Rat Model Of Pilocarpine-induced Epilepsy Research Articles

ATF5-regulated Mitochondrial Unfolded Protein Response Attenuates Neuronal Damage in Epileptic Rat by Reducing Endoplasmic Reticulum Stress Through Mitochondrial ROS.

Endoplasmic reticulum (ER) dysfunction caused by excessive ER stress is a crucial mechanism underlying seizures-induced neuronal injury. Studies have shown that mitochondrial reactive oxygen species (ROS) are closely related to ER stress, and our previous study showed that activating transcription factor 5 (ATF5)-regulated mitochondrial unfolded protein response (mtUPR) modulated mitochondrial ROS generation in a hippocampal neuronal culture model of seizures. However, the effects of ATF5-regulated mtUPR on ER stress and the underlying mechanisms remain uncertain in epilepsy. In this study, ATF5 upregulation by lentivirus infection attenuated seizures-induced neuronal damage and apoptosis in a rat model of pilocarpine-induced epilepsy, whereas ATF5 downregulation by lentivirus infection had the opposite effects. ATF5 upregulation potentiated mtUPR by increasing the expression of mitochondrial chaperone heat shock protein 60 (HSP60) and caseinolytic protease proteolytic subunit (ClpP) and reducing mitochondrial ROS generation in pilocarpine-induced seizures in rats. Additionally, upregulation of ATF5 reduced the expression of glucose-regulated protein 78 (GRP78), protein kinase RNA-like endoplasmic reticulum kinase (PERK), activating transcription factor 4 (ATF4), and C/EBP homologous protein (CHOP), suggesting suppression of ER stress; Moreover, ATF5 upregulation attenuated apoptosis-related proteins such as B-cell lymphoma-2 (BCL2) downregulation, BCL2-associated X (BAX) and cleaved-caspase-3 upregulation. However, ATF5 downregulation exerted the opposite effects. Furthermore, pretreatment with the mitochondria-targeted antioxidant mito-TEMPO attenuated the harmful effects of ATF5 downregulation on ER stress and neuronal apoptosis by reducing mitochondrial ROS generation. Overall, our study suggested that ATF5-regulated mtUPR exerted neuroprotective effects against pilocarpine-induced seizures in rats and the underlying mechanisms might involve mitochondrial ROS-mediated ER stress.

The Traf2 and NcK interacting kinase inhibitor NCB-0846 suppresses seizure activity involving the decrease of GRIA1

Epilepsy, one of the most common neurological disorders, is characterized by spontaneous recurrent seizures. Temporal lobe epilepsy (TLE) is one of the most common medically intractable seizure disorders. Traf2-and NcK-interacting kinase (TNIK) has recently attracted attention as a critical modulation target of many neurological and psychiatric disorders, but its role in epilepsy remains unclear. In this study, we hypothesized the involvement of TNIK in epilepsy and investigated TNIK expression in patients with intractable TLE and in a pilocarpine-induced rat model of epilepsy by western blotting, immunofluorescence, and immunohistochemistry. A pentylenetetrazole (PTZ)-induced epilepsy rat model was used to determine the effect of the TNIK inhibitor NCB-0846 on behavioral manifestations of epilepsy. Coimmunoprecipitation (Co-IP)/mass spectrometry (MS) was used to identify the potential mechanism. Through Co-IP, we detected and confirmed the main potential TNIK interactors. Subcellular fractionation was used to establish the effect of NCB-0846 on the expression of the main interactors in postsynaptic density (PSD) fractions. We found that TNIK was primarily located in neurons and decreased significantly in epilepsy model rats and TLE patients compared with controls. NCB-0846 delayed kindling progression and decreased seizure severity. Co-IP/MS identified 63 candidate TNIK interactors in rat hippocampi, notably CaMKII. Co-IP showed that TNIK might correlate with endogenous GRIA1, SYN2, PSD-95, CaMKIV, GABRG1, and GABRG2. In addition, the significant decrease in GRIA1 in hippocampal total lysate and PSDs after NCB-0846 treatment might help modify the progression of PTZ kindling. Our results suggest that TNIK contributes to epileptic pathology and is a potential antiepileptic drug target.

