Rat Model Of Status Epilepticus Research Articles

CoQ10 targeted hippocampal ferroptosis in a status epilepticus rat model

Status epilepticus (SE), the most severe form of epilepsy, leads to brain damage. Uncertainty persists about the mechanisms that lead to the pathophysiology of epilepsy and the death of neurons. Overloading of intracellular iron ions has recently been identified as the cause of a newly recognized form of controlled cell death called ferroptosis. Inhibiting ferroptosis has shown promise as a treatment for epilepsy, according to recent studies. So, the current study aimed to assess the possible antiepileptic impact of CoQ10 either alone or with the standard antiepileptic drug sodium valproate (SVP) and to evaluate the targeted effect of COQ10 on hippocampal oxidative stress and ferroptosis in a SE rat model. Using a lithium-pilocarpine rat model of epilepsy, we evaluated the effect of SVP, CoQ10, or both on seizure severity, histological, and immunohistochemical of the hippocampus. Furthermore, due to the essential role of oxidative stress and lipid peroxidation in inducing ferroptosis, we evaluated malonaldehyde (MDA), reduced glutathione (GSH), glutathione peroxidase 4 (GPX4), and ferritin in tissue homogenate. Our work illustrated that ferroptosis occurs in murine models of lithium-pilocarpine-induced seizures (epileptic group). Nissl staining revealed significant neurodegeneration. A significant increase in the number of astrocytes stained with an astrocyte-specific marker was observed in the hippocampus. Effective seizure relief can be achieved in the seizure model by administering CoQ10 alone compared to SVP. This was accomplished by lowering ferritin levels and increasing GPX4, reducing MDA, and increasing GSH in the hippocampus tissue homogenate. In addition, the benefits of SVP therapy for regulating iron stores, GPX4, and oxidative stress markers were amplified by incorporating CoQ10 as compared to SVP alone. It was concluded that CoQ10 alone has a more beneficial effect than SVP alone in restoring histological structures and has a targeted effect on hippocampal oxidative stress and ferroptosis. In addition, COQ10 could be useful as an adjuvant to SVP in protecting against oxidative damage and ferroptosis-related damage that result from epileptic seizures.

SREBP1 deficiency diminishes glutamate-mediated HT22 cell damage and hippocampal neuronal pyroptosis induced by status epilepticus

Status epilepticus (SE) is a life-threatening disorder that can result in death or severe brain damage, and there is a substantial body of evidence suggesting a strong association between pyroptosis and SE. Sterol regulatory element binding protein 1 (SREBP1) is a significant transcription factor participating in both lipid homeostasis and glucose metabolism. However, the function of SREBP1 in pyroptosis during SE remains unknown. In this study, we established a SE rat model by intraperitoneal injection of lithium chloride and pilocarpine in vivo. Additionally, we treated HT22 hippocampal cells with glutamate to create neuronal injury models in vitro. Our results demonstrated a significant induction of SREBP1, inflammasomes, and pyroptosis in the hippocampus of SE rats and glutamate-treated HT22 cells. Moreover, we found that SREBP1 is regulated by the mTOR signaling pathway, and inhibiting mTOR signaling contributed to the amelioration of SE-induced hippocampal neuron pyroptosis, accompanied by a reduction in SREBP1 expression. Furthermore, we conducted siRNA-mediated knockdown of SREBP1 in HT22 cells and observed a significant reversal of glutamate-induced cell death, activation of inflammasomes, and pyroptosis. Importantly, our confocal immunofluorescence analysis revealed the co-localization of SREBP1 and NLRP1. In conclusion, our findings suggest that deficiency of SREBP1 attenuates glutamate-induced HT22 cell injury and hippocampal neuronal pyroptosis in rats following SE. Targeting SREBP1 may hold promise as a therapeutic strategy for SE.

A Pediatric Rat Model of Organophosphate-Induced Refractory Status Epilepticus: Characterization of Long-Term Epileptic Seizure Activity, Neurologic Dysfunction and Neurodegeneration.

