lithium-chloride and Status-Epilepticus

Status epilepticus (SE) promotes neuronal proliferation and differentiation in the adult and developing rodent hippocampus. However, the effect of SE on other neurogenic brain regions such as the cerebellum has been less explored. To determine whether SE induced by pentylentetrazole (PTZ-SE) and lithium-pilocarpine (Li-Pilo-SE) increases cell proliferation and neurogenesis in the developing rat cerebellum. SE was induced in 14-day-old (P14) Wistar rat pups (both sexes). One hour after SE and the following day rats were injected intraperitoneally with 5-bromo-2'-deoxyuridine (BrdU, 50 mg/kg). Seven days after SE, immunohistochemistry was performed to detect BrdU-positive (BrdU+) cells or BrdU/NeuN+ cells in the cerebellar vermis. SE induced by PTZ or Li-Pilo statistically significant increased the number of cerebellar BrdU+ cells when compared with the control group (58% and 40%, respectively); maximal cell proliferation occurred in lobules II, III, VIb, VIc, VIII, IXa, and IXb of PTZ-SE group and II, V, VIc, VII, and X of Li-Pilo-SE group. An increased number of BrdU/NeuN+ cells was detected in lobules V (17 ± 1.9), VIc (25.8 ± 2.7), and VII (26.2 ± 3.4) after Li-Pilo-SE compared to their control group (9.8 ± 1.7, 12.8 ± 2.8, and 11 ± 1.7, respectively), while the number of BrdU/NeuN+ cells remained the same after PTZ-induced SE or control conditions. SE induced in the developing rat by different experimental models increases cell proliferation in the granular layer of the cerebellar vermis, but only SE of limbic seizures increases neurogenesis in specific cerebellar lobes.

Topics: Animals; Cell Proliferation; Cerebellum; Female; Lithium Chloride; Male; Neurogenesis; Pentylenetetrazole; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Seizure-induced neurogenesis has a widely recognized pro-epileptogenic function. Given the critical role of Notch signaling during the maintenance and neurogenesis of neural stem cells, we hypothesized that Notch may affect epileptogenesis and its progression through its role in neurogenesis in the adolescent rat brain. We used the lithium-pilocarpine-induced epilepsy model in adolescent Sprague-Dawley rats in order to evaluate hippocampal neurogenesis and epileptogenesis following the onset of status epilepticus (SE). We used western blotting analyses and qPCR to measure levels of Notch signaling at different phases after seizures and immunofluorescence to detect the proliferation and differentiation of neural stem cells after seizure. Following the administration of DAPT, a Notch γ-secretase inhibitor, into the lateral ventricles, we observed a suppression of abnormal neurogenesis in the acute phase and a reduction of gliosis in the chronic phase after SE. Accordingly, the frequency and duration of spontaneous seizures in chronic phase were decreased. Our results clarify the basic concept regarding the involvement of Notch signaling in the regulation of hippocampal neurogenesis and epileptogenesis, thereby potentially offering a novel and alternative treatment for epilepsy.

Topics: Animals; Cell Differentiation; Disease Models, Animal; Epilepsy; Hippocampus; Lithium Chloride; Neural Stem Cells; Neurogenesis; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptor, Notch1; Signal Transduction; Status Epilepticus

Topics: Analgesics, Opioid; Animals; Anticonvulsants; Dose-Response Relationship, Drug; Indazoles; Ligands; Lithium Chloride; Male; Morphine; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Pentylenetetrazole; Pilocarpine; Rats; Rats, Wistar; Signal Transduction; Status Epilepticus

Mood and anxiety disorders, as well as memory impairments, are important factors affecting quality of life in patients with epilepsy and can influence the antiepileptic therapy. Clinical studies of psychiatric comorbidities are quite complicated to design and interpret, so animal studies of behavioral impairments associated with seizures can be of use. We investigated the effect of early administration of endocannabinoid receptor agonist WIN-55,212-2 on the development of spontaneous seizures, long-term behavioral and memory impairments, and neurodegeneration in the hippocampus on the lithium-pilocarpine model of status epilepticus (SE). We also studied the role of spontaneous seizures in the development of pathologic consequences of the SE. Our results showed that behavioral impairments found in the elevated plus maze test depended mostly on the consequences of SE itself and not on the development of spontaneous seizures while hyperactivity in the open-field test and light-dark chamber was more prominent in rats with spontaneous seizures. Administration of WIN-55,212-2 decreased emotional behavior in the elevated plus maze but did not affect hyperactive behavior in the open-field test. Spatial memory impairment developed both in the presence or absence of spontaneous seizures and was not affected by administration of WIN-55,212-2. Both administration of endocannabinoid receptor agonist WIN-55,212-2 and the presence of spontaneous seizures affected SE-induced neuronal loss in the hippocampus.

Topics: Animals; Anticonvulsants; Benzoxazines; Disease Models, Animal; Endocannabinoids; Hippocampus; Lithium Chloride; Locomotion; Male; Maze Learning; Morpholines; Naphthalenes; Pilocarpine; Rats; Rats, Wistar; Seizures; Status Epilepticus

Increasing evidence supports that the efflux transporters, especially P-glycoprotein (P-gp), have vital roles on drug resistance in epilepsy. Overexpression of P-gp in the brain could reduce the anti-epileptic drugs (AEDs) concentration in the epileptogenic zone, resulting in drug resistance. Studies have demonstrated that recurrent seizures induce the expression of P-gp and status epilepticus (SE) could upregulate the expression of P-gp, resulting in drug resistance. MicroRNAs (miRNAs), as endogenous regulators, represent small regulatory RNA molecules that have been shown to act as negative regulators of gene expression in different biological processes. We investigated the impact of miR-146a-5p on the expression of P-gp in status epilepticus rat model. The expression of miR-146a-5p in rat cortex and hippocampus was measured by quantitative RT-PCR at 2 weeks after induction of SE. Meanwhile, we detected the expression of P-gp in the brain of SE rats using Western blotting and immunohistochemistry. Upregulation of miR-146a-5p and overexpression of P-gp were evident at 2 weeks after SE. Moreover, the expression of P-gp was downregulated by injection of miR-146a mimic into the hippocampus. We also detected the expression of interleukin-1 receptor-associated protein kinases-1 (IRAK1) and tumor necrosis factor receptor-associated factor 6 (TRAF6) and nuclear factor-kappaB (NF-κB) p65 using Western blotting and immunohistochemistry, which indicated the expression of IRAK1, TRAF6 and NF-κB p-p65/p65 increased in the brain of SE rats, and overexpression of miR-146a-5p could downregulate the expression of IRAK1, TRAF6, NF-κB p-p65/p65 and P-gp. Our study indicated that miR-146a-5p may decrease the expression of P-gp in status epilepticus rats via NF-κB signaling pathway.

Topics: Animals; ATP Binding Cassette Transporter, Subfamily B, Member 1; Cerebral Cortex; Down-Regulation; Hippocampus; Interleukin-1 Receptor-Associated Kinases; Lithium Chloride; Male; MicroRNAs; Pilocarpine; Rats, Sprague-Dawley; Status Epilepticus; TNF Receptor-Associated Factor 6; Transcription Factor RelA

This study aimed to investigate whether 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU), a soluble epoxide hydrolase inhibitor with anti-inflammatory effects, could alleviate spontaneous recurrent seizures (SRS) and epilepsy-associated depressive behaviours in the lithium chloride (LiCl)-pilocarpine-induced post-status epilepticus (SE) rat model.. The rats were intraperitoneally (IP) injected with LiCl (127 mg/kg) and pilocarpine (40 mg/kg) to induce SE. A video surveillance system was used to monitor SRS in the post-SE model for 6 weeks (from the onset of the 2nd week to the end of the 7th week after SE induction). TPPU (0.1 mg/kg/d) was intragastrically given for 4 weeks from the 21st day after SE induction in the SRS + 0.1 TPPU group. The SRS + PEG 400 group was given the vehicle (40% polyethylene glycol 400) instead, and the control group was given LiCl and PEG 400 but not pilocarpine. The sucrose preference test (SPT) and forced swim test (FST) were conducted to evaluate the depression-like behaviours of rats. Immunofluorescent staining, enzyme-linked immunosorbent assay, and western blot analysis were performed to measure astrocytic and microglial gliosis, neuronal loss, and levels of soluble epoxide hydrolase (sEH), cytokines [tumour necrosis factor alpha (TNF-α), interleukin (IL)-1β, and IL-6], and cyclic adenosine monophosphate (cAMP)-response element binding protein (CREB).. The frequency of SRS was significantly decreased at 6 weeks and 7 weeks after SE induction in the 0.1TPP U group compared with the SRS + PEG 400 group. The immobility time (IMT) evaluated by FST was significantly decreased, whereas the climbing time (CMT) was increased, and the sucrose preference rate (SPR) evaluated by SPT was in an increasing trend. The levels of sEH, TNF-α, IL-1β, and IL-6 in the hippocampus (Hip) and prefrontal cortex (PFC) were all significantly increased in the SRS + PEG 400 group compared with the control group; neuronal loss, astrogliosis, and microglial activation were also observed. The astrocytic and microglial activation and levels of the pro-inflammatory cytokines in the Hip and PFC were significantly attenuated in the TPPU group compared with the SRS + PEG 400 group; moreover, neuronal loss and the decreased CREB expression were significantly alleviated as well.. TPPU treatment after SE attenuates SRS and epilepsy-associated depressive behaviours in the LiCl-pilocarpine induced post-SE rat model, and it also exerts anti-inflammatory effects in the brain. Our findings suggest a new therapeutic approach for epilepsy and its comorbidities, especially depression.

Topics: Animals; Astrocytes; Brain; Depression; Depressive Disorder; Disease Models, Animal; Epilepsy; Epoxide Hydrolases; Hippocampus; Lithium Chloride; Male; Microglia; Neurons; Phenylurea Compounds; Pilocarpine; Piperidines; Rats; Rats, Sprague-Dawley; Seizures; Status Epilepticus; Tumor Necrosis Factor-alpha

Ifenprodil as a specific antagonist of NMDA receptors containing a dominant NR2B subunit exhibits age-dependent anticonvulsant action. Possible changes of this action due to status epilepticus (SE) elicited at early stage of development were studied using cortical epileptic afterdischarges (ADs) as a model.. Lithium-pilocarpine SE was induced at postnatal day 12 and effects of ifenprodil were studied 3, 6, 9, and 13 days after SE in rat pups with implanted epidural electrodes. Controls (LiPAR) received saline instead of pilocarpine. ADs were elicited by low frequency stimulation of sensorimotor cortex. Intensity of stimulation current increased in 18 steps from 0.2 to 15 mA. Ifenprodil (20 mg/kg) was administered intraperitoneally (ip) after the stimulation with 3.5-mA current. Threshold for four different phenomena as well as duration of ADs were evaluated.. The threshold for the transition into the limbic type of ADs was higher in 15-day-old SE rats than in LiPAR controls. Opposite difference was found in 18-day-old animals, older rats did not exhibit any difference. Isolated significant changes in total duration of ADs were found after high stimulation intensities. These changes appeared in 18-day-old rats where ADs were shorter in SE than in control LiPAR rats.. Changes in ifenprodil action were found only in the first week after SE but not in the second week. Interpretation of the results is complicated by failure of significant differences between SE and LiPAR rats probably due to a high dose of paraldehyde.

Topics: Age Factors; Animals; Animals, Newborn; Anticonvulsants; Brain Waves; Cerebral Cortex; Disease Models, Animal; Electric Stimulation; Excitatory Amino Acid Antagonists; Lithium Chloride; Male; Pilocarpine; Piperidines; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Status Epilepticus

Up to this day, the roles of PEA15 expression and its phosphorylation in seizure-related events have not been still unclear. In the present study, we found that PEA15 was distinctly phosphorylated in reactive astrocytes and apoptotic astrocytes in the rat hippocampus following LiCl-pilocarpine-induced status epilepticus (SE, a prolonged seizure activity). PEA15-serine (S) 104 phosphorylation was up-regulated in reactive astrocytes following SE, although PEA15 expression and its S116 phosphorylation were unaltered. Bisindolylmaleimide (BIM), a protein kinase C (PKC) inhibitor, attenuated SE-induced reactive astrogliosis, but phorbol 12-myristate 13-acetate (PMA, a PKC activator) aggravated it. Unlike reactive astrocytes, PEA15-S116 phosphorylation was reduced in apoptotic astrocytes. However, PEA15 expression and its S104 phosphorylation were unchanged in apoptotic astrocyte. Neither BIM nor PMA affected SE-induced astroglial apoptosis. PEA15 expression and its phosphorylations were not relevant to SE-induced CA1 neuronal death. These findings indicate that PEA15-S104 and S116 phosphorylations may play a role in reactive astrogliosis and prevention of astroglial apoptosis, respectively. Therefore, we suggest that the selective manipulation of PEA15 phosphorylations may regulate apoptotic and/or proliferative signals in astrocytes.

Topics: Animals; Apoptosis; Apoptosis Regulatory Proteins; Astrocytes; Benzylamines; Fluorescent Antibody Technique; Gliosis; Hippocampus; Indoles; Lithium Chloride; Male; Maleimides; Phosphoproteins; Phosphorylation; Pilocarpine; Protein Kinase C; Rats; Rats, Sprague-Dawley; Signal Transduction; Status Epilepticus; Sulfonamides; Tetradecanoylphorbol Acetate

Several works have demonstrated that status epilepticus (SE) induced-neurodegeneration appears to involve an overactivation of N-methyl-d-aspartate receptors and treatment with high-affinity NMDAR antagonists is neuroprotective against this brain damage. However, these compounds display undesirable side effects for patients since they block physiological NMDA receptor dependent-activity. In this context, memantine (MN), a well tolerable low-affinity NMDAR channel blocker, will be a promising alternative, since it does not compromise the physiological role of NMDA receptors on synaptic transmission. The aim of the present study was to investigate if MN could attenuate seizure severity and neuronal cell death caused by SE induced early in life. Wistar rats (15 days old; n = 6-8 per group) received memantine (20 mg/kg i.p.) in six different treatments: 6 and 3 h before SE onset; concomitant with pilocarpine; 15min and 1h after SE onset; and four consecutive administrations (15 min, 6 h, 12 h, and 18 h) after pilocarpine injection. Neurodegeneration was quantified by fluoro-jade C staining. Treatment with memantine increase latency to SE onset only in groups treated 3 h before or concomitant with pilocarpine. In CA1 hippocampal subfield, memantine significantly reduced neurodegeneration at the following times: 3 h prior SE-onset, concomitant with pilocarpine, and 15 min after pilocarpine injection. For amygdala and thalamus, all post-SE onset treatments were able to decrease neurodegeneration. In conclusion, the present study showed that MN was neuroprotective against SE-induced neuronal death and this neuroprotection appears to be time- and region-dependent.

Topics: Animals; Brain; Excitatory Amino Acid Antagonists; Female; Lithium Chloride; Male; Memantine; Neurons; Neuroprotective Agents; Pilocarpine; Rats, Wistar; Status Epilepticus

Clinical factors contributing to benzodiazepine failure in treating status epilepticus (SE) include suboptimal dosing and seizure duration. As many benzodiazepine-refractory episodes of SE arise from acute etiologies, we sought to determine whether etiology impacts SE treatment.. The potency of diazepam to terminate SE induced by lithium-pilocarpine (LiPilo-SE) or kainic acid (KA-SE) in 3-week-old rats was studied by video-electroencephalography. Synaptic γ-aminobutyric acid type A receptor (GABAR)-mediated currents were recorded from dentate granule cells using voltage-clamp electrophysiology. Surface expression of γ2 subunit-containing GABARs and Kv4.2 potassium channels in hippocampal slices was determined using a biotinylation assay. Expression of phosphorylated forms of β2/3 and γ2 subunits was determined using phosphospecific antibodies and Western blotting.. Diazepam failed to terminate late SE in LiPilo-SE animals but was successful in terminating KA-SE of 1- and 3-hour duration. One hour after SE onset, GABAR-mediated synaptic inhibition and γ2 subunit-containing GABAR surface expression were reduced in LiPilo-SE animals. These were unchanged in KA-SE animals at 1 and 3 hours. Phosphorylation of γ2 subunit residue S327 was unchanged in both models, although GABAR β3 subunit S408/409 residues were dephosphorylated in the LiPilo-SE animals. Kv4.2 potassium channel surface expression was increased in LiPilo-SE animals but reduced in KA-SE animals.. SE-model-dependent differences support a novel hypothesis that the development of benzodiazepine pharmacoresistance may be etiologically predetermined. Further studies are required to investigate the mechanisms that underlie such etiological differences during SE and whether etiology-dependent protocols for the treatment of SE need to be developed. Ann Neurol 2018;83:830-841.