Dynorphin/KOR inhibits neuronal autophagy by activating mTOR signaling pathway to prevent acute seizure epilepsy.

In previous studies, we found that dynorphin exerts antiepileptic effect by activating the kappa opioid receptor (KOR). However, the role of neuronal autophagy in dynorphin/KOR-mediated antiepileptic is still unclear. This study aimed to investigate the molecular mechanism of dynorphin's antiepileptic effect by inhibiting autophagy and reducing neuronal apoptosis. Here, a pilocarpine-induced rat model of epilepsy was established and hippocampal neurons were treated with Mg2+ -free exposed for epileptiform activity induction. The real-time polymerase chain reaction and Western blot analysis were used to evaluate messengerRNA and protein expression. The TdT-mediated dUTP-biotin nick end labeling staining and flow cytometry were used to analyze cell apoptosis in vivo and in vitro. Neuron cells viability was detected by Cell Counting Kit-8 assay. Immunofluorescent staining and green fluorescent protein-light chain 3 immunofluorescence were used to measure autophagy in vivo and in vitro. Results showed that overexpression ofprodynorphinalleviated neuronal apoptosis, activated the mammalian target of rapamycin (mTOR) signaling pathway, and inhibited neuronal autophagy in epileptic rats. Dynorphin inhibited Mg2+ -free-induced seizure-like neuron apoptosis, partially reversing the effect of Mg2+ -free on the mTOR signaling pathway and seizure-like neuron autophagy. Further, using rapamycin, we found that dynorphin inhibited Mg2+ -free-induced seizure-like neuron autophagy and apoptosis by activating the mTOR signaling pathway. In conclusion, dynorphin inhibits autophagy by activating the mTOR signaling pathway and has a protective effect on epilepsy acute seizure and epilepsy-induced brain injury.

Evaluation of anxiolytic and antidepressant effect of Saraswata Churna in the pilocarpine induced rat model of epilepsy

The aim of our present study was to assess Saraswata Churna's anxiolytic and anti-depressant efficacy in a Pilocarpine-induced epilepsy rat model. Depression and anxiety are major psychological symptoms in epilepsy patients, and they have a significant impact on health-related quality of life. Many cases of neurological problems have been treated using Saraswata Churna, an ayurvedic medicine. The elevated plus maze (EVPM) was used to assess anxiolytic action, while the forced swimming test (FST) was used to assess antidepressant activity. The EVPM test has been shown to be effective in determining the anxiolytic effects of pharmaceuticals and various steroid as well as non-steroid hormones, as well as identifying the brain locations and mechanisms that underpin anxiety-related behaviour in rodents. Saraswata churna was reported to be effective in reducing anxiety and depression in rats at a dosage of 500 mg/kg.b.w by increasing open arm time and decreasing closed arm entrance in the EVPM paradigm and increasing the duration of immobility in the FST model. As a result, Saraswata Churna could be used as an effective therapeutic agent for anxiety and depression.

The Role of the Negative Regulation of Microglia-Mediated Neuroinflammation in Improving Emotional Behavior After Epileptic Seizures.

