Status Epilepticus Group Research Articles

Effects of baicalin on the expression of Toll-like receptor 2/4 signaling pathway in kainate-induced epileptic mice

Objective To study the effects of baicalin on the expression of Toll-like receptor 2/4 (TLR2/4) signaling pathway in Kainate-induced epileptic mice, and explore the neuroprotective mechanism of baicalin after status epilepticus (SE). Methods Forty-eight ICR male mice were randomly divided into three groups: control group, SE group and baicalin group, to which baicalin was administered at doses of 100 mg/kg. To establish animal model of SE, 12 mg/kg kainic acid was administered to mice by intraperitoneal injection. Hematoxylin and eosin (HE) stain was used to observe the pathological changes. Nissl staining was used to assess the necrosis of hippocampal neurons. Immunohistochemistry was used to observe the positive expression of TLR2 and TLR4. Reverse transcription polymerase chain reaction (RT-PCR) and Western blot was used to detect the expression of the mRNA and protein 1eveI of TLR2, TLR4 and nuclear factor-κB (NF-κB). Results Baicalin can significantly relieve the injury and inhibit granule cells losts (114.27±15.53), and at the same time decreas the expression of TLR2, TLR4 and NF-κB (P=0.043). Conclusion TLR2/4 signaling pathway could be activated after SE and Baicalin could inhibit TLR2/4-mediated signaling pathway to protect the hippocampal neurons after SE in mice. Key words: Epilepsy; Kainic acid; Baicalin; Toll-like receptor 2; Toll-like receptor 4

Alterations in the Neurobehavioral Phenotype and ZnT3/CB-D28k Expression in the Cerebral Cortex Following Lithium-Pilocarpine-Induced Status Epilepticus: the Ameliorative Effect of Leptin.

Zinc transporter 3 (ZnT3)-dependent "zincergic" vesicular zinc accounts for approximately 20% of the total zinc content of the mammalian telencephalon. Elevated hippocampal ZnT3 expression is acknowledged to be associated with mossy fiber sprouting and cognitive deficits. However, no studies have compared the long-term neurobehavioral phenotype with the expression of ZnT3 in the cerebral cortex following status epilepticus (SE). The aim of this study was to investigate changes in the long-term neurobehavioral phenotype as well as the expression of ZnT3 and calcium homeostasis-related CB-D28k in the cerebral cortex of rats subjected to neonatal SE and to determine the effects of leptin treatment immediately after neonatal SE. Fifty Sprague-Dawley rats (postnatal day 6, P6) were randomly assigned to two groups: the pilocarpine hydrochloride-induced status epilepticus group (RS, n = 30) and control group (n = 20). Rats were further divided into the control group without leptin (Control), control-plus-leptin treatment group (Leptin), RS group without leptin treatment (RS), and RS-plus-leptin treatment group (RS + Leptin). On P6, all rats in the RS group and RS + Leptin group were injected intraperitoneally (i.p.) with lithium chloride (5mEq/kg). Pilocarpine (320mg/kg, i.p.) was administered 30min after the scopolamine methyl chloride (1mg/kg) injection on P7. From P8 to P14, animals of the Leptin group and RS + Leptin group were given leptin (4mg/kg/day, i.p.). The neurological behavioral parameters (negative geotaxis reaction reflex, righting reflex, cliff avoidance reflex, forelimb suspension reflex, and open field test) were observed from P23 to P30. The protein levels of ZnT3 and CB-D28k in the cerebral cortex were detected subsequently by the western blot method. Pilocarpine-treated neonatal rats showed long-term abnormal neurobehavioral parameters. In parallel, there was a significantly downregulated protein level of CB-D28k and upregulated protein level of ZnT3 in the cerebral cortex of the RS group. Leptin treatment soon after epilepticus for 7 consecutive days counteracted these abnormal changes. Taken together with the results from our previous reports on another neonatal seizure model, which showed a significant positive inter-relationship between ZnT3 and calcium/calmodulin-dependent protein kinase IIα (CaMKIIα), the data here suggest that ZnT3/CB-D28k-associated Zn (2+)/Ca(2+) signaling might be involved in neonatal SE-induced long-term brain damage in the aspects of neurobehavioral impairment. Moreover, consecutive leptin treatment is effect at counteracting these hyperexcitability-related changes, suggesting a potential clinical significance.