Children are highly vulnerable to the neurotoxic effects of organophosphates (OPs), which can cause neuronal developmental defects, including intellectual disability, autism, epilepsy, and related comorbidities. Unfortunately, no specific pediatric OP neurotoxicity model currently exists. In this study, we developed and characterized a pediatric rat model of status epilepticus (SE) induced by the OP diisopropylfluorophosphate (DFP) and examined its impact on long-term neurological outcomes. Postnatal day 21 rats were exposed to a DFP regimen with standard antidotes. Progressive behavioral deteriorations were assessed over a three-month period. Development of epileptic seizures, ictal discharges, high-frequency oscillations (HFOs), and interictal spikes were monitored by video-electroencephalography recordings. Histology-stereology analysis was performed to assess neurodegeneration, neuroinflammation, and morphologic abnormalities. DFP-exposed, post-SE animals exhibited significantly elevated levels of anxiety and depression than age-matched controls at 1, 2, and 3 months post-exposure. DFP-exposed animals displayed aggressive behavior and a marked decline in object recognition memory, as well as prominent impairment in spatial learning and memory. DFP-exposed animals had striking electrographic abnormalities with the occurrence of displayed epileptic seizures, ictal discharges, HFOs, and interictal spikes, suggesting chronic epilepsy. Neuropathological analysis showed substantially fewer principal neurons and inhibitory interneurons with a marked increase in reactive microglia and neuroinflammation in the hippocampus and other brain regions. DFP-exposed animals also exhibited mossy fiber sprouting indicating impaired network formations. Long-term epileptic seizures and neuropsychiatric functional deficits induced by DFP were consistent with neuropathological defects. Collectively, this pediatric model displays many hallmarks of chronic sequelae reminiscent of children exposed to OPs, suggesting that it will be a valuable tool for investigating pathologic mechanisms and potential treatment strategies to attenuate long-term OP neurotoxicity. SIGNIFICANCE STATEMENT: Millions of children are exposed to organophosphates (OPs) used in agriculture or chemical incidents. This study investigated the long-term impact of neonatal exposure to the OP chemical diisopropylfluorophosphate (DFP) on neurobehavioral and neurodevelopmental outcomes in adulthood. DFP exposure caused long-lasting behavioral abnormalities, epileptic seizures, and bilateral brain defects with an array of neurological sequelae seen in children's OP neurotoxicity. Thus, this model provides a novel tool to explore therapeutic interventions that mitigate long-term neurotoxic effects of children exposed to OP-induced seizures and status epilepticus.

Cannabidiol Anticonvulsant Effects Against Lithium-Pilocarpine-Induced Status Epilepticus in Male Rats Are Mediated by Neuroinflammation Modulation and Cannabinoids 1 (CB1), But Not CB2 and GABAA Receptors.

Background: Status epilepticus (SE) is a series of seizures that can lead to serious neurological damages. Cannabidiol (CBD) is extracted from the cannabis plant, which has been approved as an antiseizure medication. This study aimed to determine the efficacy of various doses of CBD on lithium-pilocarpine-induced SE in rats and possible involvement of multiple pharmacological pathways. We hypothesized that cannabinoid receptors type 1 (CB1) and CB2, as well as GABAA receptors, might have important roles in the anticonvulsant effects of CBD against SE by its anti-inflammatory effects. Methods: SE was induced by intraperitoneal (i.p.) injection of lithium (127 mg/kg, i.p.) and pilocarpine (60 mg/kg, i.p., 20 h after lithium). Forty-two male rats were divided into seven groups (including control and sham groups), and the treated groups received different doses of CBD (1, 3, 5, 10, and 25 mg/kg, i.p.). SE score was recorded over the next 2 h following pilocarpine injection. Then, we measured the levels of pro-inflammatory cytokines, including interleukin (IL)-lβ and tumor necrosis factor (TNF)-α, using ELISA kits. Also we analyzed the expression of CB1, CB2, and GABAA receptors using the Western blot technique. Results: CBD at 5 mg/kg significantly reduced Racine's scale and duration of seizures, and increased the onset time of seizure. Moreover, CBD 5 mg/kg caused significant reductions in the elevated levels of IL-lβ and TNF-α, as well as a significant increase in the decreased level of CB1 receptor expression compared to the control group. In other word, CBD reverted the effects of SE in terms of neuroinflammation and CB1 receptor. Based on the obtained results, CBD was not able to restore the declined levels of CB2 or GABAA receptors. Conclusion: Our study found anticonvulsant effects of CBD on the SE rat model induced by lithium-pilocarpine with probable involvement of CB1 receptors and anti-inflammatory effects by reducing IL-1β and TNF-α markers independent of CB2 and GABAA receptors.

