identify the possible roles of SOX21 in epileptogenesis. We found that SOX21 localized in neurons is upregulated, especially in TLE patients. SOX21 is present in the hippocampus or adjacent temporal cortex in the PTZ-kindled epileptic rat model. In addition, the P-gpMAbNano-structured material (PNM) nanoparticles carrying anti-epileptic drugs (AEDs) were injected into the epileptic model rats using an intravenous injection. The expression of tumor necrosis factor peptide in the rats was detected to verify whether the drug-carrying nanoparticles could bypass macrophages and reach the target for treatment. We also found an interaction between SOX21 and SOX2 in PTZ-kindled rats. These results indicate that SOX21 is mainly located in neurons and may regulate the pathogenesis of epilepsy, possibly in association with SOX2. Moreover, PNM nanoparticles equipped with AEDs can reach the target through macrophages in vivo, providing a new approach for the clinical treatment of epilepsy.

The role of REST/NRSF, TrkB and BDNF in neurobiological mechanisms of different susceptibility to seizure in a PTZ model of epilepsy

Global transcriptional disturbances are believed to play a major role in the course of epilepsy. Due to the high complexity, the neurobiological mechanisms underlying different susceptibility to seizure and epilepsy are not well known. A transcription factor called REST/NRSF (repressor element 1-silencing transcription factor/neuron-restrictive silencer factor) is believed to contribute to processes associated with seizure development. Its downstream genes, those encoding BDNF (brain-derived neurotrophic factor) and TrkB (BDNF receptor; tropomyosin receptor kinase B), are also thought to play a role. To verify this hypothesis, we used a PTZ kindling model of epilepsy and divided animals into groups according to their different susceptibility to seizure. The concentrations of REST/NRSF, BDNF, and TrkB protein and mRNA were measured in hippocampal homogenates.The level of REST/NRSF protein measured 24 h after the last PTZ injection was increased in animals resistant to kindling and was unchanged in groups of rats kindled after 5, 10 and 20 in.ections of PTZ. In contrast, TrkB protein concentration was enhanced in all kindled rats and was unchanged in the resistant rats. There were no changes in the protein concentration of BDNF in rats with different susceptibility to kindling; however, data from the combined kindled groups vs. the resistant group revealed an increased level of BDNF in resistant animals.In sum, the increased level of protein REST/NRSF in resistant animals may reflect its neuroprotective role against seizure development. The increased concentration of TrkB protein in kindled animals indicates its pivotal role in the process of epileptogenesis. We propose that in resistant rats, REST/NRSF could contribute to the prevention of TrkB activation related to seizures.

Expression of HIF-1α, TFRC-1 and TIM-2 Relative mRNA Levels in PTZ-Kindling Model of Epilepsy

Epilepsy is a neurological disease characterized by abnormal electrical activity and recurrent seizures in the central nervous system (CNS). Changes in hypoxia and iron metabolism can stimulate seizures through the CNS and cardiac system. Cardiovascular system disorders such as arrhythmias also accompany this process. It was aimed to examine the genes of TFRC-1 and TIM-2, which contribute to ion homeostasis by providing hypoxia inducible factor (HIF) -1 and intracellular iron flow, through the kindled model of pentylenetetrazol (PTZ). HIF-1α, TFRC-1 and TIM-2 gene expressions were investigated in both brain and heart tissue by RT-PCR method. As a result of the data, TIM-2 expression significantly decreased in the brain (p

Neuroprotective mechanisms of sildenafil and selenium in PTZ-kindling model: Implications in epilepsy

Epilepsy is one of the furthermost common neurodegenerative diseases affecting above 50 million individuals worldwide. The pathogenesis of epileptic seizures is not satisfactorily explored, and hence more effective anti-convulsive therapies are indispensable. Current study aimed to investigate the mechanisms of the potential neuroprotective effects of sildenafil/selenium on chemically-induced convulsions in mice.Kindling model was induced using pentylenetetrazol (PTZ; 35 mg/Kg, 11 doses, intraperitoneally, every other day). PTZ-insulted groups were treated intraperitoneally with sildenafil (20 mg/Kg), selenium (0.2 mg/Kg) or their combination; 30 min before PTZ administration.PTZ-kindled model showed a significant loss of neuronal cells concurrently with nitrative/oxidative stress and lipid peroxidation. This was associated with enhanced expression of inducible nitric oxide synthase (iNOS), hemeoxygenase-1 (HO-1) and vascular endothelial growth factor (VEGF) along with increased activity of thioredoxin reductase (TrxR) in hippocampal tissue. Individual treatment with sildenafil or selenium showed partial neuroprotection, simultaneously with lower hippocampal expression of 4-hydroneonenal (4-HNE), nitrotyrosine, iNOS and HO-1, yet without reaching normal levels. Sildenafil, but not selenium, enhanced the expression of VEGF and the endothelial cell marker CD34. The joint treatment with sildenafil and selenium preserved hippocampal neuronal count, improved kindling score, blunted lipid peroxides and nitrotyrosine levels, concomitantly with iNOS inhibition, normalization of TrxR activity and HO-1 expression, and evident neo-angiogenesis.Current study demonstrated the roles of several central signalling cascades in the sildenafil/selenium-evoked neuroprotection represented in, at least in part, amelioration of nitrative/oxidative stress alongside modulation of angiogenesis. Thus, sildenafil combined with selenium could be repurposed as a potential therapeutic regimen for delaying epilepsy progression.