Topics: Animals; Benzodiazepines; Brain Waves; Convulsants; Disease Models, Animal; Electroencephalography; Gene Expression Regulation; Hippocampus; Kainic Acid; Lipoproteins; Lithium Chloride; Male; Phosphorylation; Pilocarpine; Protein Transport; Rats; Rats, Sprague-Dawley; Receptors, GABA-A; Shal Potassium Channels; Statistics, Nonparametric; Status Epilepticus; Time Factors

Synaptic vesicle protein 2A (SV2A) has become an attractive target of investigation because of its role in the pathophysiology of epilepsy; SV2A is expressed ubiquitously throughout the brain in all nerve terminals independently of their neurotransmitter content and plays an important but poorly defined role in neurotransmission. Previous studies have shown that modifications in the SV2A protein expression could be a direct consequence of disease severity. Furthermore, these SV2A modifications may depend on specific changes in the nerve tissue following the induction of epilepsy and might be present in both excitatory and inhibitory terminals. Thus, we evaluated SV2A protein expression throughout the hippocampi of lithium-pilocarpine rats after status epilepticus (SE) and during early and late epilepsy. In addition, we determined the γ-aminobutyric acid (GABA)ergic or glutamatergic nature associated with SV2A modifications. Wistar rats were treated with lithium-pilocarpine to induce SE and subsequently were shown to present spontaneous recurrent seizures (SRS). Later, we conducted an exhaustive semi-quantitative analysis of SV2A optical density (OD) throughout the hippocampus by immunohistochemistry. Levels of the SV2A protein were substantially increased in layers formed by principal neurons after SE, mainly because of GABAergic activity. No changes were observed in the early stage of epilepsy. In the late stage of epilepsy, there were minor changes in SV2A OD compared with the robust modifications of SE; however, SV2A protein expression generally showed an increment reaching significant differences in two dendritic layers and hilus, without clear modifications of GABAergic or glutamatergic systems. Our results suggest that the SV2A variations may depend on several factors, such as neuronal activity, and might appear in both excitatory and inhibitory systems depending on the epilepsy stage.

Topics: Animals; Disease Models, Animal; Gene Expression; Hippocampus; Lithium Chloride; Male; Membrane Glycoproteins; Nerve Tissue Proteins; Neurons; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Temporal lobe epilepsy is usually associated with cognitive decline and memory deficits. Despite numerous existing studies on various animal models, the mechanisms of these deficits remain largely unclear. A specific form of long-term synaptic efficacy changes-long-term depression (LTD)-is thought to play an important role in memory formation and learning. However, extremely little is known about the possible alteration of LTD induction and dynamics after a status epilepticus (SE). In this work, we investigated the acute and delayed effects of lithium-pilocarpine-induced SE on NMDAR-dependent and NMDAR-independent hippocampal LTD in vitro. We found that SE affected the NMDAR-dependent and NMDAR-independent forms of LTD in different manners. The NMDAR-dependent form of LTD was almost intact 3 days after SE, but it switched from a predominantly presynaptic to a more postsynaptic locus of expression. In contrast, the NMDAR-independent LTD in the hippocampal Schaffer collaterals-CA1 synapses was fully abolished 3 days after SE. Our results emphasize the role of non-NMDA-dependent synaptic plasticity changes in the processes of epileptogenesis and the potential for therapy development.

Topics: 2-Amino-5-phosphonovalerate; Animals; CA1 Region, Hippocampal; CA3 Region, Hippocampal; Disease Models, Animal; Epilepsy, Temporal Lobe; Excitatory Amino Acid Antagonists; Excitatory Postsynaptic Potentials; Lithium Chloride; Long-Term Synaptic Depression; Male; Pilocarpine; Random Allocation; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Synapses; Time Factors; Tissue Culture Techniques

Status epilepticus (SE), is characterized by high mortality and morbidity, which can cause neuronal injury, neuronal death and alteration of neuronal networks, Recently, inflammation was shown to play a significant role in SE pathogenesis. And miRNA-146a has been shown to be involved in inflammation and to inhibit inflammatory cytokines through NF-κB pathway. In our study, we investigated the relationship between inflammation and miR-146a expression.. The SE rat model was induced by lithium-pilocarpine. Hematoxylin and eosin staining (H&E) was performed to observe the histopathology of the rat hippocampus. The expression of COX-2, TNF-α, IL-6 and IL-1β were respectively measured by Western blot and Bio-Plex ProTM Assays. The miR-146a expression in hippocampus tissue was measured by Quantitative real-time PCR.. microRNA-146a was highly expressed in the hippocampus of SE rats coupled with increased level of inflammatory cytokines than the normal group. And TQ can attune the expression of inflammatory cytokines, meanwhile, miR-146a was lower in TQ group. The expression of miRNA-146a were positively correlated with the level of inflammatory reaction.. TQ may alleviate the inflammatory reaction by inhibiting the NF-κB signaling pathway. Our study shows that miRNA-146a was involved in the inflammatory response and indicated inflammation severity in SE rats. Therefore, miRNA-146a may serve as a potential biomarker or a therapeutic target in SE.

Topics: Animals; Anticonvulsants; Benzoquinones; Cytokines; Disease Models, Animal; Gene Expression Regulation; Hippocampus; Inflammation; Lithium Chloride; Male; MicroRNAs; Muscarinic Agonists; NF-kappa B; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

Status epilepticus (SE) causes irreversible neurodegeneration if not terminated quickly. Perampanel (PER), a potent AMPA receptor antagonist, has previously been shown to terminate seizures in the lithium-pilocarpine SE model. In the present study, we assessed whether PER would also prevent neuronal damage in this model.. SE was induced in rats using lithium chloride and pilocarpine. Initiation of SE was defined as continuous seizures that exhibited as rearing accompanied by bilateral forelimb clonus (Racine score 4). Either PER (0.6, 2, or 6mg/kg) or diazepam (DZP, 10mg/kg) was administered intravenously 30min after SE initiation. Histopathological samples from treated and seizure-naive rats were taken one week after treatment and then stained with an anti-neuronal nuclei (NeuN) antibody. The sections were analyzed by using a pixel-counting algorithm to quantify the amount of staining in the CA1 subregion of the hippocampus, piriform cortex (Pir), and mediodorsal thalamic nucleus (MD).. DZP administration did not suppress seizures or the degeneration of neurons in the examined areas. Seizures were terminated in 100% of rats treated with 6mg/kg PER (n=8) and in 47% (7/15) of rats treated with 2mg/kg PER, and neurons in the analyzed areas of these animals were preserved to the level seen in naive rats. In the eight animals in which 2mg/kg PER did not terminate the seizures, neuronal loss was partially attenuated in CA1 and Pir, and neurons were fully preserved in MD. Treatment with 0.6mg/kg PER did not terminate the seizures or significantly preserve neurons. The anti-seizure effect of PER correlated well with the degree of neuroprotection in each analyzed area.. PER exhibited a strong neuroprotective effect in a drug-refractory SE model, and this effect was correlated with its attenuation of seizure.

Topics: Animals; Anticonvulsants; Antigens, Nuclear; Brain; Cell Death; Diazepam; Dose-Response Relationship, Drug; Immunohistochemistry; Lithium Chloride; Male; Nerve Tissue Proteins; Neurodegenerative Diseases; Neurons; Neuroprotective Agents; Nitriles; Pilocarpine; Pyridones; Rats, Sprague-Dawley; Status Epilepticus

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 (25 mg/kg/day) pretreated group, vit-D (1.5 mcg/kg/day) pretreated group, and group pretreated with vit-D and lamotrigine for 2 weeks. At the end of treatment, SE was induced by single intraperitoneal injection of LiCl (127 mg/kg), followed 24 h later by pilocarpine (30 mg/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.

Topics: Animals; Anticonvulsants; Antioxidants; Disease Models, Animal; Drug Therapy, Combination; Lamotrigine; Lithium Chloride; Male; Neuroprotective Agents; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Treatment Outcome; Triazines

Acute seizures induced dendritic formation and synaptogenesis promotes aberrant circuitry development and further aggravates underlying conditions towards chronic epilepsy. The G protein-coupled receptor kinase-5 (GRK5) served as a key modulator in neurogenesis and the establishment of functional neuronal circuitry. This included dendritic development, as its dysfunction could cause different central nervous system disorders, including Alzheimer's disease. However, the involvement of GRK5 in the progression of epilepsy remains unclear. The purpose of this study is to investigate the involvement of GRK5 in epilepsy, as well as its potential correlation with dendritic formation after status epilepticus.. 120 rats were divided into control and model groups. The rats in the model group were injected intraperitoneally with lithium chloride-pilocarpine hydrochloride to establish the rat model of status epilepticus (SE). The brain and hippocampus were collected at 1, 3, 7, 14 and 28days post SE induction. The expression and distribution of GRK5 and the dendritic marker microtubule-associated protein-2 (MAP-2) were detected in the hippocampus via western blot or immunohistochemistry. The co-localization of GRK5 with MAP-2 was examined via laser confocal double immunofluorescence staining. The interactions between GRK5 and MAP-2 during epileptogenesis were evaluated via immunoprecipitation.. GRK5 was distributed in all areas of the hippocampus. Its expression was significantly up-regulated in the hippocampal CA1, DG, and H areas at 7d and 14d after SE. After 14d it began to reduce. and then reduced. MAP-2 primarily existed in the neuronal dendrites of the hippocampal subregion. Its expression was enhanced at 3d. It reached its maximum level at 14d after SE, where it then began to fall. The confocal microscope analysis revealed that GRK5 was co-located well within MAP-2 positive cells. The interaction between GRK5 and MAP-2 became enhanced at 7d and 14d after SE.. GRK5 was involved in the development of epilepsy. It was associated with dendritic formation in epilepsy. This study provides a new perspective for elucidating the epilepsy pathogenesis. The concrete mechanisms of the GRK5 within epileptogenesis require further research.

Topics: Analysis of Variance; Animals; Disease Models, Animal; G-Protein-Coupled Receptor Kinase 5; Hippocampus; Immunoprecipitation; Lithium Chloride; Male; Microscopy, Confocal; Microtubule-Associated Proteins; Muscarinic Agonists; Muscarinic Antagonists; Pilocarpine; Rats; Rats, Sprague-Dawley; Scopolamine; Status Epilepticus; Time Factors

Status epilepticus (SE) in rats, along with chronic epilepsy, leads to the development of behavioral impairments resembling depressive disorder and/or attention deficit/hyperactivity disorder (ADHD), thus reflecting respective comorbidities in epilepsy patients. Suppressed neurotransmitter tone in the raphe nucleus (RN)-prefrontal cortex (PFC) serotonergic pathway and in the locus coeruleus (LC)-PFC noradrenergic pathway underlies depressive- and impulsive-like behavioral deficits respectively. We examined possible mechanisms leading to the monoamine dysfunction in brainstem efferents, namely modulatory effects of the neuropeptide galanin on serotonin (5-HT) and norepinephrine (NE) signaling. SE was induced in young adult male Wistar rats by LiCl and pilocarpine. Epileptic rats were categorized vis-à-vis behavioral deficits as not impaired, "depressed" and "impulsive". Depressive- and impulsive-like behaviors were examined in the forced swimming test (FST). The strength of serotonergic transmission in RN-PFC and of noradrenergic transmission in LC-PFC was analyzed using in vivo fast scan cyclic voltammetry. Galanin receptor type 1 (GalR1)/type 2 (GalR2) antagonist M40, and a preferential GalR2 antagonist M871 were administered over 3days locally into either RN or LC by means of ALZET osmotic minipumps connected to locally implanted infusion cannulas. Intra-RN injection of M40 improved serotonergic tone and depressive-like behavior in epileptic "depressed" rats. Intra-LC injection of M40 improved noradrenergic tone and impulsive-like behavior in epileptic "impulsive" rats. The effects of M40 were only observed in impaired subjects. The treatment did not modify neurotransmission and behavior in naïve and epileptic not impaired rats; in "depressed" rats the effects were limited to serotonergic transmission and immobility, while in "impulsive" rats - to noradrenergic transmission and struggling behavior. Intra-RN administration of M871 exacerbated depressive-like behavior, but had no effects on any other of the examined parameters in any category of animals. These findings suggest that endogenous galanin, acting through GalR1 may be involved in the pathophysiology of epilepsy-associated depression and ADHD via inhibiting RN-PFC serotonergic and LC-PFC noradrenergic transmissions respectively.

Topics: Animals; Antidepressive Agents; Biogenic Monoamines; Depression; Disease Models, Animal; Drug Delivery Systems; Galactolipids; Galanin; Impulsive Behavior; Lithium Chloride; Locus Coeruleus; Male; Muscarinic Agonists; Peptide Fragments; Pilocarpine; Raphe Nuclei; Rats; Rats, Wistar; Serotonin; Status Epilepticus; Swimming

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.

Topics: Animals; Anticonvulsants; Cerebral Cortex; Convulsants; Disease Models, Animal; Lithium Chloride; Male; Paraldehyde; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Acetylcholine (ACh) and ATP are rapidly acting neurotransmitters with a putative role in epileptic seizures. In the present study we investigated extracellular concentrations of both neurotransmitters in parallel by microdialysis in rat hippocampus. We found that infusion of neostigmine increases, while calcium-free perfusion and infusion of tetrodotoxin (TTX) decreases, ACh levels. Calcium-free perfusion also decreased ATP levels which were, however, not affected by neostigmine or TTX. During status epilepticus, ACh levels were increased threefold but returned to baseline after the termination of seizures by diazepam. ATP levels were unchanged during status epilepticus but a several-fold increase was seen when AOPCP, an inhibitor of 5'-endonucleotidase, was infused. The results demonstrate an increase of ATP levels during epileptic seizures which, however, was not of neuronal origin.

Topics: Acetylcholine; Adenosine Triphosphate; Animals; Anticonvulsants; Diazepam; Extracellular Space; Hippocampus; Lithium Chloride; Male; Neurons; Pilocarpine; Rats, Sprague-Dawley; Status Epilepticus

The goal of this study was to determine whether there are region-specific or time-dependent changes in GABA-mediated synaptic inhibition of principal neurons in the hippocampus during in vivo status epilepticus. Standard whole cell patch clamp electrophysiological techniques were used to characterize miniature inhibitory postsynaptic currents (mIPSCs) in recordings from the principal neurons (PNs) of the dentate gyrus, CA1, and CA3 in acutely-obtained hippocampal slices from control and lithium/pilocarpine-induced status epilepticus(SE)-treated animals. The reduction in mIPSC amplitude was pervasive across the 3 regions examined in hippocampal slices obtained after 60 min (late) or just 15 min after the onset of SE. The mIPSC frequency was reduced in all 3 regions after 60 min and 2 regions (dentate, CA1) after 15 min. These findings lend further support to the hypothesis that a rapid modification of the postsynaptic GABAA receptor population leads to a widespread decline in GABA-mediated inhibition that, in part, contributes to both the self-sustaining nature of SE and to the decrease in the efficacy of benzodiazepines.

Topics: Animals; Disease Models, Animal; gamma-Aminobutyric Acid; Hippocampus; Inhibitory Postsynaptic Potentials; Lithium Chloride; Male; Miniature Postsynaptic Potentials; Neurons; Patch-Clamp Techniques; Pilocarpine; Rats, Sprague-Dawley; Status Epilepticus; Time; Tissue Culture Techniques

MicroRNAs (miRNAs) are noncoding small RNAs that control gene expression at the posttranscriptional level. Some dysregulated miRNAs have been shown to play important roles in epileptogenesis. The aim of this study was to determine if miR-199a-5p regulates seizures and seizure damage by targeting the antiapoptotic protein silent information regulator 1 (SIRT1).. Hippocampal expression levels of miR-199a-5p, SIRT1, and acetylated p53 were quantified by quantitative real-time polymerase chain reaction (RT-PCR) and Western blotting in the acute, latent, and chronic stages of epilepsy in a rat lithium-pilocarpine epilepsy model. Silencing of miR-199a-5p expression in vivo was achieved by intracerebroventricular injection of antagomirs. The effects of targeting miR-199a-5p and SIRT1 protein on seizure and epileptic damage post-status epilepticus were assessed by electroencephalography (EEG) and immunohistochemistry, respectively.. miR-199a-5p expression was up-regulated, SIRT1 levels were decreased, and neuron loss and apoptosis were induced in epilepsy model rats compared with normal controls, as determined by up-regulation of acetylated p53 and cleaved caspase-3 expression. In vivo knockdown of miR-199a-5p by an antagomir alleviated the seizure-like EEG findings and protected against neuron damage, in accordance with up-regulation of SIRT1 and subsequent deacetylation of p53. Furthermore, the seizure-suppressing effect of the antagomir was partly SIRT1 dependent.. The results of this study suggest that silencing of miR-199a-5p exerts a seizure-suppressing effect in rats, and that SIRT1 is a direct target of miR-199a-5p in the hippocampus. The effect of miR-199a-5p on seizures and seizure damage is mediated via down-regulation of SIRT1. The miR-199a-5p/SIRT1 pathway may thus represent a potential target for the prevention and treatment of epilepsy and epileptic damage.