Differential Regulation of Wnt/β-catenin Signaling in Acute and Chronic Epilepsy in Repeated Low Dose Lithium-Pilocarpine Rat Model of Status Epilepticus

Epilepsy is a chronic neurological complication characterized by unprovoked seizure episodes due to the imbalance between excitatory and inhibitory neurons. The epileptogenesis process has been reported to be involved in chronic epilepsy however, the mechanism underlying epileptogenesis remains unclear. Recent studies have shown the possible involvement of Wnt/β-catenin signaling in the neurogenesis and neuronal reorganization in epileptogenesis. In this study, we used repeated low dose lithium-pilocarpine model of status epilepsy (SE) to study the involvement of Wnt/β-catenin signaling at acute and chronic stages post SE induction. The acute study ranged from day 0 to day 28 post SE induction and the chronic study ranged from day 0 to day 56 post SE induction. Several neurobehavioral parameters and seizure score and seizure frequency was analysed until the end of the study. The proteins involved in the regulation of Wnt/β-catenin signaling and downstream cascading were analysed using western blot and quantitative real-time PCR analysis. The Wnt/β-catenin pathway was found inactive in acute SE, while the same was found activated at the chronic stage. Our findings suggest that the activated Wnt/β-catenin signaling in chronic epilepsy might be the possible mechanism underlying epileptogenesis as indicated by increased neuronal count, increased synaptic density, astrogliosis and apoptosis in chronic epilepsy. These findings can help target the Wnt/β-catenin pathway differentially depending upon the type of epilepsy. The acute stage characterized by SE can be improved by targeting GSK-3β levels and the chronic stage characterized by temporal lobe epilepsy can be improved by targeting β-catenin and disheveled proteins.

Effect of intranasal and oral administration of levetiracetam onthe temporal and spatial distributions of SV2A in the KA-induced rat model of SE.

To investigate the effectiveness of nasal delivery of levetiracetam (LEV) on the distributions of synaptic vesicle protein 2 isoform A (SV2A) in epileptic rats with injection of kainic acid (KA) into amygdala. A total of 138 rats were randomly divided into four groups, including the Sham surgery group, the epilepsy group (EP), and the LEV oral administration (LPO) and nasal delivery (LND) groups. The rat intra-amygdala KA model of epilepsy was constructed. Pathological changes of rat brain tissue after status epilepticus (SE) were detected using haematoxylin and eosin staining. Expression of SV2A in rat hippocampus after SE was evaluated using the western blotting analysis. Expression and distribution of SV2A in rat hippocampus after SE were detected based on immunofluorescence staining. The EP group showed evident cell loss and tissue necrosis in the CA3 area of hippocampus, whereas the tissue damage in both LPO and LND groups was significantly reduced. Western blotting analysis showed that the expressions of SV2A in the hippocampus of both EP and LND groups were significantly decreased 1 week after SE, increased to the similar levels of the Sham group in 2 weeks, and continuously increased 4 weeks after SE to the level significantly higher than that of the Sham group. Results of immunofluorescence revealed largely the same expression patterns of SV2A in the CA3 area of hippocampus as those in the entire hippocampus. Our study revealed the same antiepileptic and neuronal protective effects by the nasal and oral administrations of LEV, without changing the expression level of SV2A.

Time- and region-dependent blood-brain barrier impairment in a rat model of organophosphate-induced status epilepticus

Acute organophosphate (OP) intoxication can trigger seizures that progress to status epilepticus (SE), and survivors often develop chronic morbidities, including spontaneous recurrent seizures (SRS). The pathogenic mechanisms underlying OP-induced SRS are unknown, but increased BBB permeability is hypothesized to be involved. Previous studies reported BBB leakage following OP-induced SE, but key information regarding time and regional distribution of BBB impairment during the epileptogenic period is missing. To address this data gap, we characterized the spatiotemporal progression of BBB impairment during the first week post-exposure in a rat model of diisopropylfluorophosphate-induced SE, using MRI and albumin immunohistochemistry. Increased BBB permeability, which was detected at 6 h and persisted up to 7 d post-exposure, was most severe and persistent in the piriform cortex and amygdala, moderate but persistent in the thalamus, and less severe and transient in the hippocampus and somatosensory cortex. The extent of BBB leakage was positively correlated with behavioral seizure severity, with the strongest association identified in the piriform cortex and amygdala. These findings provide evidence of the duration, magnitude and spatial breakdown of the BBB during the epileptogenic period following OP-induced SE and support BBB regulation as a viable therapeutic target for preventing SRS following acute OP intoxication.