ObjectiveStudies have long shown that uncontrolled inflammatory responses in the brain play a key role in epilepsy pathogenesis. Microglias play an important role in epileptic-induced neuroinflammation, but their role after epileptic seizures is still poorly understood. Alleviating epilepsy and its comorbidities has become a key area of interest for pediatricians.MethodsA pilocarpine-induced rat model of epilepsy was established. The rats were randomly divided into four groups: a control group, epilepsy group, TLR4 inhibitor group (epilepsy+TAK-242), and NF-κB antagonist group (epilepsy+BAY11–7082).Results1. The results of TUNEL staining showed that the expression in rats in the epilepsy group was the most obvious and was significantly different from that in rats in the control, EP+BAY and EP+TAK groups. 2. The expression of TLR4 and NF-κB was highest in rats in the epilepsy group and was significantly different from that in rats in the control, EP+BAY and EP+TAK groups. 3. The fluorescence intensity and number of IBA-1-positive cells in rats in the epilepsy group were highest and significantly different from those in rats in the control, EP+BAY and EP+TAK groups. Western blot analysis of IBA-1 showed that the expression in rats in the epilepsy group was the highest and was statistically significant. 4. CD68 was the highest in rats in the epilepsy group and was statistically significant. 5. In the open-field experiment, the central region residence time of rats in the EP group was delayed, the central region movement distance traveled was prolonged, the total distance traveled was prolonged, and the average speed was increased. Compared with rats in the EP group, rats in the EP+BAY and EP+ TAK groups exhibited improvements to different degrees.ConclusionAt the tissue level, downregulation of the TLR4/NF-κB inflammatory pathway in epilepsy could inhibit microglial activation and the expression of the inflammatory factor CD68, could inhibit hyperphagocytosis, and inhibit the occurrence and exacerbation of epilepsy, thus improving cognitive and emotional disorders after epileptic seizures.

MiR-101a-3p Attenuated Pilocarpine-Induced Epilepsy by Downregulating c-FOS.

This study aimed to explore the effects and function of microRNA-101a-3p (miR-101a-3p) in epilepsy. Rat model of pilocarpine-induced epilepsy was established and the seizure frequency was recorded. Expression of miR-101a-3p and c-Fos in hippocampus tissues of Rat models were detected by qRT-PCR and western blot. Besides, we established a hippocampal neuronal culture model of acquired epilepsy using Mg2+ free medium to evaluate the effects of miR-101a-3p and c-Fos in vitro. Cells were transfected with miR-101a-3p mimic, si-c-FOS, miR-101a-3p + c-FOS and its corresponding controls. MTT assay was used to detect cell viability upon transfection. Flow cytometry was performed to determine the apoptosis rate. Western blot was performed to measure the protein expression of apoptosis-related proteins (Bcl-2, Bax, and cleaved caspase 3), autophagy-related proteins (LC3 and Beclin1) and c-FOS. The targeting relationship between miR-101a-3p and c-FOS was predicted and verified by TargetScan software and dual-luciferase reporter assay. The role of miR-101a-3p was validated using epilepsy rat models in vivo. Another Rat models of pilocarpine-induced epilepsy with miR-NC or miR-101a-3p injection were established to evaluate the effect of miR-101a-3p overexpression on epilepsy in vivo. MiR-101a-3p was downregulated while c-FOS was increased in hippocampus tissues of Rat model of pilocarpine-induced epilepsy. Overexpression of miR-101a-3p or c-FOS depletion promoted cell viability, inhibited cell apoptosis and autophagy. C-FOS was a target of miR-101a-3p and miR-101a-3p negatively regulated c-FOS expression to function in epilepsy. Overexpression of miR-101a-3p attenuated pilocarpine-induced epilepsy in Rats in vivo. This study indicated that miR-101a-3p could attenuate pilocarpine-induced epilepsy by repressing c-Fos expression.

Neurochemical effects of sleep deprivation in the hippocampus of the pilocarpine-induced rat model of epilepsy

Objective(s): The present study aims to investigate the pathological mechanisms mediating the effect of paradoxical sleep deprivation (PSD) for 48 hr on the spontaneous recurrent seizures (SRS) stage of the pilocarpine rat model of temporal lobe epilepsy. Materials and Methods: This was carried out through assessment of amino acid neurotransmitter levels, the main oxidative stress parameters, and the levels of tumor necrosis factor-I± (TNF-I±), interleukin-1I² (IL-1I²), and interleukin-6 (IL-6) in the hippocampus. The experimental animals were divided into 4 groups: control, epileptic, PSD, and epileptic㴠 groups.Results: Data indicated that PSD in epileptic rats induced a significant decrease in GSH levels. TNF-I± increased significantly in the PSD group and decreased significantly in both epileptic rats and epileptic rats deprived of paradoxical sleep. PSD induced a significant increase in glutamine, glutamate, and aspartate and a significant decrease in GABA. In epileptic rats and epileptic rats deprived of PS, a significant increase in aspartate and a significant decrease in GABA and taurine were recorded. Â Conclusion: The present data suggest that exposure to PSD for 48 hr did not worsen the alterations produced in the present epileptic model. However, epileptic, PSD, epileptic + PSD groups showed a state of hyperexcitability and oxidative stress. PSD may increase the susceptibility of animals to the development of epilepsy.Â