Topics: Animals; Anticonvulsants; Apoptosis; Argonaute Proteins; Carbazoles; Convulsants; Disease Models, Animal; Hippocampus; Lithium Chloride; Male; MicroRNAs; Neurons; Oligonucleotides, Antisense; Pilocarpine; Rats; Rats, Sprague-Dawley; Signal Transduction; Sirtuin 1; Status Epilepticus; Tumor Suppressor Protein p53; Up-Regulation

AMPA receptors (AMPARs) are responsible for fast excitatory neurotransmission, and their prolonged activation can result in the generation and spread of epileptic seizures. At early stages of postnatal development, the majority of AMPARs are permeable to both Na(+) and Ca(2+) ions. This permeability, which increases neuronal excitability, is due to the lack of the GluA2 subunit, encoded by the GRIA2A gene, and/or the presence of an unedited GluA2 subunit Q/R site (glutamine instead of arginine). Lithium chloride- and pilocarpine-induced status epilepticus (LiCl/Pilo-SE) in rodents represents a model of severe seizures that result in development of temporal lobe epilepsy (TLE). The aim of this study was to determine how LiCl/Pilo-SE induced early in life (at postnatal day 12; P12) alters normal expression of the GRIA2A gene and GluA2 protein. SE was interrupted by an injection of paraldehyde (Para). Control groups were 1) naïve animals, and 2) siblings of SE rats receiving only LiCl and paraldehyde (LiCl/Para). The expression profile of GRIA2A mRNA was determined via qPCR, and GluA2 protein levels were measured by western blotting. The analysis was performed at 3h (protein levels), and then 3-, 6-, 13-, and 60days, following LiCl/Pilo-SE or LiCl/Para injection (i.e. at P12, P15, P18, P25, P72 respectively). Six different brain regions were analyzed: frontal (CXFR), parietal (CXPAR), and occipital (CXOC) cortex, dorsal (HD) and ventral (HV) hippocampus, and thalamus (TH). There was a significant increase in GRIA2A mRNA expression in the CXFR, CXPAR, and CXOC of P18 SE animals. In CXFR and HD, increased expression of GluA2 AMPAR subunit protein was detected, as well as a surge in GRIA2A mRNA and GluA2 protein expression especially at P18. In HD the surge was detected not only during development (P18), but also later in life (P72). Since high levels of GluA2 can be neuroprotective (by decreasing Ca(2+) permeability), our data suggest that the neocortex and dorsal hippocampus are able to activate endogenous antiepileptic mechanisms. A marked decrease in the overall expression of GluA2 protein in the HV in the LiCl/Pilo-SE and LiCl/Para rats, suggests that the HV is predisposed to excitotoxicity, not only during development, but even in adulthood. Interestingly, LiCl in combination with paraldehyde can also strongly alter the normal ontogeny of GRIA2A mRNA as well as GluA2 subunit protein expression.

Topics: Age Factors; Animals; Animals, Newborn; Brain; Convulsants; Disease Models, Animal; Fluoresceins; Gene Expression Regulation, Developmental; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Wistar; Receptors, AMPA; RNA, Messenger; Status Epilepticus

Pharmacoresistance remains an unsolved therapeutic challenge in status epilepticus (SE) and in cholinergic SE induced by nerve agent intoxication. SE triggers a rapid internalization of synaptic γ-aminobutyric acid A (GABAA ) receptors and externalization of N-methyl-d-aspartate (NMDA) receptors that may explain the loss of potency of standard antiepileptic drugs (AEDs). We hypothesized that a drug combination aimed at correcting the consequences of receptor trafficking would reduce SE severity and its long-term consequences.. A severe model of SE was induced in adult Sprague-Dawley rats with a high dose of lithium and pilocarpine. The GABAA receptor agonist midazolam, the NMDA receptor antagonist ketamine, and/or the AED valproate were injected 40 min after SE onset in combination or as monotherapy. Measures of SE severity were the primary outcome. Secondary outcomes were acute neuronal injury, spontaneous recurrent seizures (SRS), and Morris water maze (MWM) deficits.. Midazolam-ketamine dual therapy was more efficient than double-dose midazolam or ketamine monotherapy or than valproate-midazolam or valproate-ketamine dual therapy in reducing several parameters of SE severity, suggesting a synergistic mechanism. In addition, midazolam-ketamine dual therapy reduced SE-induced acute neuronal injury, epileptogenesis, and MWM deficits.. This study showed that a treatment aimed at correcting maladaptive GABAA receptor and NMDA receptor trafficking can stop SE and reduce its long-term consequences. Early midazolam-ketamine dual therapy may be superior to monotherapy in the treatment of benzodiazepine-refractory SE.

Topics: Animals; Anticonvulsants; Brain; Cholinergic Agents; Disease Models, Animal; Drug Synergism; Drug Therapy, Combination; Ketamine; Learning Disabilities; Lithium Chloride; Male; Maze Learning; Midazolam; N-Methylscopolamine; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Valproic Acid

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.

Topics: Animals; Anti-Inflammatory Agents; Benzoquinones; Cerebral Cortex; Cytokines; Electroencephalography; Hippocampus; Lithium Chloride; Male; Maze Learning; Memory; NF-kappa B; Pilocarpine; Rats, Sprague-Dawley; Seizures; Signal Transduction; Status Epilepticus

The pilocarpine rat model, in which status epilepticus (SE) leads to epilepsy with spontaneous recurrent seizures (SRS), is widely used to study the mechanisms of epileptogenesis and develop strategies for epilepsy prevention. SE is commonly interrupted after 30-90min by high-dose diazepam or other anticonvulsants to reduce mortality. It is widely believed that SE duration of 30-60min is sufficient to induce hippocampal damage and epilepsy. However, resistance to diazepam develops during SE, so that an SE that is longer than 30min is difficult to terminate, and SE typically recurs several hours after diazepam, thus forming a bias for studies on epileptogenesis or antiepileptogenesis. We developed a drug cocktail, consisting of diazepam, phenobarbital, and scopolamine that allows complete and persistent SE termination in the lithium-pilocarpine model. A number of novel findings were obtained with this cocktail. (a) In contrast to previous reports with incomplete SE suppression, a SE of 60min duration did not induce epilepsy, whereas epilepsy with SRS developed after 90 or 120min SE; (b) by comparing groups of rats with 60 and 90min of SE, development of epilepsy could be predicted by behavioral hyperexcitability and decrease in seizure threshold, indicating that these read-outs are suited as biomarkers of epileptogenesis; (c) CA1 damage was prevented by the cocktail, but rats exhibited cell loss in the dentate hilus, which was related to development of epilepsy. These data demonstrate that the duration of SE needed for induction of epileptogenesis in this model is longer than previously thought.

Topics: Animals; Anticonvulsants; Diazepam; Disease Models, Animal; Electrodes, Implanted; Electroencephalography; Female; Hippocampus; Lithium Chloride; Neurons; Phenobarbital; Pilocarpine; Polypharmacy; Rats, Sprague-Dawley; Scopolamine; Seizures; Status Epilepticus; Time Factors; Treatment Outcome

This study tightly controlled seizure duration and severity during status epilepticus (SE) in postnatal day 10 (P10) rats, in order to isolate hyperthermia as the main variable and to study its consequences. Body temperature was maintained at 39 ± 1 °C in hyperthermic SE rats (HT+SE) or at 35 ± 1 °C in normothermic SE animals (NT+SE) during 30 min of SE, which was induced by lithium-pilocarpine (3 mEq/kg, 60 mg/kg) and terminated by diazepam and cooling to NT. All video/EEG measures of SE severity were similar between HT+SE and NT+SE pups. At 24h, neuronal injury was present in the amygdala in the HT+SE group only, and was far more severe in the hippocampus in HT+SE than NT+SE pups. Separate groups of animals were monitored four months later for spontaneous recurrent seizures (SRS). Only HT+SE animals developed convulsive SRS. Both HT+SE and NT+SE animals developed electrographic SRS (83% vs. 55%), but SRS frequency and severity were higher in hyperthermic animals (12.5 ± 3.5 vs. 4.2 ± 2.0 SRS/day). The density of hilar neurons was lower, thickness of the amygdala and perirhinal cortex was reduced, and lateral ventricles were enlarged in HT+SE over NT+SE littermates and HT/NT controls. In this model, hyperthermia greatly increased the epileptogenicity of SE and its neuropathological sequelae.

Topics: Adjuvants, Immunologic; Animals; Animals, Newborn; Anticonvulsants; Apoptosis; Brain; Cell Death; Diazepam; Disease Models, Animal; Hyperthermia, Induced; Lithium Chloride; Male; Muscarinic Agonists; Nerve Degeneration; Neurons; Neuropil; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Time Factors

In the present study, we investigated whether endoplasmic reticulum (ER) stress is associated with neuronal- and astroglial-death in the hippocampus using LiCl-pilocarpine-induced status epilepticus (SE) rat model. Glucose-related protein (GRP) 78 and protein disulfide isomerase (PDI) expressions were transiently increased in CA1 neurons and dentate granule cells, and subsequently decreased in these cells following SE. GRP94 and calnexin (CNX) expression was gradually reduced in CA1 neurons, not in dentate granule cells. Phospho-protein kinase RNA (PKR)-like ER kinase (pPERK), phospho-eukaryotic initiation factor 2α (peIF2A) and CCAAT/enhancer-binding protein homologous protein (CHOP) immunoreactivities were observed in 17%, 12% and 7% of degenerating CA1 neurons, respectively. GRP 78 and PDI expressions were also up-regulated in reactive astrocytes within the CA1-3 regions. In the molecular layer of the dentate gyrus, PDI-positive astrocytes showed TUNEL signal, nuclear apoptosis inducing factor translocation and pPERK/peIF2A/CHOP immunoreactivities. Four weeks after SE, clasmatodendritic astrocytes showed pPERK peIF2A and CNX immunoreactivities without CHOP expression. These findings indicate that SE-induced ER stress may be associated with astroglial apoptosis and autophagic astroglial death in the regional-specific pattern.

Topics: Animals; Apoptosis Inducing Factor; Astrocytes; Calnexin; Disease Models, Animal; Endoplasmic Reticulum Chaperone BiP; Endoplasmic Reticulum Stress; Hippocampus; HSP70 Heat-Shock Proteins; In Situ Nick-End Labeling; Lithium Chloride; Male; Membrane Proteins; Nerve Tissue Proteins; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Time Factors; Transcription Factor CHOP; Up-Regulation

Status epilepticus (SE) is a life-threatening neurological emergency associated with a high mortality rate. The serotonin 1A (5-HT1A) receptor is a possible target for the treatment of SE, but its role in animal models and the precise area of brain involved remain controversial. The hippocampus is a candidate site due to its key role in the development of SE and the existence of a high density of 5-HT1A receptors. Therefore, we investigated the effects of subcutaneous and intrahippocampal activation of 5-HT1A receptors in lithium-pilocarpine-induced SE, and tested whether the hippocampus is a true effector site. We developed SE in male Sprague-Dawley rats by giving lithium chloride (LiCl; 3 meq/kg, i.p.) 22-24 h prior to pilocarpine (25 mg/kg, i.p.), and found that 8-OH-DPAT, a 5-HT1A receptor agonist administered subcutaneously (s.c.) at 0.5 or 1.0 mg/kg 1 h before pilocarpine injection increased the latency to the first epileptiform spikes, the electrographic SE, and the behavioral generalized seizures (GS), while reducing the total EEG seizure time (P <0.01). The duration of GS was shortened only by 1.0 mg/kg 8-OH-DPAT s.c. (P <0.05). All these effects were inhibited by combined administration of WAY-100635 (1.0 mg/kg, s.c.) (P <0.05), an antagonist of the 5-HT1A receptor, but WAY-100635 alone and low doses of 8-OHDPAT (0.01 and 0.1 mg/kg) did not alter seizure activity. Furthermore, intrahippocampal 8-OH-DPAT only shortened the GS duration (P <0.05). These findings imply that the 5-HT1A receptor is a promising therapeutic target against the generation and propagation of SE, and hippocampal receptors are involved in reducing the seizure severity.

Topics: 8-Hydroxy-2-(di-n-propylamino)tetralin; Action Potentials; Analysis of Variance; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Electroencephalography; Hippocampus; Lithium Chloride; Male; Muscarinic Agonists; Pilocarpine; Piperazines; Pyridines; Rats; Rats, Sprague-Dawley; Reaction Time; Receptor, Serotonin, 5-HT1A; Serotonin Antagonists; Status Epilepticus; Time Factors

Recent studies have shown that transforming growth factor β (TGFβ) signaling participates in the epileptogenesis. Serine-threonine kinase receptor-associated protein (STRAP) and Smad7 synergize in the inhibition of the TGFβ signaling. The aim of the present study was to determine the expression pattern of STRAP and Smad7 in the hippocampus and temporal lobe cortex of pilocarpine-induced rats models of epilepsy.. Lithium chloride-pilocarpine-induced rats with status epilepticus (SE) were established. Total of 60 male Sprague-Dawley rats was used as control (n = 10), 24 h (n = 10), 72 h (n = 10), 1 week (n = 10), 1 month (n = 10), and 2 months (n = 10) after pilocarpine-induced SE, respectively. We detected the expression levels of STRAP and Smad7 in the hippocampus and temporal lobe cortex of rats at the aforementioned time points using western blotting and immunohistochemistry.. STRAP level was significantly decreased in 24 h, 72 h (acute stage), 1 week (latent stage), 1 month, 2 months (chronic stage), respectively, in the rat models compared with the control rats by using both western blotting and immunohistochemistry. Smad7 had similar reduced pattern as STRAP.. Our results indicate that STRAP and Smad7 proteins might be involved in the development of temporal lobe epilepsy.

Topics: Adaptor Proteins, Signal Transducing; Animals; Blotting, Western; Chronic Disease; Disease Models, Animal; Epilepsy, Temporal Lobe; Gene Expression; Hippocampus; Immunohistochemistry; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Sprague-Dawley; Smad7 Protein; Status Epilepticus; Temporal Lobe; Time Factors

Recently, the role of inflammation has attracted great attention in the pathogenesis of mesial temporal lobe epilepsy (MTLE), and microRNAs start to emerge as promising new players in MTLE pathogenesis. In this study, we investigated the dynamic expression patterns of tumor necrosis factor alpha (TNF-α) and microRNA-155 (miR-155) in the hippocampi of an immature rat model of status epilepticus (SE) and children with MTLE. The expressions of TNF-α and miR-155 were significantly upregulated in the seizure-related acute and chronic stages of MTLE in the immature rat model and also in children with MTLE. Modulation of TNF-α expression, either by stimulation using myeloid-related protein (MRP8) or lipopolysaccharide or inhibition using lenalidomide on astrocytes, leads to similar dynamic changes in miR-155 expression. Our study is the first to focus on the dynamic expression pattern of miR-155 in the immature rat of SE lithium-pilocarpine model and children with MTLE and to detect their relationship at the astrocyte level. TNF-α and miR-155, having similar expression patterns in the three stages of MTLE development, and their relationship at the astrocyte level may suggest a direct interactive relationship during MTLE development. Therefore, modulation of the TNF-α/miR-155 axis may be a novel therapeutic target for the treatment of MTLE.

Topics: Adolescent; Animals; Astrocytes; Case-Control Studies; Child; Epilepsy, Temporal Lobe; Female; Hippocampus; Humans; Lenalidomide; Lipopolysaccharides; Lithium Chloride; Male; MicroRNAs; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Thalidomide; Tumor Necrosis Factor-alpha

Experiments were conducted to evaluate the effects of transcranial focal electrical stimulation (TFS) applied via tripolar concentric ring electrodes, alone and associated with a sub-effective dose of diazepam (DZP) on the expression of status epilepticus (SE) induced by lithium-pilocarpine (LP) and subsequent neuronal damage in the hippocampus. Immediately before pilocarpine injection, male Wistar rats received TFS (300Hz, 200-μs biphasic square charge-balanced 50-mA constant current pulses for 2min) alone or combined with a sub-effective dose of DZP (0.41mg/kg, i.p.). In contrast with DZP or TFS alone, DZP plus TFS reduced the incidence of, and enhanced the latency to, mild and severe generalized seizures and SE induced by LP. These effects were associated with a significant reduction in the number of degenerated neurons in the hippocampus. The present study supports the notion that TFS combined with sub-effective doses of DZP may represent a therapeutic tool to induce anticonvulsant effects and reduce the SE-induced neuronal damage.

Topics: Analysis of Variance; Animals; Anticonvulsants; Cell Count; Deep Brain Stimulation; Diazepam; Disease Models, Animal; Fluoresceins; Hippocampus; Lithium Chloride; Male; Muscarinic Agonists; Neurons; Pilocarpine; Rats; Rats, Wistar; Reaction Time; Status Epilepticus

The present study examined whether status epilepticus (SE) induced by LiCl-pilocarpine in immature rats (postnatal day [P]12) interferes with normal development; leads to progressive epileptogenesis, or cognitive decline and to pathology similar to that seen in human temporal lobe epilepsy. We correlated the extent of pathologic changes with the severity of functional alterations or epilepsy. SE-induced changes were compared with those of rats with SE induced at P25. Animals of both ages were exposed to a battery of behavioral tests for up to 3months after SE. Rats with SE at P12 showed mild retardation of psychomotor development and delayed habituation, whereas rats with SE at P25 showed no habituation. Assessment in adulthood using the Morris water maze test revealed that SE at both P12 and P25 led to cognitive impairment and that the severity of the impairment increased with age. A handling test revealed increased aggression in rats with SE at P25, but not in rats with SE at P12. Epilepsy was diagnosed with continuous video-electroencephalographic (EEG) monitoring for up to 7d. P25 rats were monitored at 5months after SE and seizures were detected in 83.3% of animals. P12 animals were divided into two groups and monitored at 5 or 7months after SE. Both the severity and incidence of spontaneous recurrent seizures tended to progress with time, and their incidence increased from 50% to 87.5% at 5 and 7months, respectively. Morphometric analysis and stereologic assessment of hilar neurons performed after video-EEG monitoring revealed atrophy of temporal brain structures, enlargement of lateral ventricles, and loss of hilar neurons in both age groups. In P12 rats, morphologic damage also tended to progress over time. Performance of animals in the Morris water maze correlated with the severity of damage, but not with seizure parameters.