Dendritic reorganization in the hippocampus, anterior temporal lobe, and frontal neocortex of lithium-pilocarpine induced Status Epilepticus (SE)

Status Epilepticus (SE) is a distributed network disorder, which involves the hippocampus and extra-hippocampal structures. Epileptogenesis in SE is tightly associated with neurogenesis, plastic changes and neural network reorganization facilitating hyper-excitability. On the other hand, dendritic spines are known to be the excitatory synapse in the brain. Therefore, dendritic spine dynamics could play an intricate role in these network alterations. However, the exact reason behind these structural changes in SE are elusive. In the present study, we have investigated the aforementioned hypothesis in the lithium-pilocarpine treated rat model of SE. We have examined cytoarchitectural and morphological changes using hematoxylin-eosin and Golgi-Cox staining in three different brain regions viz. CA1 pyramidal layer of the dorsal hippocampus, layer V pyramidal neurons of anterior temporal lobe (ATL), and frontal neocortex of the same animals. We observed macrostructural and layer-wise alteration of the pyramidal layer mainly in the hippocampus and ATL of SE rats, which is associated with sclerosis in the hippocampus. Sholl analysis exhibited partial dendritic plasticity in apical and basal dendrites of pyramidal cells as compared to the saline-treated weight-/age-matched control group. These findings indicate that region-specific alterations in dendritogenesis may contribute to the development of independent epileptogenic networks in the hippocampus, ATL, and frontal neocortex of SE rats.

Dynamic effects of miR-20a-5p on hippocampal ripple energy after status epilepticus in rats.

To determine the dynamic effects of miR-20a-5p on hippocampal ripple energy in rats after status epilepticus (SE). A lithium pilocarpine (LiCl-PILO)-induced rat model of status epilepticus (SE) was established, and the rats were divided into the normal control (Control, CTL), epileptic control (PILO), valproic acid (VPA + PILO), miR-20a-5p overexpression lentivirus vector (miR + PILO), sponges blocking lentivirus vector (Sponges + PILO), and scramble sequence negative control (Scramble + PILO) groups (n = 6). Electroencephalograms (EEGs) were used to analyze changes in hippocampal ripple energy before and after SE. Quantitative polymerase chain reaction (q-PCR) analysis showed that miR-20a-5p levels gradually increased after miR-20a-5p overexpression lentivirus vector injection into the lateral ventricle, and the miR-20a-5p levels were significantly higher than that in CTL group on days 7 and 36 (P < 0.001). The miR-20a-5p levels decreased significantly on days 7 and 36 after blocking by sponges lentivirus vector injected into the lateral ventricle (P < 0.001). After injection of PILO, the average ripple energy expression in each group gradually increased, and reached the peak before chloral hydrate injection (compared with 1day before SE, P < 0.05). The ripple energy in the VPA + PILO and Sponges + PILO groups was significantly lower than that in the PILO group at 60min and 70min after PILO injection and before chloral hydrate injection (P < 0.05), and maintained lower until 2h after chloral hydrate injection in VPA + PILO (P < 0.05). Compared with the VPA + PILO group, the mean ripple energy of the Sponges + PILO group had no difference at all time points (P ≥ 0.05). After SE, ripple distribution of space and energy is closely related to the occurrence of epilepsy. Inhibition of miR20a-5p expression can downregulate ripple oscillation energy during seizure.