Neurochemical effects of sleep deprivation in the hippocampus of the pilocarpine-induced rat model of epilepsy

Objective(s):The present study aims to investigate the pathological mechanisms mediating the effect of paradoxical sleep deprivation (PSD) for 48 hr on the spontaneous recurrent seizures (SRS) stage of the pilocarpine rat model of temporal lobe epilepsy. Materials and Methods:This was carried out through assessment of amino acid neurotransmitter levels, the main oxidative stress parameters, and the levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) in the hippocampus. The experimental animals were divided into 4 groups: control, epileptic, PSD, and epileptic+PSD groups.Results:Data indicated that PSD in epileptic rats induced a significant decrease in GSH levels. TNF-α increased significantly in the PSD group and decreased significantly in both epileptic rats and epileptic rats deprived of paradoxical sleep. PSD induced a significant increase in glutamine, glutamate, and aspartate and a significant decrease in GABA. In epileptic rats and epileptic rats deprived of PS, a significant increase in aspartate and a significant decrease in GABA and taurine were recorded. Conclusion:The present data suggest that exposure to PSD for 48 hr did not worsen the alterations produced in the present epileptic model. However, epileptic, PSD, epileptic + PSD groups showed a state of hyperexcitability and oxidative stress. PSD may increase the susceptibility of animals to the development of epilepsy.

Dynorphin activation of kappa opioid receptor promotes microglial polarization toward M2 phenotype via TLR4/NF-\u03baB pathway

BackgroundMicroglia-mediated neuroinflammation is associated with epilepsy. Switching microglial polarization from the pro-inflammatory M1 phenotype to the anti-inflammatory M2 phenotype represents a novel therapeutic strategy for mitigating epileptogenesis. We previously found that dynorphins protected against epilepsy via activation of kappa opioid receptor (KOR). Here, this study aims to investigate the role and the mechanism of dynorphin in regulating microglial polarization.MethodsA pilocarpine-induced rat model of epilepsy was established and lipopolysaccharide (LPS)-activated BV-2 microglial cells were used as an inflammatory model to explore the mechanism of dynorphin regulating microglial polarization.ResultsOverexpression of the dynorphin precursor protein prodynorphin (PDYN) alleviated the pilocarpine-induced neuronal apoptosis, promoted microglial polarization to the M2 phenotype, and inhibited pilocarpine-induced Toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB) pathway in the hippocampi of epileptic rats. Dynorphin activation of KOR promoted microglial M2 polarization via inhibiting TLR4/NF-κB pathway in LPS-stimulated BV-2 microglial cells. Moreover, dynorphin/KOR regulated microglial M2 polarization inhibited apoptosis of the primary mouse hippocampal neurons.ConclusionIn conclusion, dynorphin activation of KOR promotes microglia polarization toward M2 phenotype via inhibiting TLR4/NF-κB pathway.

Activation of microglia in rat model of epilepsy during persistent state and analysis of temporal and spatial expression of different types of microglia