Topics: Aggression; Aging; Animals; Animals, Newborn; Antimanic Agents; Atrophy; Behavior, Animal; Brain; Cerebral Ventricles; Cognition Disorders; Disease Progression; Electroencephalography; Hippocampus; Lithium Chloride; Male; Maze Learning; Motor Activity; Muscarinic Agonists; Pilocarpine; Psychomotor Performance; Rats; Rats, Wistar; Status Epilepticus

Epilepsy is commonly associated with cognitive impairment. Astrocyte activation and oxidative stress occur following seizures, and play a role in the pathological injury of epilepsy with cognitive impairment. The peroxisome proliferator-activated receptor gamma (PPARγ) has been shown to exhibit neuroprotective and antioxidative effects in CNS diseases. Thus, we hypothesized that rosiglitazone, a PPARγ agonist, would prevent cognitive impairment by inhibiting astrocyte activation and regulating glutathione (GSH) homeostasis after status epilepticus (SE). Using a lithium pilocarpine-induced SE model, we found that rosiglitazone significantly prevented cognitive impairment induced by SE, and potently inhibited astrocyte activation with maintenance of GSH homeostasis in the hippocampus after SE. These protective effects were significantly reversed by co-treatment with the PPARγ antagonist T0070907. These data suggest that rosiglitazone can improve cognitive impairment, and inhibit astrocyte activation and oxidative damage following SE. Rosiglitazone may be a promising agent for treatment of epilepsy involving SE-induced cognitive impairment.

Topics: Analysis of Variance; Animals; Astrocytes; Benzamides; Cell Count; Cognition Disorders; Disease Models, Animal; Glial Fibrillary Acidic Protein; Glutathione; Glutathione Disulfide; Hippocampus; Lithium Chloride; Male; Maze Learning; Oxidative Stress; Pilocarpine; PPAR gamma; Pyridines; Rats; Rats, Sprague-Dawley; Rosiglitazone; Status Epilepticus; Systole; Thiazolidinediones

The aim of this study was to investigate whether NMDA receptor was involved in the upregulation of multidrug resistance protein 2 (Mrp2) expression during status epilepticus (SE).. The alterations in the expression of Mrp2 at various time points after SE, and the inhibition of glutamate N-methyl-D-aspartate (NMDA) receptor on Mrp2 expression in hippocampus were both tested by quantitative real-time polymerase chain reaction and western blot. Moreover, immunofluorescence was also used to analyze the impact of the NMDA receptor antagonist, MK-801, on the distribution of Mrp2 in different brain areas.. The results showed that gene encoding Mrp2 was upregulated in hippocampus at 6 hours after the end of SE, and this initial increase was followed by gradual normalization. While between 3 and 72 hours after the end of SE, the protein level of Mrp2 was upregulated in hippocampus, with the highest level emerging at 24 hours. The increment of Mrp2 gene and protein induced by SE was prevented by MK-801 at 6 and 24 hours respectively after the end of SE in the hippocampus. Moreover, immunofluorescence showed that seizures-induced increase of Mrp2 expression was attenuated by the administration of MK-801 mainly in capillaries. Rats after SE exhibited a significant upregulation of Mrp2 in the capillary endothelial cells of the cerebral cortex, piriform cortex, and hippocampus, compared with those in control at 24 hours after the end of SE.. The results indicated that the NMDA receptor plays an important role in the upregulation of Mrp2 expression in the blood-brain barrier.

Topics: Analysis of Variance; Animals; ATP Binding Cassette Transporter, Subfamily B; ATP-Binding Cassette Sub-Family B Member 4; Brain Waves; Disease Models, Animal; Dizocilpine Maleate; Electric Stimulation; Electroencephalography; Female; Gene Expression Regulation; Hippocampus; Lithium Chloride; Muscarinic Agonists; Neuroprotective Agents; Pilocarpine; Rats; Rats, Wistar; RNA, Messenger; Status Epilepticus; Time Factors; von Willebrand Factor

A Ketogenic Diet (KD) mimics the anticonvulsant effects of fasting, which are known to suppress seizures. The purinergic system has been investigated in the matter of epilepsy development, especially the nucleoside adenosine, which has been considered a natural brain anticonvulsant. During epileptic seizures, extracellular adenosine concentration rises rapidly to micromolar levels. Adenosine can exert its anticonvulsant functions, after its release by nucleoside bidirectional transport, or by production through the sequential catabolism of ATP by ectonucleotidases, such as E-NTPDases (ectonucleoside triphosphate diphosphohydrolases) and ecto-5'-nucleotidase. Here, we have investigated the effect of a ketogenic diet on the nucleotide hydrolysis and NTPDases expression in the lithium-pilocarpine (Li-Pilo) model of epilepsy. For the induction of Status Epileticus (SE), 21-day-old female Wistar rats received an i.p. injection of lithium chloride (127 mg/kg) and 18-19 h later an i.p. injection of pilocarpine hydrochloride (60 mg/kg). The control groups received an injection of saline. After induction of SE, the control and Li-Pilo groups received standard or ketogenic diets for 6 weeks. The lithium-pilocarpine exposure affected the ATP (a decrease of between 8 % and 16 %) and ADP (an increase of between 18 % and 22 %) hydrolysis in both groups whereas the diet did not impact the nucleotide hydrolysis. NTPDase2 and 3 mRNA expressions decreased in the Li-Pilo group (41 % and 42 %). This data highlights the participation of the purinergic system in the pathophysiology of this model of epilepsy, since nucleotide hydrolysis and NTPDase expressions were altered by Li-Pilo exposure, with no significant effects of the ketogenic diet. However, the interaction between purinergic signaling and a ketogenic diet on epilepsy still needs to be better elucidated.

Topics: 5'-Nucleotidase; Adenosine Diphosphate; Adenosine Monophosphate; Adenosine Triphosphate; Animals; Antimanic Agents; Diet, Ketogenic; Female; Hydrolysis; Ketones; Lithium Chloride; Muscarinic Agonists; Nucleotides; Pilocarpine; Rats; Rats, Wistar; Real-Time Polymerase Chain Reaction; Status Epilepticus; Subcellular Fractions

Transcranial direct current stimulation (tDCS) is a recently available, noninvasive brain stimulation technique. The effects of cathodal tDCS on convulsions and spatial memory after status epilepticus (SE) in immature animals were investigated.. Rats underwent lithium-pilocarpine-induced SE at postnatal day (P) 20-21 and received daily 30-min cathodal tDCS for 2 weeks at P23-36 through a unilateral epicranial electrode at 200μA. After tDCS, convulsions over 2 weeks were estimated by 20-h/day video monitoring. The rats were tested in a water maze for spatial learning at P50-53 and the brains were examined for cell loss and mossy fiber sprouting.. Long-term treatment with weak cathodal tDCS reduced SE-induced hippocampal cell loss, supragranular and CA3 mossy fiber sprouting, and convulsions (reduction of 21%) in immature rats. The tDCS treatment also rescued cognitive impairment following SE.. These findings suggested that cathodal tDCS has neuroprotective effects on the immature rat hippocampus after pilocarpine-induced SE, including reduced sprouting and subsequent improvements in cognitive performance. Such treatment might also have an antiepileptic effect.

Topics: Animals; Disease Models, Animal; Electric Stimulation Therapy; Hippocampus; Lithium Chloride; Male; Maze Learning; Memory Disorders; Mossy Fibers, Hippocampal; Nerve Degeneration; Pilocarpine; Rats; Rats, Wistar; Seizures; Status Epilepticus

Status epilepticus-induced hippocampal neuronal loss is mainly associated with excitotoxicity induced by increased levels of extracellular glutamate which is normally neutralized by high-affinity uptake mechanism. The energy source for the glutamate uptake is the electrochemical Na(+) gradient maintained by Na(+)/K(+) ATPase pump. In this study, we investigated the effect of early-life-induced status epilepticus on hippocampal Na(+)/K(+) ATPase activity and glutamate uptake. Rat pups 15 days old were injected i.p. with LiCl (3 mEq/kg) 12-18 h prior to s.c. pilocarpine administration (60 mg/kg). Hippocampal Na(+)/K(+) ATPase activity and glutamate uptake were evaluated 1.5, 12 and 24 h after SE induction. LiCl-pilocarpine-induced SE decreased Na(+)/K(+) ATPase activity and glutamate uptake by 42 and 38%, respectively, 1.5 h after SE induction. However, 12 and 24 h after SE induction the pump activity and glutamate uptake returned to control levels. SE early in life increased hippocampal number of degenerating neurons in the CA1 subfield and dentate gyrus 24 h after SE induction. In conclusion, SE induced early in life causes short-term disruption in hippocampal Na(+)/K(+) ATPase activity and glutamate uptake, which may be related to neuronal death found in CA1 subfield.

Topics: Animals; Convulsants; Enzyme Activation; Glutamic Acid; Hippocampus; Lithium Chloride; Male; Nerve Degeneration; Pilocarpine; Rats; Rats, Wistar; Sodium-Potassium-Exchanging ATPase; Status Epilepticus

In this study, we performed immunohistochemistry for somatostatin (SS), neuropeptide Y (NPY), and parvalbumin (PV) in LiCl-pilocarpine-treated rats to observe quantitative changes and axonal sprouting of GABAergic interneurons in the hippocampus, especially in the sclerotic hippocampus. Fluoro-Jade B (FJB) was performed to detect the specific degeneration of GABAergic interneurons. Compared with age-matched control rats, there were fewer SS/NPY/PV-immunoreactive (IR) interneurons in the hilus of the sclerotic hippocampus in pilocarpine-treated rats; hilar dentritic inhibitory interneurons were most vulnerable. FJB stain revealed degeneration was evident at 2 months after status epilepticus. Some SS-IR and NPY-IR interneurons were also stained for FJB, but there was no evidence of degeneration of PV-IR interneurons. Axonal sprouting of GABAergic interneurons was present in the hippocampus of epileptic rats, and a dramatic increase of SS-IR fibers was observed throughout all layers of CA1 region in the sclerotic hippocampus. These results confirm selective loss and degeneration of a specific subset of GABAergic interneurons in specific subfields of the hippocampus. Axonal sprouting of inhibitory GABAergic interneurons, especially numerous increase of SS-IR neutrophils within CA1 region of the sclerotic hippocampus, may constitute the aberrant inhibitory circum and play a significant role in the generation and compensation of temporal lobe epilepsy.

Topics: Animals; Axons; Disease Models, Animal; gamma-Aminobutyric Acid; Hippocampus; Interneurons; Lithium Chloride; Male; Nerve Degeneration; Neuropeptide Y; Parvalbumins; Pilocarpine; Rats; Rats, Sprague-Dawley; Sclerosis; Somatostatin; Status Epilepticus

Interictal spikes have been implicated in epileptogenesis and cognitive dysfunction in epilepsy. Unfortunately, antiepileptic drugs have shown poor efficacy in suppressing interictal discharges; novel therapies are needed. Surface charge on neuronal membranes provides a novel target for abolishing interictal spikes. This property can be modulated through the use of neuraminidase, an enzyme that decreases the amount of negatively charged sialic acid. In the present report we determined whether applying neuraminidase to brains of rats with a history of status epilepticus would reduce number of interictal discharges. Following pilocarpine-induced status epilepticus, rats received intrahippocampal injections of neuraminidase, which significantly decreased the number of interictal spikes recorded in the CA1 region. This study provides evidence that sialic acid degradation can reduce the number of interictal spikes. Furthermore, the results suggest that modifying surface charge created by negatively charged sialic acid may provide new opportunities for reducing aberrant epileptiform events in epilepsy.

Topics: Animals; Anticonvulsants; Convulsants; Electroencephalography; Epilepsy, Temporal Lobe; Evoked Potentials; Gliosis; Hippocampus; Injections; Injections, Intraperitoneal; Lithium Chloride; Male; N-Acetylneuraminic Acid; Neuraminidase; Pilocarpine; Rats; Rats, Sprague-Dawley; Signal Processing, Computer-Assisted; Status Epilepticus

Diabetic hyperglycemia is associated with seizure severity and may aggravate brain damage after status epilepticus. Our earlier studies suggest the involvement of ATP-sensitive potassium channels (K(ATP)) in glucose-related neuroexcitability. We aimed to determine whether K(ATP) agonist protects against status epilepticus-induced brain damage. Adult male Sprague-Dawley rats were divided into two groups: the streptozotocin (STZ)-induced diabetes (STZ) group and the normal saline (NS) group. Both groups were treated with either diazoxide (15 mg/kg, i.v.) (STZ + DZX, NS + DZX) or vehicle (STZ + V, NS + V) before lithium-pilocarpine-induced status epilepticus. We evaluated seizure susceptibility, severity, and mortality. The rats underwent Morris water-maze tests and hippocampal histopathology analyses 24 h post-status epilepticus. A multi-electrode recording system was used to study field excitatory postsynaptic synaptic potentials (fEPSP). RNA interference (RNAi) to knockdown Kir 6.2 in a hippocampal cell line was used to evaluate the effect of diazoxide in the presence of high concentration of ATP. Seizures were less severe (P < 0.01), post-status epilepticus learning and memory were better (P < 0.05), and neuron loss in the hippocampal CA3 area was lower (P < 0.05) in the STZ + DZX than the STZ + V group. In contrast, seizure severity, post-status epilepticus learning and memory, and hippocampal CA3 neuron loss were comparable in the NS + DZX and NS + V groups. fEPSP was lower in the STZ + DZX but not in the NS + DZX group. The RNAi study confirmed that diazoxide, with its K(ATP)-opening effects, could counteract the K(ATP)-closing effect by high dose ATP. We conclude that, by opening K(ATP), diazoxide protects against status epilepticus-induced neuron damage during diabetic hyperglycemia.

Topics: Adenosine Triphosphate; Analysis of Variance; Animals; Blood Glucose; Cell Line, Transformed; Diabetes Mellitus, Experimental; Diazoxide; Disease Models, Animal; Dose-Response Relationship, Drug; Excitatory Postsynaptic Potentials; Gene Expression Regulation; Hippocampus; In Situ Nick-End Labeling; In Vitro Techniques; Lithium Chloride; Male; Maze Learning; Neurons; Patch-Clamp Techniques; Potassium Channels, Inwardly Rectifying; Rats; Rats, Sprague-Dawley; RNA, Small Interfering; Status Epilepticus; Transfection; Vasodilator Agents

To evaluate the effects of high-frequency electrical stimulation (HFS) in both ventral hippocampi, alone and combined with a subeffective dose of antiepileptic drugs, during the status epilepticus (SE) induced by lithium-pilocarpine (LP).. Male Wistar rats, stereotactically implanted in both ventral hippocampi, were injected with pilocarpine (30 mg/kg, i.p.) 24 h after lithium (3 mEq/kg) administration. One minute following pilocarpine injection, HFS (pulses of 60 mus width at 130 Hz at subthreshold intensities and applied during 3 h) was applied alone or combined with subeffective doses of antiepileptic drugs.. HFS alone reduced the incidence of severe generalized seizures. This effect was not evident when HFS was combined with phenytoin (33.3 mg/kg, i.p.). HFS combined with diazepam (0.41 mg/kg, i.p.) or phenobarbital (10 mg/kg, i.p.) reduced the incidence of severe generalized seizures and mortality rate, and augmented the latency to first forelimb clonus, generalized seizure, and status epilepticus (SE). When combined with gabapentin (46 mg/kg, i.p.), HFS reduced the incidence of severe generalized seizures, enhanced latency to SE, and decreased mortality rate.. Subeffective doses of antiepileptic drugs that increase the gamma-aminobutyric acid (GABA)ergic neurotransmission may represent a therapeutic tool to augment the HFS-induced anticonvulsant effects.

Topics: Amines; Animals; Anticonvulsants; Cyclohexanecarboxylic Acids; Disease Models, Animal; Electric Stimulation; Electrodes, Implanted; Gabapentin; gamma-Aminobutyric Acid; Hippocampus; Lithium Chloride; Male; Phenytoin; Pilocarpine; Rats; Rats, Wistar; Seizures; Status Epilepticus; Synaptic Transmission

Depression represents one of the most common comorbidities of temporal lobe epilepsy (TLE), and has profound negative impact on the quality of life of patients with TLE. However, causes and mechanisms of depression in TLE remain poorly understood, and effective therapies are lacking. We examined whether a commonly used model of TLE in rats could be used as a model of comorbidity between epilepsy and depression suitable for both mechanistic studies and for the development of mechanism-based antidepressant therapies. We established that animals that had been subjected to lithium chloride and pilocarpine status epilepticus (SE) and developed spontaneous recurrent seizures, exhibited a set of impairments congruent with a depressive state: behavioral equivalents of anhedonia and despair, dysregulation of the hypothalamus-pituitary-adrenal (HPA) axis, and compromised raphe-hippocampal serotonergic transmission. Pharmacologic studies have suggested that depressive impairments following SE develop as a result of enhanced interleukin-1beta signaling in the hippocampus, which leads to depression via inducing perturbations in the HPA axis and subsequent deficit in the raphe-hippocampal serotonergic transmission.