Status epilepticus triggers long-lasting activation of complement C1q-C3 signaling in the hippocampus that correlates with seizure frequency in experimental epilepsy

Status epilepticus (SE) triggers a myriad of neurological alterations that include unprovoked seizures, temporal lobe epilepsy (TLE), and cognitive deficits. Although SE-induced loss of hippocampal dendritic structures and synaptic remodeling are often associated with this pathophysiology, the underlying mechanisms remain elusive. Recent evidence points to the classical complement pathway as a potential mechanism. Signaling through the complement protein C1q to C3, which is cleaved into smaller biologically active fragments including C3b and iC3b, contributes to the elimination of synaptic structures in the normal developing brain and in models of neurodegenerative disorders. We recently found increased protein levels of C1q and iC3b fragments in human drug-resistant epilepsy. Thus, to identify a potential role for C1q-C3 in SE-induced epilepsy, we performed a temporal analysis of C1q protein levels and C3 cleavage in the hippocampus along with their association to seizures and hippocampal-dependent cognitive functions in a rat model of SE and acquired TLE. We found significant increases in the levels of C1q, C3, and iC3b in the hippocampus at 2-, 3- and 5-weeks after SE relative to controls (p<0.05). In the SE group, greater iC3b levels were significantly correlated with higher seizure frequency (p<0.05). Together, these data support that hyperactivation of the classical complement pathway after SE parallels the progression of epilepsy. Future studies will determine whether C1q-C3 signaling contributes to epileptogenic synaptic remodeling in the hippocampus.

Differences in the Levels of Interleukin-1β (IL-1β) and Interleukin-1 Receptor Antagonist (IL-1ra) in Children with Status Epilepticus and Febrile Seizure

Proinflammatory cytokines are elevated in status epilepticus and febrile seizure and associated with tissue damage. This study aimed to investigate the differences in interleukin 1 beta (IL-1β) and interleukin 1 receptor agonist (IL-1ra) levels in status epilepticus as compared with febrile seizure and febrile. This cross-sectional study was designed to include 45 subjects divided equally into three groups (status epilepticus, febrile seizure, and febrile). Both IL-1β and IL-1ra were measured by using an ELISA method. Results showed that IL-1β levels were significantly higher in the status epilepticus group as compared with the febrile seizure and febrile groups (p < 0.05). IL-1ra levels in the status epilepticus group were significantly lower compared with the febrile seizure group (p = 0.04). Consistently, the IL-1β/IL-1ra ratio in the status epilepticus group was significantly higher as compared with the febrile seizure group (p = 0.01). We concluded that IL-1β and the IL-1β/IL-1ra ratio were significantly higher in status epilepticus. IL-1ra levels were significantly higher in the febrile seizure group.

Novel insights into the effect of paroxetine administration in pilocarpine‑induced chronic epileptic rats.