Comparison of intranasal and intraperitoneal administration of Eugenia caryophyllata (clove) essential oil on spatial memory, anxiety-like behavior and locomotor activity in a pilocarpine-induced status epilepticus rat model

BackgroundEpilepsy induces behavioral effects and histological changes in the hippocampus. Eugenol, the main component of clove essential oil, has modulatory effects on seizure. We aimed to investigate the effect of intraperitoneal (IP) and intranasal (IN) clove essential oil on cognitive and histological changes during the chronic phase of temporal lope epilepsy.MethodsMale Wistar rats were divided into eight groups of seven including control, pilocarpine (PLC), clove oil (IP and IN), sesame oil (IP and IN), phenobarbital (positive control) and saline. Rats were injected with 350 mg/kg PLC to induce status epilepticus (SE). We evaluated the effects of 14 days IP (0.1 ml/kg) and IN (0.02 ml/kg) administration of clove essential oil on locomotor/explorative activity, anxiety-like behavior, spatial recognition memory, and hyperexcitability, as well as hippocampal cell survival in PLC-treated rats.ResultsOur findings indicated that clove oil could effectively ameliorate PLC-induced behavioral deficits, and also alleviate neuronal death in the cornu ammonis 1 (CA1) region of the hippocampus. Behavioral results as in the Y-maze, Open field and elevated plus maze featured significant differences between control and treated groups. Post-seizure behavioral battery (PBSS) results explicated that behavioral hyperexcitability were less in clove oil groups (both IN and IP) compared to PLC-treated rats. Moreover, results of this study demonstrated that IN administration of clove oil was more potent in alleviating behavioral impairment at a lower dosage than by IP route. The results of this study, also demonstrated that intranasal administration of clove oil could reduce duration of recurrent seizures.ConclusionIn summary, clove oil treatment ameliorated histopathological and behavioral consequences of PLC-induced SE.

Downregulation of miR-210 Promoted Apoptosis of Hippocampal Neurons by Negatively Regulating the TLR4/NF-кB1 Signaling Pathway in a Rat Model of Status Epilepticus.

PurposeStatus epilepticus (SE) is a life-threatening condition causing brain damage, hippocampal necrosis and apoptosis. This study aimed to determine whether microRNA-210 regulates seizure and apoptosis by targeting the TLR4 /NF-κB1 associated signaling pathway.MethodsIn a pilocarpine-induced epileptic rat model, the expressions of microRNA-210 (miR-210), TLR4, NF-κB1 and caspase-3 were assessed by a quantitative polymerase chain reaction and Western blotting. Tunel detects hippocampal neuron apoptosis.ResultsWe found that miR-210, TLR4, NF-κB1 and caspase-3 were upregulated in the hippocampus of the rat model compared with that of control. The knockdown of miR-210 significantly restored the expression levels of TLR4, NF-κB1 and caspase-3 and increased hippocampal apoptosis.ConclusionThese findings showed that the downregulation of miR-210 promoted apoptosis of hippocampal neurons by negatively regulating the TLR4/NF-кB1 signaling pathway.

Long non-coding RNA Tug1 regulates inflammation in microglia and in status epilepticus rats through the NF-κB signaling pathway

BackgroundInflammation plays an important role in the pathogenesis of status epilepticus (SE). The long non-coding RNA (lncRNA) taurine up-regulated gene1 (Tug1) plays a well-defined role in inflammatory diseases. However, the molecular mechanism of Tug1 in SE progression remains unknown. In present study, we investigated whether Tug1 is involved in microglial inflammation in SE rats.MethodsThe SE rat model was established via intraperitoneal injection of lithium chloride-pilocarpine. RNA-binding protein immunoprecipitation (RIP) and RIP sequencing were carried out in rat microglia (RM). Tug1 cloned into the adenovirus was overexpressed in the microglia. Knockdown of Tug1 was performed via siRNA transfection. The level of Tug1 and inflammatory factors IL-1β and TNF-α was examined by real-time polymerase chain reaction (RT-PCR) and western blotting. Protein levels of p65, p-p65, p-ΙκΒα and ΙκΒα were assessed by western blotting.ResultsThe RIP-seq result showed 14 lncRNAs that bound to the NF-κB p65 protein in RM. The lncRNA Tug1 directly interacted with p65. The level of declined Tug1 was decreased in the hippocampus of SE rats. Overexpression of Tug1 reduced the LPS-induced inflammation and M1/M2 polarization of microglia, while knockdown of Tug1 aggravated the inflammatory response in microglia. Accordingly, the protein levels of p-p65/p65 and p-ΙκΒα/ΙκΒα were reduced in the Tug1-overexpression microglia and elevated in the Tug1-knockdown microglia.ConclusionsThese findings indicate that Tug1 modulates the inflammation in microglia through the NF-κB signal pathway, and the Tug1/P65 axis are like to play a significant role in the inflammatory processes, providing a valid target for the therapy of SE.