Topics: Animals; Convulsants; Depressive Disorder; Disease Models, Animal; Epilepsy, Temporal Lobe; Hippocampus; Humans; Hypothalamo-Hypophyseal System; Interleukin-1beta; Lithium Chloride; Pilocarpine; Pituitary-Adrenal System; Raphe Nuclei; Rats; Receptors, Glucocorticoid; Serotonin; Signal Transduction; Status Epilepticus; Synaptic Transmission

Although the number of antiepileptic drugs (AEDs) is increasing, none displays neuroprotective or antiepileptogenic properties that could prevent status epilepticus (SE)-induced drug-resistant epilepsy. Ketogenic diet (KD) and calorie restriction (CR) are proposed as alternative treatments in epilepsy. Our goal was to assess the neuroprotective or antiepileptogenic effect of these diets in a well-characterized model of mesial temporal lobe epilepsy following initial SE induced by lithium-pilocarpine in adult rats.. Seventy-five P50 male Wistar rats were fed a specific diet: normocalorie carbohydrate (NC), hypocalorie carbohydrate (HC), normocalorie ketogenic (NK), or hypocalorie ketogenic (HK). Rats were subjected to lithium-pilocarpine SE, except six NC to constitute a control group for histology (C). Four rats per group were implanted with epidural electrodes to record electroencephalography (EEG) during SE and the next six following days. From the seventh day, the animals were video-recorded 10 h daily to determine latency to epilepsy onset. Neuronal loss in hippocampus and parahippocampal cortices was analyzed 1 month after the first spontaneous seizure.. After lithium-pilocarpine injection, neither KD nor CR modified SE features or latency to epilepsy. In hippocampal layers, KD or CR exhibited a neuroprotective potential without cooperative effect. Parahippocampal cortices were not protected by the diets.. The antiepileptic effect of KD and/or CR is overwhelmed by lithium-pilocarpine injection. The isolated protection of hippocampal layers induced by KD or CR or their association failed to modify the course of epileptogenesis.

Topics: Animals; Anticonvulsants; Caloric Restriction; Cerebral Cortex; Diet, Ketogenic; Dietary Carbohydrates; Disease Models, Animal; Drug Resistance; Electroencephalography; Epilepsy, Temporal Lobe; Hippocampus; Humans; Lithium Chloride; Male; Neuroprotective Agents; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Status epilepticus (SE) leads to serious damage in hippocampus of the adult brain. Much less is known about immature brain where neuronal degeneration may have different localization and time course. Lithium-pilocarpine SE was induced in 12-day-old male Wistar rats. Six different intervals after SE (from 4 h to 1 week) were studied using Fluoro-Jade B staining. Three to four animals were used for every interval. Severity of damage in individual parts of hippocampal formation was semi-quantified. A consistent neuronal damage occurred in all hippocampal fields (CA 1, CA 3, dentate gyrus) at all survival intervals. Hippocampal fields CA 1 and CA 3 exhibited degeneration of interneurons located mainly in stratum oriens and pyramidale at shorter intervals (4-12h). Massive degeneration of pyramidal cells started at 24h in CA 1 and at 48 h in CA 3. Dentate gyrus exhibited degenerating neurons in granular layer with a peak at short intervals (4-8 h), and molecular layer was spared. The lower blade of dentate gyrus was more affected than the upper blade. Damage of hilar neurons was negligible. Our results demonstrate that SE elicited in immature rats causes acute neurodegeneration in the hippocampus. Time course of this degeneration is different for individual parts of hippocampal formation and for individual cell types.

Topics: Animals; Animals, Newborn; Dentate Gyrus; Disease Models, Animal; Hippocampus; Lithium Chloride; Male; Nerve Degeneration; Pilocarpine; Pyramidal Cells; Rats; Rats, Wistar; Status Epilepticus; Time Factors

In the present study we analyzed aquaporin-4 (AQP4) immunoreactivity in the piriform cortex (PC) and the hippocampus of pilocarpine-induced rat epilepsy model to elucidate the roles of AQP4 in brain edema following status epilepticus (SE). In non-SE-induced animals, AQP4 immunoreactivity was diffusely detected in the PC and the hippocampus. AQP4 immunoreactivity was mainly observed in the endfeet of astrocytes. Following SE the AQP4-deleted area was clearly detected in the PC, not in the hippocampus. Decreases in dystrophin and α-syntrophin immunoreactivities were followed by reduction in AQP4 immunoreactivity. These alterations were accompanied by the development of vasogenic edema and the astroglial loss in the PC. In addition, acetazolamide (an AQP4 inhibitor) treatment exacerbated vasogenic edema and astroglial loss both in the PC and in the hippocampus. These findings suggest that SE may induce impairments of astroglial AQP4 functions via disruption of the dystrophin/α-syntrophin complex that worsen vasogenic edema. Subsequently, vasogenic edema results in extensive astroglial loss that may aggravate vasogenic edema.

Topics: Animals; Aquaporin 4; Astrocytes; Brain Edema; Calcium-Binding Proteins; Cell Count; Cerebral Cortex; Disease Models, Animal; Dystrophin; Gene Expression Regulation; Glial Fibrillary Acidic Protein; Hippocampus; Lithium Chloride; Male; Membrane Proteins; Muscle Proteins; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

Annexin A7 (synexin, ANXA7) is a member of annexins, which plays an essential role in the regulation of calcium homeostasis. Considerable evidence shows that the pathogenetic mechanism of acquired epilepsy (AE) has been related to the imbalance of calcium homeostasis. The aim of this study was to investigate ANXA7 expression and cellular localization in the cortex and hippocampus in the rat lithium-pilocarpine model of AE.. Totally 81 adult healthy male Wistar rats were randomly divided into control group (n = 9) and experimental group (n = 72), the experimental group contained eight subgroups according to sacrifice time (n = 9) (6-hour, 24-hour, 48-hour, 72-hour, 7-day, 15-day, 1-month, and 2-month). In the experimental group, rats were intraperitoneally injected by lithium-pilocarpine to induce AE model. We examined the expression and localization of ANXA7 via immunohistochemistry, double-label immunofluorescence with the use of neuron specific enolase (NSE) antibody, glial fibrillary acidic protein (GFAP) antibody and propidium iodide (PI), respectively. The data of optical density value were analyzed by analysis of variance.. ANXA7 expression increased significantly in the experimental groups especially in the acute period (6 hours, 24 hours, and 48 hours after the onset of seizure) using immunohistochemistry. Double-label immunofluorescence and confocal microscopy disclosed that ANXA7 localized in the neurons but not in astrocytes and did not localize in the nucleus, which were performed with anti-NSE, anti-GFAP and PI respectively.. ANXA7 may play a potential role in the pathogenetic mechanisms of the rat lithium-pilocarpine model of AE.

Topics: Animals; Annexin A7; Calcium; Cerebral Cortex; Disease Models, Animal; Fluorescent Antibody Technique; Hippocampus; Immunohistochemistry; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Status epilepticus (SE) activates the autonomic nervous system, increasing sympathetic nervous system control of cardiac function during seizure activity. However, lasting effects of SE on autonomic regulation of the heart, which may contribute to mortality following seizure activity, are unknown. Therefore, autonomic control of cardiac function was assessed following SE.. Using Sprague-Dawley rats after 1-2 weeks of recovery from lithium-pilocarpine-induced SE or control procedures, we tested overall sympathovagal control of the heart, the individual contributions of the sympathetic and parasympathetic components of the autonomic nervous system, and baroreflex sensitivity.. SE induced a chronic shift in sympathovagal balance toward sympathetic dominance resulting from decreased parasympathetic activity. Baroreflex sensitivity to increased blood pressure was also decreased, likely resulting from diminished vagal activation.. Chronic alterations in autonomic regulation of cardiac function, characterized by increased sympathetic dominance, occur following SE and likely contribute to subsequent increased cardiac risk and mortality.

Topics: Analysis of Variance; Animals; Atropine; Baroreflex; Blood Pressure; Disease Models, Animal; Heart Rate; Lithium Chloride; Male; Muscarinic Antagonists; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Sympathetic Nervous System; Time Factors; Vagus Nerve

Topiramate, an antiepileptic drug with multiple mechanisms of action, was assessed as a neuroprotective agent following status epilepticus. We administered topiramate or normal saline chronically beginning 1 hour after cessation of lithium pilocarpine-induced status epilepticus. Control animals not subjected to status epilepticus were also treated with topiramate or normal saline. Following completion of the topiramate treatment, animals were tested in the water maze to assess spatial learning and underwent in vivo single-cell place cell recordings. Spontaneous seizure frequency following status epilepticus in the topiramate-treated rats was similar to that in the rats treated with saline. Following status epilepticus, rats had profound deficits in water maze performance and place cell function. Rats subjected to status epilepticus and treated with topiramate were also severely impaired in the water maze, but performed slightly better than rats treated with saline. Following status epilepticus, topiramate-treated rats did not differ from rats treated with normal saline in the platform switch, a test of prefrontal function. Although place cell firing patterns were similar in both the topiramate- and saline-treated rats, rats treated with topiramate had higher information content scores than rats treated with saline. Topiramate-treated animals had less supragranular sprouting following status epilepticus than nontreated rats. Control animals treated with topiramate did not differ from saline-treated controls on any measures. Taken together, this study shows that topiramate administered following status epilepticus has modest neuroprotective effects.

Topics: Animals; Anticonvulsants; Convulsants; Electrodes, Implanted; Electroencephalography; Electrophysiology; Fructose; Hippocampus; Lithium Chloride; Male; Maze Learning; Neurons; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Status Epilepticus; Topiramate

Status epilepticus (SE) is one of the most serious manifestations of epilepsy. Systemic inflammation and damage of blood-brain barrier (BBB) are etiologic cofactors in the pathogenesis of pilocarpine SE while acute osmotic disruption of the BBB is sufficient to elicit seizures. Whether an inflammatory-vascular-BBB mechanism could apply to the lithium-pilocarpine model is unknown. LiCl facilitated seizures induced by low-dose pilocarpine by activation of circulating T-lymphocytes and mononuclear cells. Serum IL-1beta levels increased and BBB damage occurred concurrently to increased theta EEG activity. These events occurred prior to SE induced by cholinergic exposure. SE was elicited by lithium and pilocarpine irrespective of their sequence of administration supporting a common pathogenetic mechanism. Since IL-1beta is an etiologic trigger for BBB breakdown and its serum elevation occurs before onset of SE early after LiCl and pilocarpine injections, we tested the hypothesis that intravenous administration of IL-1 receptor antagonists (IL-1ra) may prevent pilocarpine-induced seizures. Animals pre-treated with IL-1ra exhibited significant reduction of SE onset and of BBB damage. Our data support the concept of targeting systemic inflammation and BBB for the prevention of status epilepticus.

Topics: Action Potentials; Animals; Anti-Inflammatory Agents, Non-Steroidal; Blood-Brain Barrier; Brain; Enzyme-Linked Immunosorbent Assay; Gliosis; Interleukin 1 Receptor Antagonist Protein; Interleukin-1beta; Leukocytes; Lithium Chloride; Male; Microelectrodes; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Theta Rhythm

Recent evidence suggests that aberrant neuro/gliogenesis and/or inflammation play critical roles in epileptogenesis. Although the plastic and inflammatory changes have been described in the postseizure hippocampus, little data is available concerning extrahippocampal regions, notably in the piriform and entorhinal cortices, amygdala, and parts of the thalamus. In this study, we examined histological changes in whole epileptic rat brain, with respect to cell death, cell genesis, and inflammation.. Experimental status epilepticus (SE) was induced using a lithium-pilocarpine injection. Neuronal death was evident in the amygdala, piriform, and entorhinal cortices, as well as the subfields of hippocampus. Microglial activation was observed in more extended limbic areas, such as, the hippocampus, entorhinal, perirhinal and piriform cortices, amygdala, thalamus, and hypothalamus, and a robust increase of cell genesis was noted in these damaged areas. The majority of newly generated cells in extrahippocampal areas proliferated in situ, and differentiated mainly into astrocytes or oligodendrocytes. In addition, stromal cell-derived factor-1alpha was found to be induced in close temporal and anatomical association with seizure-induced plasticity.. These findings indicate that neuronal death, inflammation, and cell genesis are substantially associated throughout the entire brain and that they may influence the epileptogenic process and clinical manifestations.

Topics: Amygdala; Animals; Astrocytes; Cell Differentiation; Cell Division; Cell Survival; Chemokine CXCL12; Entorhinal Cortex; Hippocampus; Lithium Chloride; Male; Microglia; Neurogenesis; Neuronal Plasticity; Oligodendroglia; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Thalamus; Up-Regulation

Frequent epileptic seizures or prolonged seizure activity (status epilepticus, SE) is known to increase the brain expression of drug efflux transporter genes and proteins, such as P-glycoprotein (Pgp) and members of the multidrug resistance protein (MRP) family, which might reduce brain levels of antiepileptic drugs and, therefore, be involved in drug resistance. However, the time course of alterations in Pgp or MRPs after seizures or SE is only incompletely known.. This prompted us to study the time course of alterations in the expression of different efflux transporter genes (Mdr1a, Mdr1b, MRP1, MRP2, MRP5) at various times after a pilocarpine-induced SE in limbic brain regions, using quantitative real-time polymerase chain reaction (RT-PCR) (qPCR).. Unexpectedly, between 6 and 24 h after onset of SE, genes encoding Pgp (Mdr1a, Mdr1b), Mrp1, and Mrp5 were downregulated in hippocampus, amygdala, or piriform cortex. This initial decrease in expression was followed by normalization and then increased expression, which became maximal 2 days after SE. One explanation for the initial decrease in transporter expression could be SE-induced acute inflammatory processes, because proinflammatory cytokines are known to suppress the expression of Pgp and other efflux transporters. To directly address this possibility, we quantified the hippocampal mRNA expression of interleukin-1beta, interleukin-6, and tumor necrosis factor-alpha, showing a marked SE-induced increase in these cytokines, which paralleled the decreased expression of efflux transporters.. Taken together, these findings indicate that alterations in expression of drug efflux transporters after prolonged seizure activity are more complex than previously thought.

Topics: Analysis of Variance; Animals; ATP Binding Cassette Transporter, Subfamily B, Member 1; Brain; Cyclooxygenase 2; Cytokines; Disease Models, Animal; Female; Gene Expression Regulation; Lithium Chloride; Multidrug Resistance-Associated Proteins; Pilocarpine; Rats; Rats, Wistar; RNA, Messenger; Status Epilepticus; Time Factors

Progression of severity in experimental status epilepticus (SE), defined as refractoriness to first- and second-line abortive agents, may be related to a five-stage progression of electroencephalography (EEG) patterns. This was tested in the lithium-pilocarpine rat SE model. Abortive treatment with diazepam and phenobarbital was given at EEG stages I, III, and V. In stage I, the combination therapy resulted in 100% SE termination. However, stage III corresponded to high treatment resistance (0% abortion) and stage V to an intermediate response (63%). Comparisons of time-to-treatment durations showed overlap between stage I and stage III, despite having markedly different response rates to abortive medications. Therefore, EEG patterns reflect the dynamic pathophysiology of SE and can be used as reliable and specific markers to distinguish treatment-responsive from treatment-refractory SE more accurately than time alone.

Topics: Analysis of Variance; Animals; Anticonvulsants; Diazepam; Disease Models, Animal; Disease Progression; Electroencephalography; Lithium Chloride; Male; Phenobarbital; Pilocarpine; Predictive Value of Tests; Rats; Rats, Sprague-Dawley; Reaction Time; Status Epilepticus; Time Factors

Significant increases in local cerebral blood flow during lithium-pilocarpine (Li-P) induced seizure have been reported. We recently found that both acetone and methyl ethyl ketone (MEK) showed anticonvulsive effects in status epilepticus induced by Li-P in rats. In this study, we examined whether MEK also suppressed the enhancement of local cerebral blood flow induced by Li-P with a simplified autoradiographic method using [(14)C]-para-iodo-N-isopropyl amphetamine ([(14)C]-IMP). Significant increases in local cerebral blood flow in the thalamus, hypothalamus, hippocampus and cerebellum were observed in Li-P induced status epilepticus rats. Pretreatment with MEK (8 mmol/kg) completely suppressed the enhancement of local cerebral blood flow to or below the control level in all regions.

Topics: Animals; Anticonvulsants; Autoradiography; Butanones; Carbon Radioisotopes; Cerebellum; Cerebrovascular Circulation; Hippocampus; Hypothalamus; Iofetamine; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Thalamus

A ketogenic diet has been used successfully to treat patients with intractable epilepsy, although the mechanism is unknown. Acetone has been shown to have an anticonvulsive effect in various animal models. The main purpose of this study was to determine whether other ketones, 2-butanone (methyl ethyl ketone: MEK) and 3-pentanone (diethyl ketone: DEK), also show anticonvulsive effects in lithium-pilocarpine (Li-pilocarpine)-induced status epilepticus (SE) in rats.. Anticonvulsive effects of MEK and DEK in Li-pilocarpine SE rats were measured by behavioural scoring. Anti-seizure effects of MEK were also evaluated using electroencephalography (EEG). Neuroprotective effect of MEK was investigated by haematoxylin and eosin staining 4 weeks after the treatment with pilocarpine.. Acetone, MEK and DEK showed anticonvulsant effects in Li-pilocarpine-induced SE rats. Treatment with MEK twice (8 mmol.kg(-1) and 5 mmol.kg(-1)) almost completely blocked spontaneous recurrent cortical seizure EEG up to 4 weeks after the administration of pilocarpine. MEK also showed strong neuroprotective effects in Li-pilocarpine-treated rats 4 weeks following the administration of pilocarpine. Significant neural cell death occurred in the hippocampus of Li-pilocarpine SE rats, especially in the CA1 and CA3 subfields. In contrast, normal histological characteristics were observed in these regions in the MEK-pretreated rats.. Both MEK and DEK showed strong anticonvulsive effects in Li-pilocarpine-induced SE rats. They also inhibited continuous recurrent seizure and neural damage in hippocampal region for 4 weeks after the treatment with pilocarpine. These findings appear to be of value in the investigation of epilepsy.