Objective To investigate the activation of microglia in state of continuity (SE) of epileptic rat model and the temporal and spatial expression of different types of microglia. Methods 60 healthy male Wistar rats were randomly divided into control group, SE at the beginning (NC group), SE lasting for 30 minutes (SE30min group), SE lasting for 1 hour (SE1h group), SE holding. For 2 hours (SE2h), 12 rats in each group. Behavioral changes and activation of microglia in hippocampus were observed. The expression levels of IL-6, IL-1beta and IL-10 and IL-4 were detected. Results In the SE group, with the prolongation of time, the response of MG cells increased, and the activation of MG cells was most obvious in SE2h group. The expression levels of IL-10, IL-4 protein and mRNA in SE1h group and SE2h group were significantly higher than those in NC group and SE30min group, while the expression levels of IL-6, IL-1beta protein and mRNA in M1 group were significantly lower than those in NC group and SE30min group (P<0.05). Conclusion Activation of microglia can be seen in pilocarpine-induced epilepsy rat model. With the prolongation of seizure time, polarization of microglia gradually transforms into neurotrophic M2 cells. The expression levels of anti-inflammatory factors IL-10 and IL-4 will increase, while the expression of pro-inflammatory factors IL-6 and IL-1beta will decrease. Key words: Status epilepticus; Microglia; Cell polarization; Classical activation; Alternative activation

Hippocampal low-frequency stimulation improves cognitive function in pharmacoresistant epileptic rats

ObjectiveThe aims of the present study were to observe the changes of cognitive function in a pilocarpine-induced rat model of epilepsy, and to investigate the effects of hippocampal low-frequency stimulation (Hip-LFS) on cognitive function in rats with pharmacoresistant epilepsy. MethodsA total of 100 male Sprague Dawley rats were randomly selected to establish an epilepsy model. Rats with successfully induced epilepsy were injected intraperitoneally with phenobarbital and phenytoin for pharmacoresistance selection. The selected pharmacoresistant epileptic (PRE) rats were assigned to a pharmacoresistant control group (PRC group), or a group that received Hip-LFS (LFS group). The same number of rats with pharmacosensitive epilepsy formed the PSC group, and a normal control (NCR) group was included. A novel object recognition (NOR) test, and a Morris water maze (MWM) task were used to assess cognitive function in all groups. ResultsThe epileptic rats showed decreased abilities of learning and memory compared with normal control. The rats in the LFS group displayed significantly shorter escape latency in place navigation, spent longer times in the target quadrant, and traveled longer distances on the platform in the spatial probe test than PRC group. In the NOR test, compared with the PRC group, the discrimination index of the LFS group was significantly increased. Compared with the PRC group, the average frequency and duration of seizures were also decreased in the LFS group. ConclusionsThe present pilocarpine-induced rat model of epilepsy showed impaired cognitive function, especially in rats with PRE. The Hip-LFS treatment could effectively improve the cognitive function of rats with PRE.

Medial septal stimulation increases seizure threshold and improves cognition in epileptic rats

BackgroundTemporal lobe epilepsy is most prevalent among focal epilepsies, and nearly one-third of patients are refractory to pharmacological intervention. Persistent cognitive and neurobehavioral comorbidities also occur due to the recurrent nature of seizures and medication-related side effects. HypothesisElectrical neuromodulation is an effective strategy to reduce seizures both in animal models and clinically, but its efficacy to modulate cognition remains unclear. We hypothesized that theta frequency stimulation of the medial septum would increase septohippocampal oscillations, increase seizure threshold, and improve spatial learning in a rat model of pilocarpine-induced epilepsy. MethodsSham and pilocarpine rats were implanted with electrodes in the medial septum, hippocampus and prefrontal cortex. EEG was assessed days prior to and following stimulation. Sham and pilocarpine-treated rats received either no stimulation, continuous (throughout each behavior), or pre-task (one minute prior to each behavior) 7.7 Hz septal stimulation during the Barnes maze spatial navigation test and also during assessment of flurothyl-induced seizures. ResultsBoth continuous and pre-task stimulation prevented epilepsy-associated reductions in theta oscillations over time. Additionally, both stimulation paradigms significantly improved spatial navigation in the Barnes maze, reducing latency and improving search strategy. Moreover, stimulation led to significant increases in seizure threshold in pilocarpine-treated rats. There was no evidence of cognitive enhancement or increased seizure threshold in stimulated sham rats. ConclusionThese findings have profound implications as theta stimulation of the septum represents a single frequency and target that has the potential to both improve cognition and reduce seizures for patients with refractory epilepsy.