Lacosamide Reduces Seizure Severity but Increases Seizure Frequency in PTZ-Kindled Rats

Abstract Objective: This study evaluated the anticonvulsant action of lacosamide (LCS), a novel drug that was recently approved for the treatment of partial or secondarily generalized seizures, using an animal model of generalized epilepsy induced by repetitive pentylenetetrazole (PTZ) administration in rats. The main goal was to evaluate the behavioral pattern of lacosamide action by classifying seizures according to a modi Racine-scale. Furthermore, the reproducibility of the win-PTZ kindling model of epilepsy, a recently described variant of the standard PTZ-kindling model, was also assessed. Methods: Adult male Wistar rats (n=16) were divided into two groups and underwent the win-PTZ-kindling protocol in two independent trials. After finishing the kindling procedure, all animals, which presented stage 5 seizures were tested for the anticonvulsant action of lacosamide at three different doses (3, 10, and 30 mg/kg). Results: The maximal severity of seizures decreased and the latency to stage 3-5 seizures increased when the animals were treated with lacosamide at a single dose of 10 mg/kg compared to saline pretreatment (p &lt; 0.05), both parameter reflecting an anticonvulsant action of the drug. Unfortunately, the number of stage 3-5 seizures also increased, but not significantly. The win-PTZ kindling model showed an adequate reproducibility between different trials, however, the number of fully kindled rats was lower than previously reported. Conclusions: Lacosamide showed a convincing anticonvulsant action in the win-PTZ kindling model of epilepsy by preventing the generalization of seizures. The win-PTZ kindling model was proved to be useful for studying epileptogenesis and the anticonvulsant action of drugs.

Effect of synaptic adhesion-like molecule 3 on epileptic seizures: Evidence from animal models.

Axonal sprouting and synaptic reorganization are the primary pathophysiological characteristics of epilepsy. Recent studies demonstrated that synaptic adhesion-like molecule 3 (SALM3) is highly expressed in the central nervous system and plays important roles in neurite outgrowth, branching, and axon guidance, mechanisms that are also observed in epilepsy. However, the expression of SALM3 in the epileptic brain and the effect of SALM3 in the pathogenesis of epilepsy remain unclear. The aims of this study were to investigate SALM3 expression in rat models of epilepsy and to explore the functional significance of SALM3 in epilepsy. We demonstrated that SALM3 was expressed at significantly higher levels in epileptic rats compared with controls. Inhibition of SALM3 by SALM3 shRNA inhibited status epilepticus in the acute stage of disease and decreased spontaneous recurrent seizures in the Lithium-pilocarpine model of chronic stages of epilepsy. Consistent with these findings, SALM3 shRNA significantly prolonged the latent period in the PTZ kindling model. Our study suggests that the overexpression of SALM3 might be associated with epileptogenesis and that selectively inhibiting SALM3 may have therapeutic potential in treating epilepsy.

Effectiveness of ketogenic diet in pentylenetetrazol-induced and kindling rats as well as its potential mechanisms

The effects and mechanisms of ketogenic diets (KD) are unclear. In this study, we aimed to reveal electrographic and behavioral thresholds in responses to the KD in pentylenetetrazol (PTZ)-induced seizures, as well as its antiepileptogenic effects on PTZ-kindling rats. Additionally, we investigated the potential link between KD and expression levels of two cation chloride co-transporters: K+–Cl− co-transporter 2 (KCC2) and Na+–K+–Cl− co-transporter 1 (NKCC1). The KD group had significantly higher electrographic thresholds than the control (ND) group for the first spike-and-wave, subcontinuous spike-and-wave, high amplitude spike-and-wave, and polyspikes both in the cortex and hippocampus. Compared to the ND group, the KD group had higher behavioral thresholds for behavioral absence, first jerk, first overt myoclonia, and generalized seizures. In the PTZ-kindling model, KD not only prolonged the latency of myoclonic and clonic convulsions, but shortened clonic and generalized duration. In addition, KD rats had higher KCC2 protein expression before kindling, during myoclonic jerks, and GTCS compared with ND rats. There were no significant differences in NKCC1 protein levels between both groups following the four-week dietary intervention without PTZ exposure (before kindling). Moreover, KD inhibited the upregulation of NKCC1 expression induced by kindling in myoclonic jerks and GTCS. Therefore, our findings demonstrated that KD had antiepileptic features in elevating thresholds to most electrographic and behavioral seizure patterns in PTZ-induced rats, as well as delaying the progression and alleviating the severity of seizure in PTZ-kindling model. The antiepileptogenic effects of KD may be attributed to its regulatory properties on KCC2 and NKCC1 protein expression.