Ameliorating effect of ketogenic diet on acute status epilepticus: Insights into biochemical and histological changes in rat hippocampus.

This study aimed to evaluate the potential neuroprotective effects of ketogenic diet (KD) against the neuronal disruptions induced by SE in lithium-pilocarpine rat model of status epilepticus (SE). Four groups of female rats include; groups I and III received standard diet and groups II and IV received KD for 3 weeks. Groups I and II were left untreated, while groups III and IV were injected with LiCl (127 mg/kg, i.p.) followed by pilocarpine HCl (10mg/kg, i.p.) 18-24h later, repeatedly, till induction of SE. 72 h post-SE, KD effectively ameliorated the balance between excitatory (glutamate) and inhibitory (GABA) neurotransmitters and the oxidative stress indices, increased adenine nucleotides and decreased immunoreactivity of iNOS, TNFα, glial fibrillary acidic protein, and synaptophysin. Thiswas in association with improvement in inflammatory response and neuronal tissue characteristics in hippocampus of SE rats. Histological changes showed preservation of neuronal integrity. These findings highlight the protective effects of KD in the acute phase post-SE via ameliorating biochemical and histological changes involved. PRACTICAL APPLICATIONS: Epilepsy is the fourth most common neurological disorder that requires lifelong treatment. It stigmatizes patients and their families. The use of the ketogenic diet (KD) as a therapy for epilepsy developed from observations that fasting could reduce seizures. From 1920s, the KD was a common epilepsy treatment until it was gradually superseded by anticonvulsant drugs so that by the 1980s it was rarely used. However, there has been a resurgence of interest and usage of the KD for epilepsy since the turn of the century. Despite its long history, the mechanisms by which KD exhibits its anti-seizure action are not fully understood. Our study aims to identify the mechanism of KD which may help further studies to achieve the same benefits with a drug or supplement to overcome its unpalatability and gastrointestinal side effects.

Altered Expression of Par3, aPKC-λ, and Lgl1 in Hippocampus in Kainic Acid-Induced Status Epilepticus Rat Model

Introduction: Mossy fiber sprouting (MFS) is a frequent histopathological finding in temporal lobe epilepsy (TLE) and is involved in the pathology of TLE. However, molecular signals underlying MFS remain unclear. Partitioning defective 3(Par3), atypical protein kinase C-λ(aPKC-λ), and lethal giant larvae 1(Lgl1) were involved in the neuronal polarity and axon growth. The potential roles of those proteins in MFS and epileptogenesis of TLE were investigated.Material and Methods: The epileptic rat models were established by intracerebroventricular injection of kainic acid (KA). The degree of MFS was measured by using Timm staining, Neuronal loss and the expression aPKC-λ, Par3, and Lgl1 in hippocampus were measured by using immunohistochemistry and western blot analysis.Results: The neuronal loss in CA3 region was observed from 3 days to 8 weeks, while the neuronal loss in the hilar region was observed from 1 to 8 weeks in experimental group. The Timm score in the CA3 region in experimental group was significantly higher than that in the control group from 2 to 8 weeks. Compared with control group, the expressions of Par3 and Lgl1 were upregulated and the expression of aPKC-λ was downregulated in the experimental groups. Positive correlation between the Par3 expression and Timm scores, and the negative correlation between the aPKC-λ expression and Timm scores in CA3 region were discovered in experimental group.Conclusion: The findings of the present study indicated that aPKC-λ, Par3, and Lgl1 may be involved in MFS and in the epileptogenesis of temporal lobe epilepsy.