The aim of the present study was to investigate the role of paroxetine intervention in epilepsy, and its association with the expression of serotonin transporter (SERT) and hippocampal apoptosis. Thirty adult male Sprague Dawley rats were divided into control vehicle (n=6) and epileptic (n=24) groups. Status epilepticus (SE) was induced via systemic injection of pilocarpine, and seizure activity was monitored via video electroencephalogram. The epileptic group was then randomly divided into two groups; Four weeks following SE induction, paroxetine (5mg/kg/day; SE+paroxetine group) or normal saline (SE group) was intraperitoneally injected for 4weeks. Brain tissue was collected to evaluate apoptosis via terminal deoxynucleotidyl transferase dUTP nick‑end labeling. SERT, B‑cell lymphoma‑2 (Bcl‑2) and brain derived neurotropic factor (BDNF) expression levels were evaluated by western blotting, and miR‑16 expression was evaluated by reverse transcription‑quantitative polymerase chain reaction. Paroxetine did not affect the mortality of the pilocarpine‑induced chronic epileptic rats. Spontaneous recurrent seizures (SSRs) were observed 7‑28days following SE induction. The frequency and stage of the SSRs were reduced by paroxetine administration. Apoptotic cells were observed in the epileptic hippocampus. Following paroxetine intervention, the staining intensity and number of apoptotic cells were significantly decreased. Expression levels of BDNF and Bcl‑2 were lower in the SE group compared with the vehicle group. The former was not altered by paroxetine injection; however, the latter was increased. In the SE group, SERT expression was not altered in the raphe nucleus but was decreased in the hippocampus. Following paroxetine administration, SERT expression was decreased in the raphe nucleus and increased in the hippocampus. In the SE group, miR‑16 expression was decreased in the raphe nucleus and increased in the hippocampus. Following paroxetine administration, miR‑16 expression was not altered in the raphe nucleus but was reduced in the hippocampus. In conclusion, the seizures and hippocampal apoptosis observed in chronic epileptic rats were alleviated by paroxetine treatment. This effect may be associated with the reduced Bcl‑2 and BDNF expression and the modulation of SERT expression. The alterations in miR‑16 expression may provide a potential explanation for the modulation of apoptosis; however, further research is required to determine the complete underlying molecular mechanism.

The role of necroptosis in status epilepticus-induced brain injury in juvenile rats.

To study the role of necroptosis in status epilepticus (SE)-induced injury in the developing brain and the possible associations of necroptosis with epileptogenesis and cognitive dysfunction. The lithium-pilocarpine epilepsy model was reproduced in male rats at postnatal day 25. Propidium iodide (PI) staining was used to detect cell death after SE. Transmission electron microscopy (TEM) was performed to observe morphological changes in injured neurons. Western blot and immunofluorescence (IF) staining were used to investigate the expression of receptor interacting protein kinase-3 (RIP3), mixed lineage kinase domain-like (MLKL), and p-MLKL after SE. EEG was monitored during the chronic epileptic period. The Morris water maze test was performed to evaluate spatial learning and memory in juvenile rats after SE. Massive PI-positive (PI+) neurocytes were observed mainly in the amygdala and piriform cortex 24h to 7days after SE, with the most prominent changes observed after 72h. Injured neurons observed via TEM exhibited necroptotic morphological features, including loss of ribosomes, autophagosome formations, deformed nuclei with condensed and marginated chromatin, and disruptive cell membranes. The expression of RIP3 and p-MLKL increased after 24h, peaked at 72h, and decreased 7days after SE. In addition, IF staining revealed that MLKL was expressed in cell plasma membranes present in the amygdala and piriform cortex. This finding was concomitant with the fact that MLKL is involved in executing necroptosis by binding and disrupting the plasma membrane. During the chronic epileptic period, spontaneous recurrent seizures were observed behaviorally and interictal spikes and sharp waves were recorded by EEG in the SE group. The Morris water maze test revealed that in the place navigation test, the escape latency of the SE group was longer than that of the control group (p<0.05). In the spatial probe test, the number of times the rats in the SE group passed through the original platform site was lesser than that of the rats in the control group (p<0.05). SE-induced brain injury leads to neuronal necroptosis in juvenile rats. MLKL may play a significant role in the execution of SE-induced necroptosis. Further studies are required to determine whether inhibiting necroptosis can prevent chronic epileptogenesis and improve cognitive ability for juvenile rats.

Epilepsy is associated with ventricular alterations following convulsive status epilepticus in children.