Topics: Acetone; Animals; Anticonvulsants; Butanones; Diet, Ketogenic; Hippocampus; Lithium Chloride; Male; Neuroprotective Agents; Pentanones; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Status epilepticus (SE) is a seizure or series of seizures that persist for >30 min and often results in mortality. Death rarely occurs during or immediately following seizure activity, but usually within 30 days. Although ventricular arrhythmias have been implicated in SE-related mortality, the effects of this prolonged seizure activity on the cardiac function and susceptibility to arrhythmias have not been directly investigated. We evaluated myocardial damage, alterations in cardiac electrical activity, and susceptibility to experimentally induced arrhythmias produced by SE in rats. SE resulted in seizure-related increases in blood pressure, heart rate, and the first derivative of pressure, as well as modest, diffuse myocyte damage assessed by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling staining. Ten to twelve days following seizures, electrocardiographic recordings showed arrhythmogenic alterations in cardiac electrical activity, denoted by prolonged QT interval corrected for heart rate and QT dispersion. Finally, SE increased susceptibility to experimentally induced (intravenous aconitine) cardiac arrhythmias. These data suggest that SE produces tachycardic ischemia following the activation of the sympathetic nervous system, resulting in cardiac myofilament damage, arrhythmogenic alterations in cardiac electrical activity, and increased susceptibility to ventricular arrhythmias.

Topics: Aconitine; Actin Cytoskeleton; Action Potentials; Animals; Arrhythmias, Cardiac; Biomarkers; Blood Pressure; Disease Models, Animal; Disease Susceptibility; Electrocardiography; Heart Rate; In Situ Nick-End Labeling; Lithium Chloride; Male; Myocardium; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Sympathetic Nervous System; Time Factors; Troponin I

Ca2+-stimulated protein kinase II (CaMKII) is critically involved in the regulation of synaptic function and is implicated in the neuropathology associated with ischemia and status epilepticus (SE). The activity and localization of CaMKII is regulated by multi-site phosphorylation. In the present study we investigated the effects of global ischemia followed by reperfusion and of SE on the phosphorylation of CaMKII on T253 in rat forebrains and compared this to the phosphorylation of T286. Both ischemia and SE resulted in marked increases in the phosphorylation of T253, and this was particularly marked in the postsynaptic density (PSD). Phosphorylation of T286 decreased rapidly towards basal levels following ischemia whereas phosphorylation of T253 remained elevated for between 1 and 6 h before decreasing to control values. Following SE, phosphorylation of T253 remained elevated for between 1 and 3 h before decreasing to control levels. In contrast, phosphorylation of T286 remained elevated for at least 24 h following the termination of SE. Total CaMKII associated with PSDs transiently increased 10 min following ischemia, but only several hours following SE. The results demonstrate that phoshorylation of CaMKII on T253 is enhanced following both ischemia/reperfusion and SE and indicate that the phosphorylation of T253 and T286 are differentially regulated.

Topics: Animals; Calcium; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Disease Models, Animal; Ischemia; Lithium Chloride; Male; Phosphorylation; Pilocarpine; Prosencephalon; Rats; Rats, Wistar; Status Epilepticus; Synaptosomes; Threonine

Depression represents one of the most common comorbidities in patients with epilepsy. However, the mechanisms of depression in epilepsy patients are poorly understood. Establishment of animal models of this comorbidity is critical for both understanding the mechanisms of the condition, and for preclinical development of effective therapies. The current study examined whether a commonly used animal model of temporal lobe epilepsy (TLE) is characterized by behavioural and biochemical alterations involved in depression. Male Wistar rats were subjected to LiCl and pilocarpine status epilepticus (SE). The development of chronic epileptic state was confirmed by the presence of spontaneous seizures and by enhanced brain excitability. Post-SE animals exhibited increase in immobility time under conditions of forced swim test (FST) which was indicative of despair-like state, and loss of taste preference in saccharin solution consumption test which pointed to the symptomatic equivalence of anhedonia. Biochemical studies revealed compromised serotonergic transmission in the raphe-hippocampal serotonergic pathway: decrease of serotonin (5-HT) concentration and turnover in the hippocampus, measured by high performance liquid chromatography, and decrease of 5-HT release from the hippocampus in response to raphe stimulation, measured by fast cyclic voltammetry. Administration of fluoxetine (FLX, 20 mg/kg/day for 10 days) to naive animals significantly shortened immobility time under conditions of FST, and inhibited 5-HT turnover in the hippocampus. In post-SE rats FLX treatment led to a further decrease of hippocampal 5-HT turnover; however, performance in FST was not improved. At the same time, FLX reversed SE-induced increase in brain excitability. In summary, our studies provide initial evidence that post-SE model of TLE might serve as a model of the comorbidity of epilepsy and depression. The finding that behavioural equivalents of depression were resistant to an antidepressant medication suggested that depression in epilepsy might have distinct underlying mechanisms beyond alterations in serotonergic pathways.

Topics: Animals; Behavior, Animal; Depression; Epilepsy, Temporal Lobe; Fluoxetine; Hippocampus; Lithium Chloride; Male; Models, Animal; Pilocarpine; Rats; Rats, Wistar; Selective Serotonin Reuptake Inhibitors; Serotonin; Status Epilepticus

The A-type voltage-gated potassium channels (Kv4) have been proved to play a major role as modulators of somatodendritic excitability. Recent studies indicate that neuronal hyperactivity in epilepsy is associated with changes in Kv4. However, the precise regulation of Kv4 in the development of epilepsy and its underlying mechanism remain unclear. In this study, we investigated whether the expression of the Kv4.2 channel and of its major modulator, voltage-dependent potassium channel-interacting protein (KChIP1), is altered following lithium-pilocarpine induced status epilepticus (SE) and the chronic-epilepsy phase in the rat model. We found that Kv4.2 and KChIP1 expression was transiently up-regulated following SE, whereas it was down-regulated during the chronic phase: this was most prominent in the CA1 and CA3 regions. The time-course analysis of the protein expression level showed that the peak Kv4.2 up-regulation was between 6 and 24 h after SE, whereas KChIP1 expression was increased earlier and for a shorter period. The temporospatial changes in Kv4.2 were very similar to those of its major modulator KChIP1. We compared the difference in 4-aminopyridine (4-AP)-induced intracellular calcium ([Ca(2+)]i) elevation between model and control brain slices. The results showed that the [Ca(2+)]i elevation induced by the Kv4 channel blocker 4-AP was aggravated and prolonged in the model slice after SE. The functional relevance of these changes in Ca(2+) homeostasis and Kv4.2 and KChIP1 expression may be associated with intrinsic neuronal excitability regulation and epileptogenesis.

Topics: 4-Aminopyridine; Animals; Calcium; Disease Models, Animal; Extracellular Fluid; Female; Gene Expression Regulation; Hippocampus; In Vitro Techniques; Kv Channel-Interacting Proteins; Lithium Chloride; Pilocarpine; Potassium Channel Blockers; Rats; Rats, Sprague-Dawley; Shal Potassium Channels; Status Epilepticus; Time Factors

This study investigated putative correlations among behavioral changes and: (1) neuronal loss, (2) hippocampal mossy fiber sprouting, and (3) reactive astrogliosis in adult rats submitted to early-life LiCl-pilocarpine-induced status epilepticus (SE).. Rats (P15) received LiCl (3 mEq/kg, i.p.) 12-18 h prior pilocarpine (60 mg/kg; s.c.). At adulthood, animals were submitted to behavioral tasks and after the completion of tasks biochemical and histological analysis were performed.. In SE group, it was observed an increased number of degenerating neurons in the CA1 subfield and in the hilus of animals 24 h after SE. At adulthood, SE group presented an aversive memory deficit in an inhibitory avoidance task and the animals that presented lower latency to the step down showed a higher score for mossy fiber sprouting. In the light-dark exploration task, SE rats returned less and spent less time in the light compartment and present an increased number of risk assessment behavior (RA). There was a negative correlation between the time spent in the light compartment and the score for mossy fiber sprouting and a positive correlation between score for mossy fiber sprouting and number of RA. LiCl-pilocarpine-treated animals showed higher levels of S100B immunocontent in the CSF as well as a positive correlation between the score for sprouting and the GFAP immunocontent in the CA1 subfield, suggesting an astrocytic response to neuronal injury.. We showed that LiCl-pilocarpine-induced SE during development produced long-lasting behavioral abnormalities, which might be associated with mossy fiber sprouting and elevated CSF S100B levels at adulthood.

Topics: Age Factors; Animals; Animals, Newborn; Anxiety; Behavior, Animal; Fear; Hippocampus; Lithium Chloride; Memory; Mossy Fibers, Hippocampal; Nerve Growth Factors; Pilocarpine; Rats; Rats, Wistar; S100 Calcium Binding Protein beta Subunit; S100 Proteins; Status Epilepticus

Status epilepticus (SE) is a severe neurological condition that can result in brain damage. In animals, SE is associated with cell loss and aberrant synaptogenesis. These pathological processes appear to be activity-dependent and may continue after the SE has ended. We postulated that suppression of electrical activity following SE at the site of the epileptic focus will reduce seizure-induced damage. To achieve this goal, tetrodotoxin (TTX) was used to suppress electrical activity in the hippocampi bilaterally following SE. Adult rats experienced lithium-pilocarpine-induced SE for 2h while controls underwent sham-SE with saline injections. Starting 12h after the SE or sham-SE rats received either continuous TTX (1 microM) or saline infusions through cannulas implanted in the bilateral hippocampi for 5h daily for 4 days. TTX resulted in significant EEG suppression and reduction in spikes and sharp waves. Rats were sacrificed 2 weeks after SE and the brains examined for cell loss and sprouting. Rats receiving TTX following SE had significantly more cell loss as well as a trend toward more mossy fiber sprouting than saline-treated rats following SE. TTX injection in sham-SE rats caused no cell loss or mossy fiber sprouting. These results suggest that suppression of electrical activity following SE is detrimental.

Topics: Analysis of Variance; Anesthetics, Local; Animals; Disease Models, Animal; Electroencephalography; Hippocampus; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Sprague-Dawley; Staining and Labeling; Status Epilepticus; Tetrodotoxin

Lithium-pilocarpine status epilepticus (SE) resulted in delayed changes of single cortical interhemisperic (transcallosal) responses in immature rats. Low-frequency stimulation inducing depression and/or potentiation was studied to analyze possible dynamic changes in cortical responses. Status was elicited in 12-day-old (SE12) or 25-day-old (SE25) rats. Control siblings received saline instead of pilocarpine. Interhemispheric responses were elicited by stimulation of the sensorimotor region of the cerebral cortex 3, 6, 9, 13, or 26 days after status. A series of 5 biphasic pulses with intensity equal to twofold threshold were used for stimulation. The interval between pulses was 100, 125, 160, 200 or 300 ms, eight responses were always averaged. Peak amplitude of the first positive, first negative and second positive waves was measured and responses to the second, third, fourth and fifth pulse were compared with the first one. Animals after status epilepticus as well as lithium-paraldehyde controls exhibit a frequency depression at nearly all the intervals studied. An outlined increase of responses in SE rats in comparison with the controls three days after SE stayed just below the level of statistical significance. In addition, animals in the SE12 group exhibited potentiation of responses at this interval after SE. With longer intervals after SE, the relation between SE and control animals changed twice resulting in a tendency to lower amplitude of responses in SE than in control rats 26 days after SE. Rats in the SE25 group exhibited higher responses than controls 13 days after status, but this difference was not present at the longest interval after SE. Low-frequency stimulation did not reveal increased cortical excitability as a long-lasting consequence of status epilepticus induced in immature rats. In addition, the outlined differences between SE and control rats changed with the time after SE.

Topics: Animals; Cerebral Cortex; Disease Models, Animal; Electric Stimulation; Electroencephalography; Evoked Potentials; Lithium Chloride; Long-Term Potentiation; Long-Term Synaptic Depression; Male; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Time Factors

Behavioral, electroencephalographic (EEG) and neuropathological effects of methomyl, a carbamate insecticide reversibly inhibiting acetylcholinesterase activity, were studied in naive or lithium chloride (24 h, 3 mEq/kg, s.c.) pretreated male Wistar rats. In naive animals, methomyl with equal potency produced motor limbic seizures and fatal status epilepticus. Thus, the CD50 values (50% convulsant dose) for these seizure endpoints were almost equal to the LD50 (50% lethal dose) of methomyl (13 mg/kg). Lithium pretreated rats were much more susceptible to convulsant, but not lethal effect of methomyl. CD50 values of methomyl for motor limbic seizures and status epilepticus were reduced by lithium pretreatment to 3.7 mg/kg (a 3.5-fold decrease) and 5.2 mg/kg (a 2.5-fold decrease), respectively. In contrast, lithium pretreatment resulted in only 1.3-fold decrease of LD50 value of methomyl (9.9 mg/kg). Moreover, lithium-methomyl treated animals developed a long-lasting status epilepticus, which was not associated with imminent lethality observed in methomyl-only treated rats. Scopolamine (10 mg/kg) or diazepam (10 mg/kg) protected all lithium-methomyl treated rats from convulsions and lethality. Cortical and hippocampal EEG recordings revealed typical epileptic discharges that were consistent with behavioral seizures observed in lithium-methomyl treated rats. In addition, convulsions induced by lithium-methomyl treatment were associated with widespread neurodegeneration of limbic structures. Our observations indicate that lithium pretreatment results in separation between convulsant and lethal effects of methomyl in rats. As such, seizures induced by lithium-methomyl administration may be an alternative to lithium-pilocarpine model of status epilepticus, which is associated with high lethality.

Topics: Animals; Anticonvulsants; Diazepam; Disease Models, Animal; Drug Synergism; Electroencephalography; Lithium Chloride; Male; Methomyl; Rats; Rats, Wistar; Scopolamine; Seizures; Status Epilepticus

It is well known that status epilepticus (SE) becomes increasingly difficult to control over time. Previous studies have indicated that the electroencephalographic pattern at the time of intervention is predictive of the probability of successful treatment. However, these findings are based on studies limited to the first 2h of SE onset. Little data is available on the efficacy of treating SE at later time points.. The aim of the present study was to investigate the efficacy of diazepam (DZP) treatment given at two different phases of SE in a lithium-pilocarpine rat model: during continuous ictal discharges (CIDs, phase 3), and during late periodic epileptiform discharges (late PEDs, phase 5). Changes in cortical and hippocampal electroencephalographs (EEGs) were observed continuously during the phases of SE, as well as at 24, 36, 48, and 72h after SE onset. The effects of DZP treatment during CIDs or during late PEDs were compared to control DZP-untreated rats.. In all three groups, hippocampal and cortical EEGs displayed five distinct phases of SE. There were no statistical differences in the duration of phases 1 and 2 among the three groups. Although DZP administration during CIDs did not terminate CIDs in most rats, it did significantly shorten the duration of phases 3 and 4 of SE. Importantly, DZP given during phase 5 successfully ended behavioral and electrographic seizures in most rats.. Hippocampal and cortical EEGs displayed five distinct phases of SE that were similarly responsive to DZP treatment. Termination of electrographic seizures with DZP treatment was more effective in the last phase of SE (late PEDs) than in phase 3 (CIDs). These findings suggest that previous reports of DZPs decrease in efficacy over time may not be applicable to DZP treatment at 4h-post onset.

Topics: Animals; Anticonvulsants; Behavior, Animal; Convulsants; Diazepam; Electroencephalography; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

The mode and mechanism of neuronal death induced by status epilepticus (SE) in the immature brain have not been fully characterized. In this study, we analyzed the contribution of neuronal necrosis and caspase-3 activation to CA1 damage following lithium-pilocarpine SE in P14 rat pups. By electron microscopy, many CA1 neurons displayed evidence of early necrosis 6 hours following SE, and the full ultrastructural features of necrosis at 24-72 hours. Caspase-3 was activated in injured (acidophilic) neurons 24 hours following SE, raising the possibility that they died by caspase-dependent "programmed" necrosis.

Topics: Animals; Animals, Newborn; Autophagy; Brain; Caspase 3; Cell Death; Disease Models, Animal; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Female; Lithium Chloride; Male; Microscopy, Electron; Necrosis; Neurons; Pilocarpine; Pyramidal Cells; Rats; Rats, Wistar; Status Epilepticus

An enriched environment can enhance brain recovery in animals with early-life status epilepticus (SE). The purpose of this study was to determine the effects of early-life SE on spatial memory and hippocampal extracellular signal-regulated kinase (ERK) level, and the possible therapeutic effects of the enriched environment. Rats were assigned randomly to four groups: (1) control rats (nonenriched control); (2) control rats housed in an enriched environment from Postnatal Day (P) 25 to P40 (enriched control); (3) rats in which SE was induced with lithium-pilocarpine (Li-PC) at P21 (nonenriched SE); and (4) rats in which SE was induced with Li-PC at P21 and then housed in an enriched environment from P25 to P40 (enriched SE). As adults, the rats underwent spatial learning and memory tests in the Morris water maze between P50 and P55. At P55, subsets of animals were evaluated for expression of hippocampal ERK1/2 phosphorylation immediately following completion of the Morris water maze. At ~P100, another set of animals was tested for seizure threshold. When studied as adults, only the nonenriched SE group had a spatial memory deficit. The nonenriched SE group also exhibited lower levels of phosphorylated ERK2 as compared with the nonenriched control, enriched control, and enriched SE groups. Both the nonenriched SE and enriched SE groups had reduced seizure thresholds as compared with the nonenriched control and enriched control groups. Results from this study demonstrate that an enriched environment improves spatial memory in rats subjected to early-life SE, possibly through upregulation of phosphorylated ERK2 in the hippocampus. However, an enriched environment has no effect on seizure threshold.