TPPU Pre-Treatment Rescues Dendritic Spine Loss and Alleviates Depressive Behaviours during the Latent Period in the Lithium Chloride-Pilocarpine-Induced Status Epilepticus Rat Model

Epileptogenesis may be responsible for both of recurrent seizures and comorbid depression in epilepsy. Disease-modifying treatments targeting the latent period before spontaneous recurrent seizures may contribute to the remission of seizures and comorbid depression. We hypothesized that pre-treatment with 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU), a soluble epoxide hydrolase (sEH) inhibitor, which has anti-inflammatory and neuroprotective effects might rescue status epilepticus (SE)-induced dendritic spine loss and alleviate depressive behaviours. Rats were either pre-treated with TPPU (0.1 mg/kg/d) intragastrically or with vehicle (40% polyethylene glycol 400) from 7 days before to 7 days after SE that was induced with lithium chloride and pilocarpine intraperitoneally. Rats in the Control group were given saline instead. The forced swim test (FST) was performed on the 8th day after SE to evaluate the depression-like behaviours in rats. The results showed that seizures severity during SE was significantly decreased, and the immobility time during FST was significantly increased through TPPU pre-treatment. Moreover, pre-treatment with TPPU attenuated inflammations including microglial gliosis and the level of proinflammatory cytokine IL-1β in the hippocampus; in addition, neuronal and dendritic spine loss in the subfields of hippocampus was selectively rescued, and the expression of NR1 subunit of N-methyl-D-aspartate (NMDA) receptor, ERK1/2, CREB, and their phosphorylated forms involved in the dendritic spine development were all significantly increased. We concluded that pre-treatment with TPPU attenuated seizures severity during SE and depressive behaviours during the period of epileptogenesis probably by rescuing dendritic spine loss in the hippocampus.

Effect of gap junction blockers on hippocampal ripple energy expression in rats with status epilepticus.

To study the effect of gap junction blockers, quinine (QUIN) and carbenoxolone (CBX), on hippocampal ripple energy expression in rats with status epilepticus (SE). A total of 24 rats were randomly divided into four groups: model, QUIN, valproic acid (VPA), and CBX (n=6 each). A rat model of SE induced by lithium-pilocarpine (PILO) was prepared. The QUIN, VPA, and CBX groups were given intraperitoneal injection of QUIN (50 mg/kg), VPA by gavage (200 mg/kg), and intraperitoneal injection of CBX (50 mg/kg) respectively, at 3 days before PILO injection. Electroencephalography was used to analyze the change in hippocampal ripple energy before and after modeling, as well as before and after chloral hydrate injection to control seizures. Ripple expression was observed in the hippocampal CA1, CA3, and dentate gyrus regions of normal rats. After 10 minutes of PILO injection, all groups had a gradual increase in mean ripple energy expression compared with 1 day before modeling, with the highest expression level before chloral hydrate injection in the model, VPA and CBX groups (P<0.05). The QUIN group had the highest expression level of mean ripple energy 60 minutes after PILO injection. The mean ripple energy returned to normal levels in the three intervention groups immediately after chloral hydrate injection, while in the model group, the mean ripple energy returned to normal levels 1 hour after chloral hydrate injection. The mean ripple energy remained normal till to day 3 after SE in the four groups. The changing trend of maximum ripple energy was similar to that of mean ripple energy. The change in ripple energy can be used as a quantitative indicator for early warning of seizures, while it cannot predict seizures in the interictal period. Gap junction blockers can reduce ripple energy during seizures.

Characterization of metabolic activity induced by kainic acid in adult rat whole brain at the early stage: A 18FDG-PET study

ObjectiveBrain metabolic processes are not fully characterized in the kainic acid (KA)-induced Status Epilepticus (KASE). Thus, we evaluated the usefulness of 18F-fluorodeoxyglucose positron emission tomography (FDG-PET) as an experimental strategy to evaluate in vivo, in a non-invasive way, the glucose consumption in several brain regions, in a semi-quantitative study to compare and to correlate with data from electroencephalography and histology studies. Methods.Sixteen male Wistar rats underwent FDG-PET scans at basal state and after KA injection. FDG-PET images were normalized to an MRI-based atlas and segmented to locate regions. Standardized uptake values (SUV) were obtained at several time points. EEGs and cell viability by histological analysis, were also evaluated. ResultsFDG-PET data showed changes in regions such as: amygdala, hippocampus, accumbens, entorhinal cortex, motor cortex and hypothalamus. Remarkably, hippocampal hypermetabolism was found (mean SUV = 2.66 ± 0.057) 2 h after KA administration, while hypometabolism at 24 h (mean SUV = 1.83 ± 0.056) vs basal values (mean SUV = 2.19 ± 0.057). EEG showed increased spectral power values 2 h post-KA administration. Hippocampal viable-cell counting 24 h after KA was decreased, while Fluoro-Jade B-positive cells were increased, as compared to control rats, coinciding with the hypometabolism detected in the same region by semi-quantitative FDG-PET at 24 h after KASE. ConclusionsPET is suitable to measure metabolic brain changes in the rat model of status epilepticus induced by KA (KASE) at the first 24 h, compared to that of EEG; PET data may also be sensitive to cell viability.