SummaryObjectiveConvulsive status epilepticus can exert profound cardiovascular effects in adults, including ventricular depolarization–repolarization abnormalities. Whether status epilepticus adversely affects ventricular electrical properties in children is less understood. Therefore, we sought to characterize ventricular alterations and the associated clinical factors in children following convulsive status epilepticus.MethodsWe conducted a 2‐year retrospective case–control study. Children between 1 month and 21 years of age were included if they were admitted to the pediatric intensive care unit with primary diagnosis of convulsive status epilepticus and had 12‐lead electrocardiogram (ECG) within 24 h of admission. Children with heart disease or ion channelopathy, or who were on vasoactive medications were excluded. Age‐matched control subjects had no history of seizures or epilepsy. The primary outcome was ventricular abnormalities represented by ST segment changes, abnormal T wave, QRS axis deviation, and corrected QT (QTc) interval prolongation. The secondary outcomes included QT/RR relationship, beat‐to‐beat QTc interval variability, ECG interval measurement between groups, and clinical factors associated with ECG abnormalities.ResultsOf 317 eligible children, 59 met the inclusion criteria. History of epilepsy was present in 31 children (epileptic) and absent in 28 children (nonepileptic). Compared with the control subjects (n = 31), the status epilepticus groups were more likely to have an abnormal ECG, with overall odds ratios of 3.8 and 7.0 for the nonepileptic and the epileptic groups, respectively. Simple linear regression analysis demonstrated that children with epilepsy exhibited impaired dependence and adaptation of the QT interval on heart rate. Beat‐to‐beat QTc interval variability, a marker of ventricular repolarization instability, was increased in children with epilepsy.SignificanceConvulsive status epilepticus can adversely affect ventricular electrical properties and stability in children, especially those with epilepsy. These findings suggest that children with epilepsy may be particularly vulnerable to seizure‐induced arrhythmias. Therefore, postictal cardiac surveillance may be warranted in this population.

The effect of metformin treatment on endoplasmic reticulum (ER) stress induced by status epilepticus (SE) via the PERK-eIF2α-CHOP pathway.

Status epilepticus (SE) is defined as continuous seizure activity lasting more than 5 minutes. It results in neuronal cell death, mediated by endoplasmic reticulum (ER) stress response. Previously, metformin demonstrated neuroprotective effects in primary cortical neurons. In this study, we analyzed the effect of metformin on ER stress via the pro-apoptotic protein kinase RNA-like endoplasmic reticulum kinase (PERK)-eukaryotic initiation factor 2α (eIF2α)-C/EBP homologous protein (CHOP) pathway. SE was induced in rats by pentylenetetrazole. Following SE, the rats were treated with salubrinal, GSK2656157, or metformin. In a control group (normal saline) SE was not induced. CHOP, eIF2α, and PERK expression was determined by Western blot; apoptosis was analyzed by TUNEL assay. CHOP expression was significantly increased at 6 and 24 hours following SE. At both time points, eIF2α and PERK levels were also increased. At 6 hours, CHOP expression was significantly reduced in salubrinal, GSK2656157 and metformin groups versus SE group. eIF2α and PERK levels were decreased in metformin compared to SE group. eIF2α expression was markedly decreased in salubrinal versus SE group, while PERK expression was markedly reduced in GSK2656157 versus SE group. At 6 and 24 hours, the apoptosis rate was significantly increased in SE versus control group, while it was significantly reduced in salubrinal, GSK2656157, and metformin groups compared to SE group. The apoptosis rate also decreased in salubrinal group at 24 hours, although not to the extent observed in metformin group. Overall, CHOP expression and apoptosis induced by SE in rats were reduced with metformin. Further studies are required to evaluate the clinical relevance of metformin for patients with SE.

Neuroprotective effects of vitamin D alone or in combination with lamotrigine against lithium-pilocarpine model of status epilepticus in rats.

Status epilepticus (SE) is considered one of the major serious forms of epilepsy with high mortality rate. Since the currently available antiepileptic drugs have low efficacy and high adverse effects, new more efficient and safe therapies are critically needed. There is increasing evidence supporting dietary and alternative therapies for epilepsy, including the ketogenic diet, modified Atkins diet, and omega-3 fatty acids. Recent studies have shown significant prophylactic and therapeutic potential of vitamin D (vit-D) use in many neurological disorders. Therefore, in the present study, the neuroprotective effects and mechanisms of vit-D alone or in combination with lamotrigine have been evaluated in the lithium-pilocarpine model of SE in rats. Rats were divided into five groups: normal group, SE group, lamotrigine (25mg/kg/day) pretreated group, vit-D (1.5mcg/kg/day) pretreated group, and group pretreated with vit-D and lamotrigine for 2weeks. At the end of treatment, SE was induced by single intraperitoneal injection of LiCl (127mg/kg), followed 24h later by pilocarpine (30mg/kg). Seizures' latency, cognitive performance in Morris water maze, brain oxidative stress biomarkers (glutathione, lipid peroxides, and nitric oxide), brain neurochemistry (γ-aminobutyric acid and glutamate), and brain histopathology have been evaluated. Vit-D prevented pilocarpine-induced behavioral impairments and oxidative stress in the brain; these results were improved in combination with lamotrigine. Vit-D has a promising antiepileptic, neuroprotective, and antioxidant effects. It can be provided to patients as a supportive treatment besides antiepileptic drugs. However, clinical trials are needed to establish its efficacy and safety.