Dynorphin activation of kappa opioid receptor protects against epilepsy and seizure-induced brain injury via PI3K/Akt/Nrf2/HO-1 pathway

ABSTRACT Dynorphins act as endogenous anticonvulsants via activation of kappa opioid receptor (KOR). However, the mechanism underlying the anticonvulsant role remains elusive. This study aims to investigate whether the potential protection of KOR activation by dynorphin against epilepsy was associated with the regulation of PI3K/Akt/Nrf2/HO-1 pathway. Here, a pilocarpine-induced rat model of epilepsy and Mg2+-free-induced epileptiform hippocampal neurons were established. Decreased prodynorphin (PDYN) expression, suppressed PI3K/Akt pathway, and activated Nrf2/HO-1 pathway were observed in rat epileptiform hippocampal tissues and in vitro neurons. Furthermore, dynorphin activation of KOR alleviated in vitro seizure-like neuron injury via activation of PI3K/Akt/Nrf2/HO-1 pathway. Further in vivo investigation revealed that PDYN overexpression by intra-hippocampus injection of PDYN-overexpressing lentiviruses decreased hippocampal neuronal apoptosis and serum levels of inflammatory cytokines and malondialdehyde (MDA) content, and increased serum superoxide dismutase (SOD) level, in pilocarpine-induced epileptic rats. The protection of PDYN in vivo was associated with the activation of PI3K/Akt/Nrf2/HO-1 pathway. In conclusion, dynorphin activation of KOR protects against epilepsy and seizure-induced brain injury, which is associated with activation of the PI3K/Akt/Nrf2/HO-1 pathway.

Depression comorbidity in epileptic rats is related to brain glucose hypometabolism and hypersynchronicity in the metabolic network architecture.

Temporal lobe epilepsy (TLE) is one of the most common types of epilepsy syndromes in the world. Depression is an important comorbidity of epilepsy, which has been reported in patients with TLE and in different experimental models of epilepsy. However, there is no established consensus on which brain regions are associated with the manifestation of depression in epilepsy. Here, we investigated the alterations in cerebral glucose metabolism and the metabolic network in the pilocarpine-induced rat model of epilepsy and correlated it with depressive behavior during the chronic phase of epilepsy. Fluorodeoxyglucose (18 F-FDG) was used to investigate the cerebral metabolism, and a cross-correlation matrix was used to examine the metabolic network in chronically epileptic rats using micro-positron emission tomography (microPET) imaging. An experimental model of epilepsy was induced by pilocarpine injection (320 mg/kg, ip). Forced swim test (FST), sucrose preference test (SPT), and eating-related depression test (ERDT) were used to evaluate depression-like behavior. Our results show an association between epilepsy and depression comorbidity based on changes in both cerebral glucose metabolism and the functional metabolic network. In addition, we have identified a significant correlation between brain glucose hypometabolism and depressive-like behavior in chronically epileptic rats. Furthermore, we found that the epileptic depressed group presents a hypersynchronous brain metabolic network in relation to the epileptic nondepressed group. This study revealed relevant alterations in glucose metabolism and the metabolic network among the brain regions of interest for both epilepsy and depression pathologies. Thus it seems that depression in epileptic animals is associated with a more diffuse hypometabolism and altered metabolic network architecture and plays an important role in chronic epilepsy.

2R,4R-APDC, a Metabotropic Glutamate Receptor Agonist, Reduced Neuronal Apoptosis by Upregulating MicroRNA-128 in a Rat Model After Seizures.