Anticonvulsant and neuroprotective effects of carbamazepine-levetiracetam adjunctive treatment in convulsive status epilepticus rat model: Inhibition of cholinergic transmission

This study evaluated the anticonvulsant and neuroprotective effects of carbamazepine (CBZ), levetiracetam (LEV), and CBZ + LEV adjunctive treatment in convulsive status epilepticus (CSE) rat model.Twenty-five male Wistar rats were randomized into five groups (n = 5). Groups I and II received 0.2 ml of normal saline intraperitoneally (i.p), while groups III-V received CBZ (25 mg/kg i.p), LEV (50 mg/kg i.p) or combination of sub-therapeutic doses of CBZ (12.5 mg/kg i.p) and LEV (25 mg/kg i.p). Thirty minutes later, seizure was kindled with pilocarpine hydrochloride (350 mg/kg) in group II-V rats. Seizure indices, markers of excitotoxicity, and astroglioses were determined, while the hippocampal morphometry was also evaluated. The data was analysed using descriptive and inferential statistics, while the results were presented as mean ± SEM in graphs or tables, and the level of significance was taken at p < 0.05.The anticonvulsant treatments delayed the inception of seizure indices (p = 0.0006), while the percentage mortality decreased significantly (p = 0.0001) in all the treatment groups. The hippocampal concentrations of acetylcholine, malondialdehyde, and tissue necrotic factor-alpha decreased significantly (p = 0.0077) in all the treated group relative to the positive control. The reactive astrogliosis in the hippocampus (CA 1) increased significantly (p = 0.0001) compared with the control but abrogated in all the treatment groups relative to the positive control.The anticonvulsant and neuroprotective effects are in this order: LEV < CBZ + CBZ < CBZ. The drug efficacy is attributable to the inhibition of cholinergic transmission.

Seizure-mediated iron accumulation and dysregulated iron metabolism after status epilepticus and in temporal lobe epilepsy

Neuronal dysfunction due to iron accumulation in conjunction with reactive oxygen species (ROS) could represent an important, yet underappreciated, component of the epileptogenic process. However, to date, alterations in iron metabolism in the epileptogenic brain have not been addressed in detail. Iron-related neuropathology and antioxidant metabolic processes were investigated in resected brain tissue from patients with temporal lobe epilepsy and hippocampal sclerosis (TLE-HS), post-mortem brain tissue from patients who died after status epilepticus (SE) as well as brain tissue from the electrically induced SE rat model of TLE. Magnetic susceptibility of the presumed seizure-onset zone from three patients with focal epilepsy was compared during and after seizure activity. Finally, the cellular effects of iron overload were studied in vitro using an acute mouse hippocampal slice preparation and cultured human fetal astrocytes. While iron-accumulating neurons had a pyknotic morphology, astrocytes appeared to acquire iron-sequestrating capacity as indicated by prominent ferritin expression and iron retention in the hippocampus of patients with SE or TLE. Interictal to postictal comparison revealed increased magnetic susceptibility in the seizure-onset zone of epilepsy patients. Post-SE rats had consistently higher hippocampal iron levels during the acute and chronic phase (when spontaneous recurrent seizures are evident). In vitro, in acute slices that were exposed to iron, neurons readily took up iron, which was exacerbated by induced epileptiform activity. Human astrocyte cultures challenged with iron and ROS increased their antioxidant and iron-binding capacity, but simultaneously developed a pro-inflammatory phenotype upon chronic exposure. These data suggest that seizure-mediated, chronic neuronal iron uptake might play a role in neuronal dysfunction/loss in TLE-HS. On the other hand, astrocytes sequester iron, specifically in chronic epilepsy. This function might transform astrocytes into a highly resistant, pro-inflammatory phenotype potentially contributing to pro-epileptogenic inflammatory processes.

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

ObjectiveThe 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.MethodsRats 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.ResultsAlthough 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.SignificanceNeither 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.