Protective Effects of Cannabidiol against Seizures and Neuronal Death in a Rat Model of Mesial Temporal Lobe Epilepsy.

The present study reports the behavioral, electrophysiological, and neuropathological effects of cannabidiol (CBD), a major non-psychotropic constituent of Cannabis sativa, in the intrahippocampal pilocarpine-induced status epilepticus (SE) rat model. CBD was administered before pilocarpine-induced SE (group SE+CBDp) or before and after SE (group SE+CBDt), and compared to rats submitted only to SE (SE group), CBD, or vehicle (VH group). Groups were evaluated during SE (behavioral and electrophysiological analysis), as well as at days one and three post-SE (exploratory activity, electrophysiological analysis, neuron density, and neuron degeneration). Compared to SE group, SE+CBD groups (SE+CBDp and SE+CBDt) had increased SE latency, diminished SE severity, increased contralateral afterdischarge latency and decreased relative powers in delta (0.5–4 Hz) and theta (4–10 Hz) bands. Only SE+CBDp had increased vertical exploratory activity 1-day post SE and decreased contralateral relative power in delta 3 days after SE, when compared to SE group. SE+CBD groups also showed decreased neurodegeneration in the hilus and CA3, and higher neuron density in granule cell layer, hilus, CA3, and CA1, when compared to SE group. Our findings demonstrate anticonvulsant and neuroprotective effects of CBD preventive treatment in the intrahippocampal pilocarpine epilepsy model, either as single or multiple administrations, reinforcing the potential role of CBD in the treatment of epileptic disorders.

SCN1A Does Not Determine Seizure Duration in Children Unaffected by Dravet's Syndrome

A significant proportion of children with seizures present with status epilepticus (SE), early in their disease. Our objective was to evaluate whether SCN1A variants predispose children to SE, outside of Dravet's syndrome. SCN1A analyses were performed in children aged 1 month to 16 years with at least one episode of status epilepticus (group A), and in children with short seizures only (group B). Clinical and genetic findings were compared between both groups. Total 99 children were included: 61 in group A, 38 in group B. SCN1A variants were found in 2 (3.3%) of 61 patients with status epilepticus and in 3 (7.9%) of 38 patients with short seizures. The difference between both groups was −4.6% (95% CI −18 to 5.1), and was not statistically significant ( p = 0.37, Fisher's exact test). In four of these five children, seizures were observed in a GEFS+ familial context. The last patient had a generalized genetic epilepsy not further determined. SCN1A variants were not significantly more frequent in group A than in group B. Their predicted severity was not higher. Our results indicate that SCN1A variants per se do not determine seizure duration in children not affected by Dravet's syndrome.

Which component of treatment is important for changes of cortical epileptic afterdischarges after status epilepticus in immature rats?

Role of lithium chloride and paraldehyde in acute changes after lithium-pilocarpine status epilepticus (SE) induced at postnatal day 12 was studied in 15-day-old rats. In addition to SE group four other groups were formed: naïve animals without any injection, lithium chloride group, paraldehyde group and lithium-paraldehyde group. Cortical epileptic afterdischarges (CxADs) induced by increasing intensities of stimulation current were used as a measure of excitability. SE animals did not exhibit any change in duration of CxADs with increasing stimulation intensity in contrast to naïve control with a progressive prolongation of CxAD. LiCl group was similar to SE rats whereas paraldehyde and lithium-paraldehyde groups exhibited some progress in duration of ADs. Lithium chloride participates in short-term changes of CxADs after SE. Paraldehyde and combination of lithium and paraldehyde are similar to naïve controls.