This study aimed to study the protective effect of (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylate (2R,4R-APDC), a selective metabotropic glutamate receptor agonist, against hippocampal neuronal apoptosis induced by seizures in a rat model of pilocarpine-induced epilepsy. The Morris water maze test was used to assess the spatial memory abilities of epileptic rats with or without 2R,4R-APDC treatment. TUNEL assay was performed to examine neuronal apoptosis in hippocampus. Western blot was conducted to evaluate changes in the levels of caspase-3 and caspase-9 in hippocampus. Real-time PCR was used to determine the levels of microRNA-128 (miR-128) in hippocampus. The results of the Morris water maze test showed that the 2R,4R-APDC treatment reduced the escape latencies and swimming lengths of rats after seizures. The TUNEL assay showed that 2R,4R-APDC significantly counteracted seizure-induced cell apoptosis. The western blot confirmed this finding, demonstrating that the levels of cleaved caspase-3 and cleaved caspase-9 were potently decreased by 2R,4R-APDC in rat hippocampus after seizures. In addition, 2R,4R-APDC upregulated miR-128 expression levels in the hippocampus. A miR-128 mimic or inhibitor decreased or increased the percentage of TUNEL-positive cells in rats after seizures and 2R,4R-APDC treatment, respectively. The levels of both cleaved caspase-3 and cleaved caspase-9 were decreased in hippocampus exposed to the miR-128 mimic, whereas they were markedly increased in miR-128 inhibitor-treated hippocampus. In conclusion, 2R,4R-APDC protected hippocampal cells from cell apoptosis after seizures, possibly by upregulating miR-128.

Early Seizure Detection by Applying Frequency-Based Algorithm Derived from the Principal Component Analysis

The use of automatic electrical stimulation in response to early seizure detection has been introduced as a new treatment for intractable epilepsy. For the effective application of this method as a successful treatment, improving the accuracy of the early seizure detection is crucial. In this paper, we proposed the application of a frequency-based algorithm derived from principal component analysis (PCA), and demonstrated improved efficacy for early seizure detection in a pilocarpine-induced epilepsy rat model. A total of 100 ictal electroencephalographs (EEG) during spontaneous recurrent seizures from 11 epileptic rats were finally included for the analysis. PCA was applied to the covariance matrix of a conventional EEG frequency band signal. Two PCA results were compared: one from the initial segment of seizures (5 sec of seizure onset) and the other from the whole segment of seizures. In order to compare the accuracy, we obtained the specific threshold satisfying the target performance from the training set, and compared the False Positive (FP), False Negative (FN), and Latency (Lat) of the PCA based feature derived from the initial segment of seizures to the other six features in the testing set. The PCA based feature derived from the initial segment of seizures performed significantly better than other features with a 1.40% FP, zero FN, and 0.14 s Lat. These results demonstrated that the proposed frequency-based feature from PCA that captures the characteristics of the initial phase of seizure was effective for early detection of seizures. Experiments with rat ictal EEGs showed an improved early seizure detection rate with PCA applied to the covariance of the initial 5 s segment of visual seizure onset instead of using the whole seizure segment or other conventional frequency bands.

Tangeretin alters neuronal apoptosis and ameliorates the severity of seizures in experimental epilepsy-induced rats by modulating apoptotic protein expressions, regulating matrix metalloproteinases, and activating the PI3K/Akt cell survival pathway

PurposeEpilepsy is complex neural disarray categorized by recurring seizures. Despite recent advances in pharmacotherapies for epilepsy, its treatment remains a challenge due to the contrary effects of the drugs. As a result, the identification of novel anti-epileptic drugs (AEDs) with neuroprotective properties and few side effects is of great value. Thus, the present study assessed the treatment effects of tangeretin using a rat model of pilocarpine-induced epilepsy. Materials and MethodsSeparate groups of male Wistar rats received oral administrations of tangeretin at 50, 100, or 200mg/kg for 10 days and then, on the 10th day, they received an intraperitoneal injection of pilocarpine (30mg/kg). Subsequently, neuronal degeneration and apoptosis were assessed using Nissl staining and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay procedures. Additionally, the expressions of phosphatidylinositol-3-kinase (PI3K/Akt) pathway proteins, cleaved caspase-3, Bad, Bcl-2, Bcl-xL, and Bax were determined using Western blot analyses. ResultsTangeretin reduced the seizure scores and latency to first seizure of the rats and effectively activated the pilocarpine-induced suppression of PI3K/Akt signaling. Additionally, tangeretin effectively regulated the levels of apoptosis-inducing factor (AIF) in mitochondria as well as the expressions of apoptotic pathway proteins. Seizure-induced elevations in the activities and expressions of matrix metalloproteinases (MMPs)-2 and -9 were also modulated. ConclusionThe present results indicate that tangeretin exerted potent neuroprotective effects against pilocarpine-induced seizures via the activation of PI3K/Akt signaling and the regulation of MMPs.