Topics: Analysis of Variance; Animals; Animals, Newborn; Behavior, Animal; Cognition; Disease Models, Animal; Environment; Hippocampus; Lithium Chloride; Male; Maze Learning; Mitogen-Activated Protein Kinase 1; Phosphorylation; Pilocarpine; Rats; Rats, Sprague-Dawley; Spatial Behavior; Status Epilepticus; Time Factors

To try to identify the critical structures during epileptogenesis, we used the lithium-pilocarpine model that reproduces most clinical and neuropathological features of temporal lobe epilepsy (TLE). We used imaging techniques as well as a disease modifying approach and pharmacological strategy. With [14C]-2-deoxyglucose autoradiography, we assessed changes in cerebral glucose utilization. T2-weighted magnetic resonance imaging (MRI, 4.7 T) allowed follow-up of structures involved in epileptogenesis. A potential disease-modifying effect was studied using preconditioning with brief seizures (amygdala kindling, maximal electroshocks) and pharmacological strategies including vigabatrin (250 mg/kg), caffeine (0.3 g/L in drinking water), topiramate (10-60 mg/kg), pregabalin (50 mg/kg followed by 10 mg/kg), or RWJ-333369 (10-120 mg/kg). In adult and PN21 rats that became epileptic, entorhinal, and piriform cortices were the initial structures exhibiting significant signal changes on MRI scans, from 6 h after status epilepticus (SE) onset, reflecting neuronal death. In PN21 rats that did not become epileptic, no signal occurred in parahippocampal cortices. In hippocampus, MRI signal change appeared 36-48 h after SE, and progressively worsened to sclerosis. During the latent and chronic phases, the metabolic level in the hilus of adult and PN21 epileptic rats was normal although neuronal loss reached 60-75%. Protection limited to CA1 and/or CA3 (caffeine, topiramate, vigabatrin, amygdala kindling) did not affect the latency to spontaneous seizures. Protection limited to the entorhinal and piriform cortices (pregabalin) delayed epileptogenesis. The combined protection of Ammon's horn and parahippocampal cortices (RWJ-333369) prolonged the latency before the onset of seizures in a dose-dependent manner or, in some cases, prevented the epilepsy. The entorhinal and piriform cortices are critically involved in the early phase of the epileptogenesis while the hilus may initiate and/or maintain epileptic seizures. Pharmacological protection of the basal cortices is necessary for a beneficial disease-modifying effect but this must be combined with protection of the hippocampus to prevent epileptogenesis in this model of TLE.

Topics: Animals; Animals, Newborn; Anticonvulsants; Autoradiography; Cell Count; Cerebral Cortex; Deoxyglucose; Disease Models, Animal; Electroencephalography; Electroshock; Entorhinal Cortex; Epilepsy, Temporal Lobe; Fructose; Glucose; Hippocampus; Kindling, Neurologic; Lithium Chloride; Magnetic Resonance Imaging; Olfactory Pathways; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus; Topiramate; Vigabatrin

Neurochemical studies document involvement of benzodiazepine (BDZ) and mu opioid receptors in seizure development and their possible age-related role during epileptogenesis. To study developmental changes of this role LiCl/pilocarpine status epilepticus (SE) was induced in P12, P25 and/or adult rats. This SE leads to epilepsy in all adult and subpopulation of immature rats. Using in vitro autoradiography, benzodiazepine (BDZ) and mu opioid receptor binding was evaluated 1 week (early phase of epileptogenesis) and 3 months (chronic phase) after SE in 27 brain structures involved in seizure generation and spread (amygdala, hippocampus, basal ganglia and thalamic nuclei). The pattern of receptor binding changes was related to the age at SE, interval after SE and to brain structures. Enhanced BDZ binding was found 1 week after SE in many cortical areas in P12 and also in the amygdala complex and dentate gyrus in both P12 and P25. No changes of BDZ binding occurred in adults at that time, but 3 months after SE a decrease of binding appeared in all evaluated areas in both adult and P25 but not P12 rats. This decrease did not reflect neuronal loss. mu opioid receptors were less significantly affected but clear tendency to decrease binding occurred in adult rats in various cortical, amygdala and thalamic regions early after SE. Changes were less expressed in immature rats. Our data support the hypothesis that age-related changes of receptor properties may participate in different functional consequences of SE including epileptogenesis (more common in older age groups) and behavioral changes.

Topics: Age Factors; Analysis of Variance; Animals; Brain; Disease Models, Animal; Lithium Chloride; Pilocarpine; Rats; Rats, Wistar; Receptors, GABA-A; Receptors, Opioid, mu; Statistics, Nonparametric; Status Epilepticus; Time Factors

To investigate the effects of olomoucine, a cyclin dependent protein kinase (CDK) inhibitor, on the neuronal apoptosis after status epilepticus (SE).. Lithium chloride was injected intraperitoneally, and pilocarpine was injected intraperitoneally after 18 h to 24 SD rats so as to cause SE. Twenty-two of the 24 rats developed SE and 2 of them died. The surviving 20 rats were then randomly divided into 2 equal groups: olomoucine group, injected intracerebroventricularly after the SE was terminated by diazepam and chloral hydrate once a day for 3 days, and SE group, infused intracerebroventricularly with DMSO solution Another 10 rats were injected intraperitoneally with normal saline and then infused intracerebroventricularly with DMSO solution to be used as control group. Six hours after SE attack 5 rats from each group were killed respectively with their brains taken out. Semiquantitative RT-PCR was used to detect the mRNA expression of anti-inflammatory cytokines, such as interleukin (IL)-1beta and tumor necrosis factor (TNF)-alpha. Three days later the other 5 rats in each group were killed with their entorhinal cortex and hippocampus taken out. TUNEL was used to observe the apoptosis. Immunofluorescence (IF) staining was used to detect the expression of neuronal nuclear nucleoprotein (NeuN) and cyclin B1.. TUNEL showed that apoptotic neurons were rare in the control group and were numerous in the SE group, especially in the entorhinal cortex and the hylus of dentate gyrus, and the number of apoptotic neurons in the hylus of dentate gyrus of the olomoucine group was not significantly different from that of the control group (P < 0.05), however, the number of apoptotic cells in the entorhinal cortex of the olomoucine group was still significantly higher than that of the control group (P < 0.05). IF staining demonstrated that in the control group the co-expression of NeuN and TUNEL-labeled cells was weak; and in the SE group the co-expression of NeuN and TUNEL was significantly increased compared with that in the control group (P < 0. 05). The number of cyclin B1 positive cells in the olomoucine group was 18.22 +/- 3.99, significantly lower than that of the SE group (24.57 +/- 6.78, P < 0.05). Semiquantitative RT-PCR showed that the IL-1beta and TNF-alpha mRNA expression levels of the SE group were both significantly higher than those of the control group (both P < 0.05), and the IL-1beta and TNF-alpha mRNA expression levels of the olomoucine group, except the TNF-alpha mRNA expression in the cortex, were all significantly lower than those of the SE group (all P < 0.05), and not significantly different from those of the control group (all P > 0.05).. Olomoucine treatment can inhibits cell cycle protein B1 expression, anti-inflammatory cytokines such as IL-1beta and TNF-alpha secretion, thus decreasing neuronal death and providing neuroprotection after SE, which suggests a potential promising therapeutic way for epilepsy treatment.

Topics: Animals; Apoptosis; Brain; Cyclin B; Cyclin B1; Cyclin-Dependent Kinases; Enzyme Inhibitors; Fluorescent Antibody Technique; In Situ Nick-End Labeling; Interleukin-1beta; Kinetin; Lithium Chloride; Male; Nerve Tissue Proteins; Neurons; Nuclear Proteins; Phosphopyruvate Hydratase; Pilocarpine; Random Allocation; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Status Epilepticus; Tumor Necrosis Factor-alpha

Topics: Animals; Animals, Newborn; Brain; Disease Models, Animal; Hippocampus; Inflammation; Kainic Acid; Lipopolysaccharides; Lithium Chloride; Neurons; Pilocarpine; Rats; Status Epilepticus

Whether status epilepticus (SE) in early infancy, rather than the underlying illness, leads to temporal lobe neurodegeneration and volume reduction remains controversial.. SE was induced with LiCl-pilocarpine in P12 rats. To assess acute neuronal damage, brains (five controls, five with SE) were investigated at 8 h after SE by using silver and Fluoro-Jade B staining. Some brains from the early phase were processed for electron microscopy. To assess chronic changes, brains from nine controls and 13 rats with SE at P12 were analyzed after 3 months by using histology and magnetic resonance imaging (MRI).. MRI analysis of the temporal lobe of adult animals with SE at P12 indicated that 23% of the rats had hippocampal, 15% had amygdaloid, and 31% had perirhinal volume reduction. Histologic analysis of sections from the MR-imaged brains correlated with the MRI data. Analysis of neurodegeneration 8 h after SE by using both silver and Fluoro-Jade B staining revealed degenerating neurons located in the same temporal lobe regions as the volume reduction in chronic samples. Electron microscopic analysis revealed irreversible ultrastructural alterations. As with the chronic histologic and MRI findings, interanimal variability was seen in the distribution and severity of acute damage.. Our data indicate that SE at P12 can cause acute neurodegeneration in the hippocampus as well as in the adjacent temporal lobe. It is likely that acute neuronal death contributes to volume reduction in temporal lobe regions that is detected with MRI in a subpopulation of animals in adulthood.

Topics: Amygdala; Animals; Animals, Newborn; Apoptosis; Disease Models, Animal; Entorhinal Cortex; Fluoresceins; Hippocampus; Lithium Chloride; Magnetic Resonance Imaging; Male; Microscopy, Electron; Nerve Degeneration; Neurons; Organic Chemicals; Pilocarpine; Rats; Rats, Wistar; Silver Staining; Status Epilepticus; Temporal Lobe

Glutamate excitotoxicity has been involved in the pathophysiology of epilepsy. Normal functioning of glutamate transporters clears the synaptically released glutamate to prevent excitotoxic neuronal death. Using densitometric immunohistochemical analysis, we examined the temporal expression of the neuronal glutamate transporter (EAAC1) in the lithium-pilocarpine rat model of temporal lobe epilepsy. During the acute period of lithium-pilocarpine-induced status epilepticus, EAAC1 transporter expression increased in the pyramidal neurons of cornus ammonis (CA)1, CA2 and CA3 (fields of the hippocampus), in dentate gyrus (DG) granule cells and in olfactory tubercle (Tu). During the latent period, EAAC1 expression was strongly expressed in the DG granular and molecular layers, Tu, cerebral cortex and septum, and went back to control levels in CA1, CA2 and CA3 layers. The overexpression of EAAC1 occurred mainly in structures prone to develop Fluoro-Jade-B-positive degenerating neurons. It is, however, not clear to what extent the overexpression of EAAC1 contributes to epileptogenesis and in which area it may represent a preventive or compensatory or response to injury.

Topics: Animals; Brain; Brain Chemistry; Epilepsy, Temporal Lobe; Excitatory Amino Acid Transporter 3; Fluoresceins; Immunohistochemistry; Lithium Chloride; Nerve Degeneration; Organic Chemicals; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

We developed a rat pilocarpine seizure/status epilepticus (SE) model, which closely resembles 1.6-2.0 x LD50 soman exposure, to analyse the molecular mechanism of neuronal damage and to screen effective neuroprotectants against cholinergic agonist and chemical warfare nerve agent (CWNA) exposure. Rats implanted with radiotelemetry probes capable of recording electroencephalogram (EEG), electrocardiogram (ECG), temperature, and physical activity were treated with lithium chloride (5 mEq/kg, im), followed 24 h later by (ip) doses of pilocarpine hydrochloride. Based on radiotelemetry analysis, a dose of 240 mg/kg (ip) pilocarpine generated seizure/SE analogous to 1.6-2.0 x LD50 of soman. The model was refined by reducing the peripheral convulsions without affecting the central nervous system (CNS) by administering methylscopolamine bromide (1 mg/kg, ip), an anti-cholinergic that does not cross the blood-brain barrier. However, when methylscopolamine bromide was administered, a higher dose of pilocarpine (320 mg/kg, ip) was required to generate the equivalent seizure/SE. Histopathology data indicated that pilocarpine induces significant damage to the hippocampal region of the brain, with similar neuropathology to that of 1.6-2.0 x LD50 soman exposure. There was a reduction in body temperature after the administration of pilocarpine, as observed in organophosphate (OP) nerve agents exposure. The heart-rate of pilocarpine-treated animals increased compared to the normal range. The pilocarpine seizure/SE model was also reproducible in the absence of lithium chloride. These results support that pilocarpine seizure/SE model is useful in studying the molecular mechanisms of neuropathology and screening neuroprotectants following cholinergic agonist and CWNA exposure.

Topics: Animals; Anticonvulsants; Behavior, Animal; Body Temperature; Brain; Chemical Warfare Agents; Cholinergic Antagonists; Drug Evaluation, Preclinical; Electroencephalography; Heart Rate; Lethal Dose 50; Lithium Chloride; Male; Models, Animal; Motor Activity; Muscarinic Agonists; N-Methylscopolamine; Neuroprotective Agents; Neurotoxicity Syndromes; Pilocarpine; Rats; Rats, Sprague-Dawley; Seizures; Soman; Status Epilepticus

This work was designed to study the influence of drugs during seizures and status epilepticus (SE) induced by pilocarpine and mortality in adult rats. Morphine (0.1 and 0.2 mg/kg), SCH 23390 (0.1 and 0.2 mg/kg), haloperidol (5 and 10mg/kg) and lithium (30 and 60 mg/kg) were administered intraperitoneally (i.p.), 30 min before to pilocarpine (400 mg/kg, s.c.). The animals were observed (24 h) to determine: number of peripheral cholinergic signs, tremors, stereotyped movements, seizures, SE, latency to first seizure and number of deaths after pilocarpine treatment. Morphine and haloperidol had proconvulsant effects in both doses tested. Smaller and higher doses of these drugs no protected and increased pilocarpine-induced seizures, SE and/or mortality. SCH 23390 protected against seizures, increased the latency to first seizure and reduced the mortality of the animals treated with pilocarpine Theses results suggest that dopamine receptor system receptor subtypes exert opposite functions on the regulation of convulsive activity. The morphine is proconvulsant in lower doses. The opioids in high doses tested exert an action proconvulsant during the establishment of epileptic activity induce by pilocarpine. The lithium no protected the animals against seizures induced by pilocarpine and is used which a model of epilepsy associated with lower doses of pilocarpine in several studies, suggesting absence of the effect anticonvulsants in rodents.

Topics: Analgesics, Opioid; Animals; Antimanic Agents; Dopamine Antagonists; Haloperidol; Lithium Chloride; Male; Morphine; Muscarinic Agonists; Pilocarpine; Rats; Seizures; Status Epilepticus

To study cortical excitability after status epilepticus induced in two age groups of immature rats.. Lithium-pilocarpine status epilepticus was elicited in 12- (SE12) or 25-day-old (SE25) rats. Control siblings received saline instead of pilocarpine. Interhemispheric responses were elicited by stimulation of sensorimotor region of cerebral cortex 3, 6, 9, 13, or 26 days after status. Single biphasic pulses with intensities from 0.2 to 4 mA were used for stimulation; eight responses were always averaged. Amplitude of the first positive and negative waves (i.e., monosynaptic transcallosal responses) was measured and used for construction of input-output (I/O) curves. FluoroJade B was used to visualize degenerating neurons 24 h after status in both age groups.. No significant changes were found at short intervals, but only a tendency to lower amplitudes 3 days after status in SE12 group. Marked changes appeared 26 days after status. The younger group exhibited lower amplitudes than did control rats, whereas SE25 animals generated responses with higher amplitude than did controls (i.e., the I/O curve was steeper. FluoroJade B-positive neurons were scarce in SE12 rats, whereas a substantial number of positive neurons was found in SE25 animals. The positive neurons exhibited characteristics of interneurons, and their distribution in cortical layers differed in the two groups.. Status epilepticus resulted in neuronal death in both SE12 and SE25 animals. Changes in transcallosal evoked potentials were opposite in the two age groups. Augmented amplitude of responses in SE25 rats may indicate an increased cortical excitability.