Protective Effects of Thymoquinone Against Convulsant Activity Induced by Lithium-Pilocarpine in a model of Status Epilepticus.

Inflammation plays a pivotal role in status epilepticus (SE). Thymoquinone (TQ) is a bioactive monomer extracted from black seed (Nigella sativa) oil, which has anti-inflammatory properties in the context of various diseases. This study explored the protective effects of TQ in SE and used a lithium-pilocarpine model of SE to investigate the underlying mechanism, which was related to inflammation mediated by the NF-κB signaling pathway. In the present study, latency to SE increased in the TQ-pretreated group compared with the SE group, and the incidence of SE was significantly reduced. The seizure severity score measured on the Racine scale was significantly decreased in the TQ group compared with the SE group. Moreover, the results of the behavioral tests suggested that TQ may also have a protective effect on learning and memory functions. Finally, we further investigated the protective mechanism of TQ. The results showed that TQ-pretreatment significantly downregulated the protein levels of COX-2 and TNF-α in the brain, in a manner mediated by the NF-κB signaling pathway. These findings demonstrate that TQ attenuates convulsant activity via an anti- inflammation signaling pathway in a model of SE.

Diagnostic value of amplitude-integrated electroencephalogram for epileptic seizures in pediatric intensive care unit

Objective To assess the value of amplitude-integrated electroencephalogram(aEEG) in diagnose of epileptic seizure and status epilepticus in pediatric intensive care unit(PICU). Methods Thirty-five children admitted to Children's Hospital of Fudan University from January to December 2014 with severe neurologia situation in PICU were investigated, and bedside video-EEG(VEEG) findings were recorded for more than 4 hours.VEEG signals were transformed into aEEG signals by Galileo NT PMS software: one-channel aEEG (C3-C4), mutichannel aEEG(F3-F4; C3-C4; P3-P4; T3-T4) plus original EEG.Electrical seizure activity on VEEG was signed out with respect to its occurrence, duration and localization of seizure onset; while aEEG seizure was recorded only during its occurrence.The relationship between aEEG and VEEG was analyzed by Spearman analysis. Results A total of 61 traces in 35 cases were suitable for analysis.(1) gender: 24 male and 11 female; years of age: 12 cases of 2 months-1 year old, 13 cases >1-6 years old, 10 cases >6-12 years old; etiological factors: 15 cases associated with severe viral encephalitis, 6 cases associated with epilepsy, 6 cases associated with intracranial hemorrhage, 5 cases associated with hypoxic-ischemic encephalopathy(HIE), 3 cases with other factors.(2)The results were divided into 2 groups: status epilepticus (SE) group and epileptic seizures group.There were 37 traces in SE group, 19 traces of them were convulsive SE, and severe viral encephalitis and epilepsy were the most common causes.The other 18 traces were non-convulsive SE, the most common causes were severe viral encephalitis, HIE and intracranial hemorrhage.There were 24 traces in non-SE group, and 118 frequencies of epileptic seizures were monitored.(3)The diagnostic value of aEEG for SE and epileptic seizures: 37 traces with status epilepticus on VEEG were all diagnosed as SE on aEEG.For non-SE electrical seizures, the sensitivity of aEEG for detection of electrical seizures was as follows: 79.66%(94/118 traces)for one-channel aEEG, 91.53%(108/118 traces) for multichannel aEEG.Findings from one-channel aEEG and multichannel aEEG were all related to VEEG(ρ=0.837, 0.901, all P<0.01). Conclusions The aEEG is a simple and effective method for monitoring epileptic seizures and SE in PICU.One-channel aEEG is the best method to find SE.It is helpful to increase the numbers of channel in order to find epileptic seizures. Key words: Child; Severe neurology situation; Amplitude-integrated electroencephalogram; Status epilepti-cus; Seizure

Decreased neuron loss and memory dysfunction in pilocarpine-treated rats pre-exposed to hypoxia