Design, synthesis and pharmacological evaluation of N-[4-(4-(alkyl/aryl/heteroaryl)-piperazin-1-yl)-phenyl]-carbamic acid ethyl ester derivatives as novel anticonvulsant agents

A series of alkyl/aryl/heteroaryl piperazine derivatives (37–54) were designed and synthesized as potential anticonvulsant agents. The target compounds are endowed with satisfactory physicochemical as well as pharmacokinetic properties. The synthesized compounds were screened for their in vivo anticonvulsant activity in maximal electroshock (MES) and subcutaneous pentylenetetrazole (sc-PTZ) seizure tests. Further, neurotoxicity evaluation was carried out using rotarod method. Structure activity relationship studies showed that compounds possessing aromatic group at the piperazine ring displayed potent anticonvulsant activity. Majority of the compounds showed anti-MES activity whereas compounds 39, 41, 42, 43, 44, 50, 52, and 53 exhibited anticonvulsant activity in both seizure tests. All the compounds except 42, 46, 47, and 50 did not show neurotoxicity. The most active derivative, 45 demonstrated potent anticonvulsant activity in MES test at the dose of 30mg/kg (0.5h) and 100mg/kg (4h) and also delivered excellent protection in sc-PTZ test (100mg/kg) at both time intervals. Therefore, compound 45 was further assessed in PTZ-kindling model of epilepsy which is widely used model for studying epileptogenesis. This compound was effective in delaying onset of PTZ-evoked seizures at the dose of 5mg/kg in kindled animals and significantly reduced oxidative stress better than standard drug phenobarbital (PB). In result, compound 45 emerged as a most potent and safer anticonvulsant lead molecule.

Proconvulsant effects of estriol, the third estrogen, in the mouse PTZ-kindling model.

The effect of estriol, the third estrogen, was evaluated for its effect on pentylenetetrazole (PTZ)-kindling model of epileptogenesis in mice followed by evaluation on kindling-induced changes in cognitive and motor functions. Kindling was induced by once every 2 days treatment with PTZ (25 mg/kg, i.p.) for 5 weeks. The seizure severity during induction of kindling and percentage incidence of animals kindled at the end of 5 weeks was recorded. Motor function was assessed using a grip strength meter while spatial memory was assessed in a cross maze. Estriol (0.005 and 0.01 mg/kg i.p.) reduced the time for induction of kindling from 5 weeks to 3 and 2 weeks for male and female mice respectively and enhanced the percentage incidence of seizures. Clomiphene (0.9 mg/kg i.p.) delayed the development of kindling and produced anticonvulsant effects. It also partially reversed the proconvulsant effects of estriol. On grip strength test and spontaneous alternation behaviour, a significant decline was observed in kindled mice which was further reduced by pre-treatment with estriol. Both clomiphene and diazepam were unable to reverse the reduced GS of PTZ-kindled mice but enhanced the percentage alternation of such animals. The study shows that estriol has powerful proconvulsant effects. Its administration in hormone replacement therapy or other indications, thus, requires careful monitoring in patients susceptible to epileptic seizures. The anticonvulsant effects of clomiphene requires further investigations.

EVALUATION OF ANTI EPILEPTIC ACTIVITY OF ETHANOLIC EXTRACT OF MURRAYA KAENIGII ON MES, PTZ INDUCED CONVULSIONS IN RATS

The aim of the study was to evaluate anti epileptic activity of ethanolic extract of M. kaenigii on MES, PTZ Induced convulsions in Rats. After the collection of the plant it was extracted with using ethanol solvent by maceration method. Preliminary Phytochemical analysis was done. Acute toxicity was studied. Assessments of antioxidant activity by assay of superoxide dismutase were done and also different enzyme analysis was studied. Effect of EEMK on ME induced convlusions was done in PHT treated animals. Secondary metabolites including alkaloids, saponins, cardiac glycosides and flavonoids were found to be present in the extract and these could have a wide range of compounds with pharmacological activities including those found in this study. Effect of antioxidant enzymes on MES and PTZ induced convolutions were studied. EEMK significantly inhibited the PTZ-induced convulsion, prolonged the latency for convulsion and decreased mortality. The PTZ kindling model is a widely used screening model for testing anticonvulsive compounds. Results based on the PTZ induced clonic seizures and MES induced generalized clonic-tonic seizures in rats model demonstrated that the ethanol extract of Murraya koenigii - Linn. is effective in reducing the severity of clonic seizures and generalized clonic-tonic seizures. This, therefore, supports the traditional use of the plant in the treatment of epilepsy.