Up-regulated BAFF and BAFF receptor expression in patients with intractable temporal lobe epilepsy and a pilocarpine-induced epilepsy rat model

PurposeSome studies have suggested that BAFF and BAFFR are highly expressed in the central nervous system (CNS) and participate in inflammatory and immune associated diseases. However, whether BAFF and BAFFR are involved in the pathogenesis of epilepsy remains unknown. This study aimed to investigate the expression of BAFF and BAFFR proteins in the brains of patients with temporal lobe epilepsy (TLE) and in a pilocarpine-induced rat model of TLE to identify possible roles of the BAFF–BAFFR signaling pathway in epileptogenesis. MethodsReal-time quantitative polymerase chain reaction (RT-qPCR), western blot, immunohistochemistry, and double-immunofluorescence were performed in this study. ResultsThe results showed that BAFF and BAFFR expression levels were markedly up-regulated in intractable TLE patients and TLE rats. Moreover, BAFF and BAFFR proteins mainly highly expressed in the membranes and cytoplasms of the dendritic marker MAP2 in the cortex and hippocampus. ConclusionTherefore, the significant increased in BAFF and BAFFR protein expression in both TLE patients and rats suggest that BAFF and BAFFR may play important roles in regulating the pathogenesis of epilepsy.

Anticonvulsant effect of piperine ameliorates memory impairment, inflammation and oxidative stress in a rat model of pilocarpine-induced epilepsy.

The primary active component of black pepper is piperine, which is purified and used to treat epilepsy, achieving higher efficiency when purified. The present study was conducted to evaluate whether the anticonvulsant effect of piperine ameliorates pilocarpine-induced epilepsy, and to investigate the mechanism underlying these effects. Epilepsy was induced in Sprague Dawley rats using pilocarpine. Pilocarpine-induced epilepsy in the rats was treated with 40 mg/kg piperine for 45 consecutive days. Status epilepticus and a Morris water maze test were used to analyze the anticonvulsant effects of piperine in the epileptic rats. Inflammation and oxidative stress were then measured using commercially-available kits following piperine treatment. Lastly, the activity of caspase-3 and the protein expression levels of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were evaluated using commercially-available kits and western blot analysis, respectively. The results demonstrated that treatment with piperine was able to reduce the status epilepticus and prevented memory impairment following pilocarpine-induced epilepsy in rats. The anticonvulsant effects of piperine decreased inflammation and oxidative stress following pilocarpine-induced epilepsy in rats. The upregulated activity of caspase-3 and expression levels of Bax/Bcl-2 were suppressed following treatment with piperine in the rats with pilocarpine-induced epilepsy. These results suggest that the anticonvulsant effects of piperine ameliorate memory impairment, inflammation and oxidative stress in a rat model of pilocarpine-induced epilepsy.

Expression of Glypican-4 in the brains of epileptic patients and epileptic animals and its effects on epileptic seizures

Glypican-4 (Gpc4) has been found to play an important role in enhancing miniature excitatory postsynaptic currents (mEPSCs). But, the relationship between Gpc4 and epilepsy is still a mystery. In this study, we investigated the expression patterns of Gpc4 in patients with epilepsy and in a pilocarpine-induced rat model of epilepsy. We also determined if altered Gpc4 expression resulted in increased susceptibility to seizures. Western blotting and immunofluorescent methods were utilized. Gpc4 was significantly increased in patients and epileptic rats induced by pilocarpine injection. According to behavioral studies, downregulation of Gpc4 by Gpc4 siRNA decreased spontaneous seizure frequency, while upregulation of Gpc4 by recombinant Gpc4 overexpression led to a converse result. These findings support the hypothesis that increased expression of Gpc4 in the brain is associated with epileptic seizures.