Preconditioning can induce a cascade of cellular events leading to neuroprotection against subsequent brain insults. In this study, we investigated the chronic effects of hypoxic preconditioning on spontaneous recurrent seizures (SRS), neuronal death, and spatial memory performance in rats subjected to pilocarpine (Pilo)-induced status epilepticus (SE). Rats underwent a short hypoxic episode (7% O2+93% N2; 30min on two consecutive days) preceding a 4-h SE (HSE group). Control groups were rats submitted to SE only (SE), rats subjected to hypoxia only (H) or normoxia-saline (C). Animals were monitored for the occurrence of SRS, and spatial memory performance was evaluated in the radial-arm maze. Hippocampal sections were analyzed for cell death and mossy fiber sprouting at 1 or 60days after SE. Compared to SE group, HSE had increased SE latency, reduced number of rats with SRS, reduced mossy fiber sprouting at 60days, and reduced cell death in the hilus and the CA3 region 1 and 60days after SE. Additionally, HSE rats had better spatial memory performance than SE rats. Our findings indicated that short hypoxic preconditioning preceding SE promotes long-lasting protective effects on neuron survival and spatial memory.

Does status epilepticus induced at early postnatal period change excitability after cortical epileptic afterdischarges?

Possible changes of cortical excitability after status epilepticus (SE) elicited in 12-day-old rats were studied by means of paired cortical afterdischarges (ADs). Consequences of lithium-pilocarpine status were studied in animals with implanted electrodes 3, 6, 9, 13, and 26 days after SE. Paired low-frequency stimulation with a 1-min interval was repeated after 10 min, and duration of ADs was measured. Control rats received saline instead of pilocarpine; other treatments were the same as in SE group. Postictal refractoriness (i.e., the testing response significantly shorter than the conditioning one) appeared at the age of 18 days in lithium-paraldehyde controls, whereas SE animals exhibited this phenomenon since postnatal day 21. The only significant difference between SE and lithium-paraldehyde controls was found in the second conditioning AD in the oldest group studied-it was longer in 38-day-old SE animals. Our results demonstrated moderate signs of higher excitability of SE rats in comparison with control ones long before appearance of spontaneous seizures.

Hippocampal YKL-40 expression in rats after status epilepticus

ObjectiveThis study aims to investigate rat hippocampal cartilage glycoprotein-39 (YKL-40) expression levels following status epilepticus (SE), and the potential mechanism of YKL-40 in SE-induced neuronal injury. MethodsSeventy-two healthy adult male Sprague-Dawley rats were randomly assigned into two groups: control and SE groups. A lithium chloride-pilocarpine SE model was established, and the control group was injected with sodium chloride. The hippocampus was assessed at three, six, 12, 24 and 72h after SE. YKL-40 protein was detected via immunohistochemistry (n=6). Cellular YKL-40/neuronal nuclei antigen (NeuN) expression levels were determined with double immunofluorescence. YKL-40 mRNA was detected via quantitative real-time polymerase chain reaction (qRT-PCR, n=6). ResultsYKL-40 immunoreactivity was poor and was mainly localized to the cytoplasm within hippocampal CA3/CA4 neurons with limited immunoreactivity in the nucleus in the control group. Following SE, YKL-40 immunoreactivity exhibited a diffused distribution throughout the cytoplasm in neurons and increased distribution in the nucleus. CA3 and CA4 YKL-40 protein expression significantly increased at six hours post-SE (P<0.05), peaked at 12h (P<0.01), and decreased at 24h (P<0.05) and 72h (P<0.05) in the SE group, compared with the control group. Double-label immunofluorescence of YKL-40/NeuN exhibited a hippocampal CA3/CA4 neuron location. Furthermore, SE induced a significant increase inYKL-40 mRNA at six, 24, or 72h (P<0.05); which peaked at 12h (P<0.01). ConclusionsYKL-40 was expressed in neurons, and elevated rat hippocampal YKL-40 expression levels may be involved in the pathogenesis of injury following SE.

Dual mechanisms of rapid expression of anxiety-related behavior in pilocarpine-treated epileptic mice

A mouse model of epilepsy was generated by inducing status epilepticus (SE) for either 1.5 or 4.5h with pilocarpine to study anxiety-related behaviors, changes in the electroencephalogram of the cerebral cortex and hippocampus, and expression of hippocampal proteins. The viability and rate of success of SE induction were high in C57BL/6N mice but not in C57BL/6J mice. C57BL/6N mice were immotile during the first 2days after SE; however, by the third day, most mice were recovered and exhibited strong anxiety-related behaviors in response to the light/dark preference test and open field test. There was a striking difference in the temporal appearance of anxiety-related behavior between the two SE durations: 1.5h SE mice exhibited strong anxiety-related behavior 3days after SE that gradually attenuated over the next few weeks, whereas 4.5h SE mice exhibited strong anxiety-related behavior 3days after SE that persisted even at nearly 1year after SE. Mice receiving both SE durations exhibited generalized seizures (GS) after SE; however, there was a marked difference in the timing and duration of GS appearance. Mice in the 4.5h SE group exhibited spontaneous GS from 4days to at least 96days after SE. In contrast, mice in the 1.5h SE group exhibited GS only within the first several days after SE; however, epileptic spike clusters continuously appeared in the cerebral cortex and hippocampus for up to twelve days after SE. Among the hippocampal proteins tested, only brain derived-neurotrophic factor (BDNF) exhibited altered expression in parallel with anxiety-related behavior. These results showed the possibility that BDNF expression in the hippocampus might cause anxiety-related behavior in adulthood.

Status epilepticus triggers early mitochondrial fusion in the rat hippocampus in a lithium-pilocarpine model

Many reports investigating the hippocampus have demonstrated an increase in neuronal damage, cellular loss, oxidative stress and mitochondrial DNA damage during status epilepticus (SE); however, information regarding alterations in mitochondrial fission and fusion events in SE is lacking. The aim of the present study was to examine the possible imbalance between mitochondrial fission and fusion in the hippocampus of male rats after acute seizure mediated by SE. In this study, we used ninety animals were randomly divided into control and SE groups and subjected to the lithium-pilocarpine model of epilepsy. Hippocampi were obtained at 3, 24 and 72h after SE, and the cytoplasmic and mitochondrial fractions of the cells were used to analyze changes in the Drp1 and Fis1 fission proteins and the Mfn1 and Opa1 fusion proteins by western blot analysis. Moreover, changes in the expression of fission and fusion mRNA transcripts were evaluated by real-time PCR. Mitochondrial morphology was also analyzed using standard transmission electron microscopy. Our data showed that the fission-related mRNA Drp1 was down-regulated rapidly after SE, while Fis1 did not show any significant changes in expression. Moreover, the mitochondrial fusion-associated proteins Mfn1 and Opa1 exhibited an increase in expression at 72h after SE. Electron microphotography revealed several morphological changes, such as swollen mitochondria and damage of the inner mitochondrial membrane, at 24h; at 72h elongation of some mitochondrial was also observed. Our results suggest that after the initiation of SE, the main regulator of the fission mRNA Drp1 is down-regulated, which in turn regulates mitochondrial fission and leads to an increase in the Mfn1 and Opa1 proteins to induce mitochondrial fusion, suggesting an imbalance of the fission and fusion processes.

High frequency oscillations can pinpoint seizures progressing to status epilepticus

Status epilepticus (SE) is defined as a seizure lasting more than 5min or a period of recurrent seizures without recovery between them. SE is a serious emergency condition that requires immediate intervention; therefore, identifying SE electrophysiological markers may translate in prompt care to stop it. Here, we analyzed the EEG signals recorded from the CA3 region of the hippocampus and the entorhinal cortex in rats that responded to systemic administration of 4-aminopyridine (4AP) by generating either isolated seizures or seizures progressing to SE. We found that high frequency oscillations (HFOs) – which can be categorized as ripples (80–200Hz) and fast ripples (250–500Hz) – had different patterns of occurrence in the two groups (n=5 for each group). Specifically, fast ripples in CA3 and entorhinal cortex of the SE group occurred at higher rates than ripples, both during the ictal and post-ictal periods when compared to the HFOs recorded from the isolated seizure group. Our data reveal that different patterns of HFO occurrence can pinpoint seizures progressing to SE, thus suggesting the involvement of different neuronal networks at the termination of seizure discharges.