Topics: Animals; Cerebral Cortex; Corpus Callosum; Electric Stimulation; Fluoresceins; Fluorescent Dyes; Functional Laterality; Interneurons; Lithium Chloride; Male; Nerve Degeneration; Organic Chemicals; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

To test effects of paraldehyde on behavioral outcome of status epilepticus (SE) in developing rats.. Motor SE was induced by LiCl-pilocarpine in rats on postnatal (P) day 12 or 25. Two hours after SE onset, animals were injected with a single dose of paraldehyde (0.07 and 0.3 ml/kg in the P12 group and 0.3 and 0.6 ml/kg in the P25 group). Effects on seizure severity and mortality were evaluated. Growth of animals and their motor abilities were monitored until the adulthood. Three months after SE, cognitive abilities were tested by using the Morris water maze.. Both tested doses of paraldehyde equally affected motor seizures. Convulsions continued until the paraldehyde administration, but then they quickly subsided in all groups. During the subsequent 24 h, occasional clonic seizures occurred in P25 animals treated with the lower dose of paraldehyde. Only hyperactivity and/or automatisms were observed in the other experimental groups. Mortality was not affected by the dosage of paraldehyde. The higher dosage of paraldehyde improved recovery after SE in both age groups. No difference was found in motor abilities between controls and SE animals, except shortening of time spent on the rod in the rotarod test in the P12 group. In P25 rats, treatment with a higher dosage of paraldehyde improved learning abilities compared with the lower dosage. In the P12 group, animals treated with the lower dosage exhibited slightly impaired learning compared with controls and animals receiving the higher dosage.. Paraldehyde injected 2 h after SE onset modulates long-term outcome in immature rats in a dose-related manner.

Topics: Animals; Animals, Newborn; Anticonvulsants; Behavior, Animal; Body Weight; Dose-Response Relationship, Drug; Exploratory Behavior; Lithium Chloride; Male; Maze Learning; Motor Activity; Paraldehyde; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

The piriform cortex is involved in genesis and propagation of temporal lobe seizures. Degenerating neurons demonstrated by FluoroJade B staining are visible early after status epilepticus (SE) as well as after longer intervals. Furthermore, the piriform cortex is activated during an early phase of experimental temporal seizures, as described by magnetic resonance imaging (MRI) studies. It indicates that the early activity of the piriform cortex should be accompanied by increased adenosine triphosphate (ATP) production. Cytochrome oxidase activity in the brain may be used as an endogenous metabolic marker for neurons. The present research studied activity of the cytochrome oxidase separately in the rostral and caudal parts of the piriform cortex after lithium chloride-pilocarpine-induced SE in adult rats.. SE was induced by a single dose of pilocarpine (40 mg/kg) in LiCl-pretreated adult Wistar rats. Cytochrome oxidase activity was mapped by optical density on sections stained with histochemistry separately in the rostral and caudal parts of the piriform cortex.. Optical density of the rostral part of the piriform cortex remained nearly unchanged at both 1 week (0.284 +/- 0.009 in SE group vs. 0.297 +/- 0.005 in controls) and 3 months (0.318 +/- 0.007 in SE group vs. 0.333 +/- 0.004 in controls) after SE intervals. The caudal part of the piriform cortex showed a decrease of optical density in both groups at 1 week (0.265 +/- 0.007 in SE group vs. 0.285 +/- 0.009 in controls) and 3 months after SE (0.292 +/- 0.006 in SE animals vs. 0.310 +/- 0.003 in controls), respectively. Nissl-stained sections demonstrated a marked neuronal loss and gliosis and/or necrotic cavities through the caudal piriform cortex 1 week after SE.. Our results demonstrated that damage of the piriform cortex is not homogeneous and thus that its parts are differently involved in epileptic activity.

Topics: Animals; Brain Mapping; Cerebral Cortex; Electron Transport Complex IV; Lithium Chloride; Male; Neurons; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

To explore the age character of the activity of Caspase 3 and neuron death induced by LiCl-pilocarpine status epilepticus.. LiCl-pilocarpine was injected into healthy infant rats (19 days) and adult rats (2-3 months) subcutaneously and intra-abdominally to evoke status epilepticus (SE). First, the age difference of the seizure was used to measure the sensitivity of seizure. Second, the dynamic features of the apoptotic neurons and the activity of Caspase 3 at 15, 30 min and 1, 2, 4, 8 hours after SE respectively were investigated by TUNEL, flow cytometry and fluorospectrophotometry.. (1) The average duration from the injection to seizure was (13.3 +/- 5.63) min in infant rats, and (22.5 +/- 5.66) min in adult rats. (2) The proportion of the 4th or 5th degree of severity at onset of seizure was 68% in infant rats and 18% in adult rats. (3) Although the count of died neurons (in the CA3 of hippocampus, dentate gyrus and cortex of temporal lobe) was physiologically higher in normal infant rats than in adult rats, the count of positive neurons by TUNEL stain in mature brain (524 +/- 26) remarkably increased and exceeded that in premature brain (465 +/- 26) at 30 min after SE. Although continuously observed until 8 hours after SE, the count of apoptotic neurons in mature brain was also remarkably higher than that in infant brain. Change of neurons in apoptotic early events detected by flow cytometry was the same as the result of TUNEL. (4) The increasing proportion of activity of Caspase 3 after SE for 30 min in adult rats remarkably exceeded that in infant rats; it was 0.10 +/- 0.07 in adult rats and 0.003 +/- 0.04 in infant rats. The difference between the infant rats (0.39 +/- 0.20) and adult rats (0.10 +/- 0.20) increased after SE for 2 hours.. A mechanism of inhibiting apoptotic process in premature brain during SE for the protection against brain damage was well reconfirmed by different animal SE models induced by lithium-pilocarpine. It was indicated that the protective mechanism against brain damage in premature brain could be presented in most severe seizures of different types. This protective mechanism could act on the apoptotic occurrence in the earlier period before the activation of Caspase cascade reaction.

Topics: Age Factors; Animals; Apoptosis; Caspase 3; Caspases; Female; Hippocampus; Lithium Chloride; Male; Neurons; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Prolonged seizures in early childhood are associated with an increased risk of development of epilepsy in later life. The mechanism(s) behind this susceptibility to later development of epilepsy is unclear. Increased synaptic activity during development has been shown to permanently alter excitatory neurotransmission and could be one of the mechanisms involved in this increased susceptibility to the development of epilepsy. In the present study we determine the effect of status-epilepticus induced by lithium/pilocarpine at postnatal day 10 (P10 SE) on the expression of glutamate receptor and transporter mRNAs in hippocampal dentate granule cells and protein levels in dentate gyrus of these animals in adulthood. The results revealed a decrease in glutamate receptor 2 (GluR2) mRNA expression and protein levels as well as an increase in protein levels for the excitatory amino acid carrier 1 (EAAC1) in P10 SE rats compared to controls. Expression of glutamate receptor 1 (GluR1) mRNA was decreased in both P10 SE rats and identically handled, lithium-injected littermate controls compared to naive animals, and GluR1 protein levels were significantly lower in lithium-controls than in naive rats, suggesting an effect of either the handling or the lithium on GluR1 expression. These changes in EAA receptors and transporters were accompanied by an increased susceptibility to kainic acid induced seizures in P10 SE rats compared to controls. The current data suggest that early-life status-epilepticus can result in permanent alterations in glutamate receptor and transporter gene expression, which may contribute to a lower seizure threshold.

Topics: Amino Acid Transport System X-AG; Animals; Cell Separation; Convulsants; Dentate Gyrus; Disease Models, Animal; Disease Susceptibility; Excitatory Amino Acid Transporter 2; Excitatory Amino Acid Transporter 3; Glutamate Plasma Membrane Transport Proteins; Handling, Psychological; Lithium Chloride; Male; Neurons; Pilocarpine; Rats; Receptors, AMPA; Receptors, Glutamate; RNA, Messenger; Seizures; Status Epilepticus; Symporters; Time

Long-term GABA(A) receptor alterations occur in hippocampal dentate granule neurons of rats that develop epilepsy after status epilepticus in adulthood. Hippocampal GABA(A) receptor expression undergoes marked reorganization during the postnatal period, however, and the effects of neonatal status epilepticus on subsequent GABA(A) receptor development are unknown. In the current study, we utilize single cell electrophysiology and antisense mRNA amplification to determine the effect of status-epilepticus induced by lithium-pilocarpine in postnatal day 10 rat pups on GABA(A) receptor subunit expression and function in hippocampal dentate granule neurons. We find that rats subjected to lithium-pilocarpine-induced status epilepticus at postnatal day 10 show long-term GABA(A) receptor changes including a two-fold increase in alpha1 subunit expression (compared with lithium-injected controls) and enhanced type I benzodiazepine augmentation that are opposite of those seen after status epilepticus in adulthood and may serve to enhance dentate gyrus inhibition. Further, unlike adult rats, postnatal day 10 rats subjected to status epilepticus do not become epileptic. These findings suggest age-dependent differences in the effects of status epilepticus on hippocampal GABA(A) receptors that could contribute to the selective resistance of the immature brain to epileptogenesis.

Topics: Age Factors; Animals; Animals, Newborn; Blotting, Northern; Cell Survival; Cerebral Cortex; Dose-Response Relationship, Drug; Drug Interactions; Electroencephalography; GABA Agonists; Gene Expression Regulation; Hippocampus; In Vitro Techniques; Lithium Chloride; Patch-Clamp Techniques; Pilocarpine; Pyridines; Rats; Rats, Sprague-Dawley; Receptors, GABA-A; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Status Epilepticus; Video Recording; Zolpidem

The aims of the present study were to ascertain whether nonconvulsive status epilepticus (NCSE) could give rise to long-term behavioral deficits and permanent brain damage. Two months after NCSE was elicited with pilocarpine (15 mg/kg i.p.) in LiCl-pretreated adult male rats, animals were assigned to either behavioral (spontaneous behavior, social interaction, elevated plus-maze, rotorod, and bar-holding tests) or EEG studies. Another group of animals was sacrificed and their brains were processed for Nissl and Timm staining as well as for parvalbumin and calbindin immunohistochemistry. Behavioral analysis revealed motor deficits (shorter latencies to fall from rotorod as well as from bar) and disturbances in the social behavior of experimental animals (decreased interest in juvenile conspecific). EEGs showed no apparent abnormalities. Quantification of immunohistochemically stained sections revealed decreased amounts of parvalbumin- and calbindin-immunoreactive neurons in the motor cortex and of parvalbumin-positive neurons in the dentate gyrus. Despite relatively inconspicuous manifestations, NCSE may represent a risk for long-term deficits.

Topics: Animals; Arousal; Behavior, Animal; Brain; Brain Damage, Chronic; Brain Mapping; Calbindins; Convulsants; Electroencephalography; Limbic System; Lithium Chloride; Male; Maze Learning; Motor Activity; Motor Skills; Neocortex; Parvalbumins; Pilocarpine; Postural Balance; Rats; Rats, Wistar; Reaction Time; S100 Calcium Binding Protein G; Social Behavior; Status Epilepticus; Video Recording

The neuropeptide galanin exhibits anticonvulsant effects in experimental epilepsy. Two galanin receptor subtypes, GalR1 and GalR2, are present in the brain. We examined the role of GalR1 in seizures by studying the susceptibility of GalR1 knockout (KO) mice to status epilepticus (SE) and accompanying neuronal injury. SE was induced in GalR1 KO and wild type (WT) mice by Li-pilocarpine, 60 min electrical perforant path stimulation (PPS), or systemic kainic acid (KA). Seizures were analyzed using Harmonie software. Cell injury was examined by FluoroJade B- and terminal deoxynucleotidyl transferase-mediated uridine triphosphate nick end labeling; neurogenesis was studied using bromodeoxyuridine labeling. Compared with WT littermates, GalR1 KO showed more severe seizures, more profound injury to the CA1 pyramidal cell layer, as well as injury to hilar interneurons and dentate granule cells upon Li-pilocarpine administration. PPS led to more severe seizures in KO, as compared with WT mice. No difference in the extent of neuronal degeneration was observed between the mice of two genotypes in CA1 pyramidal cell layer; however, in contrast to WT, GalR1 KO developed mild injury to hilar interneurons on the side of PPS. KA-induced seizures did not differ between GalR1 KO and WT animals, and led to no injury to the hippocampus in either of experimental group. No differences were found between KO and WT mice in both basal and seizure-induced neuronal progenitor proliferation in all seizure types. Li-pilocarpine led to more extensive glia proliferation in GalR1 KO than in WT, and in both mouse types in two other SE models. In conclusion, GalR1 mediate galanin protection from seizures and seizure-induced hippocampal injury in Li-pilocarpine and PPS models of limbic SE, but not under conditions of KA-induced seizures. The results justify the development and use of GalR1 agonists in the treatment of certain forms of epilepsy.

Topics: Animals; Dentate Gyrus; Drug Combinations; Electric Stimulation; Hippocampus; Kainic Acid; Lithium Chloride; Mice; Mice, Knockout; Neurons; Perforant Pathway; Pilocarpine; Receptor, Galanin, Type 1; Status Epilepticus; Stem Cells

Status epilepticus (SE) is more common in children than adults and has a high mortality and morbidity rate. SE in adult rats results in long-term disturbances in learning and memory, as well as an enhanced seizure susceptibility to further seizures. In contrast, a number of studies suggest that the immature brain is less vulnerable to the morphologic and physiologic alterations after SE. The goal of this study was to determine whether the long-term consequences of SE during development on hippocampal plasticity and cognitive function are age and model specific.. We used lithium-pilocarpine (Li-PC) to induce SE at different age points during development (P12, P16, P20) and evaluated the effects of this abnormal neural activity on spatial memory performance and seizure susceptibility in the animals beginning at P55, corresponding to young adulthood.. We demonstrated that SE at P12 did not result in any structural or functional changes detectable in adulthood, whereas SE at both P16 and P20 induced cell loss and mossy fiber sprouting within the hippocampus and cognitive impairment when the animals were tested as adults.. Whereas the seizure threshold to generalized seizures was not altered, animals with SE at P20 showed an increased susceptibility to kindling in adulthood.

Topics: Age Factors; Animals; Apoptosis; Convulsants; Electroencephalography; Hippocampus; Lithium Chloride; Long-Term Potentiation; Male; Maze Learning; Mental Recall; Neuronal Plasticity; Orientation; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

Acute caffeine exposure has proconvulsant effects and worsens epileptic and ischemic neuronal damage. Surprisingly, prolonged caffeine exposure decreases the susceptibility to seizures and the extent of ischemic damage. We explored whether the exposure to a low long-term dose of caffeine could protect the brain from neuronal damage and epileptogenesis in the lithium-pilocarpine model of temporal lobe epilepsy.. Rats received either plain tap water or water containing caffeine (0.3 g/L) for 15 days before the induction of status epilepticus (SE) by lithium-pilocarpine and for 7 days after SE. The extent of neuronal damage was assessed in the hippocampus and piriform and entorhinal cortices in brain sections stained with thionine and obtained from animals killed 7 days after SE. The latency to spontaneous recurrent seizures was controlled by video monitoring.. Caffeine treatment induced a marked, almost total neuroprotection in CA1 and a very limited protection in the hilus of the dentate gyrus, whereas damage in layers III-IV of the piriform cortex was slightly worsened by the treatment. All rats, whether they received caffeine or plain tap water, became epileptic after the same latency (17-19 days).. Thus these data extend the neuroprotective effects of low long-term caffeine exposure to epileptic damage and confirm that the sole protection of the Ammon's horn has no influence on the genesis of spontaneous recurrent seizures in this model.

Topics: Age Factors; Animals; Anticonvulsants; Apoptosis; Caffeine; Convulsants; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Schedule; Hippocampus; Kindling, Neurologic; Lithium Chloride; Male; Maze Learning; Mossy Fibers, Hippocampal; Nerve Regeneration; Neuroprotective Agents; Orientation; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

Spontaneous seizures in rats emerge several weeks after induction of status epilepticus with pharmacologic treatment or electrical stimulation, providing an animal model for human temporal lobe epilepsy. In this study, we investigated whether status epilepticus caused changes in the function of voltage-gated sodium channels in entorhinal cortex layer V neurons, a cellular group important for the genesis of limbic seizures.. We induced status epilepticus in rats, by using lithium-pilocarpine, and then 2-12 weeks later, used whole-cell voltage-clamp to examine voltage-activated sodium currents of acutely dissociated layer V neurons.. Transient sodium currents of entorhinal cortex layer V neurons isolated from 9- to 12-week post-status epilepticus rats were similar to currents in age-matched controls; however, low-threshold persistent sodium currents were significantly larger. This increase in persistent activity was not seen 2-3 weeks after pilocarpine treatment; thus it occurred after a delay comparable to the delay in the appearance of spontaneous seizures.. Increased persistent currents are expected to accentuate neuronal excitability and thus may contribute to the genesis of spontaneous seizures after status epilepticus.

Topics: Animals; Cells, Cultured; Disease Models, Animal; Entorhinal Cortex; Epilepsy, Temporal Lobe; Kindling, Neurologic; Lithium Chloride; Male; Membrane Potentials; Neurons; Patch-Clamp Techniques; Pilocarpine; Rats; Rats, Long-Evans; Sodium Channels; Status Epilepticus

Status epilepticus is usually initially treated with a benzodiazepine such as diazepam. During prolonged seizures, however, patients often lose their sensitivity to benzodiazepines, thus developing pharmacoresistant seizures. In rats, administration of LiCl followed 20-24 h later by pilocarpine induces a continuous, self-sustained, and reproducible form of status epilepticus that can be terminated with diazepam when it is administered soon after the pilocarpine injection. However, when administered after a 45 min delay, diazepam is less effective. Previous findings have suggested that the development of pharmacoresistance is related to the stage of status epilepticus. In the present study, we characterized the seizure stage-dependence of diazepam pharmacoresistance. Following administration of different doses of diazepam at varying time intervals after specific behaviorally- and electrographically-defined seizure stages, stage-, time-, and dose-dependent pharmacoresistance to diazepam developed. We also studied two other antiepileptic drugs commonly used in the treatment of status epilepticus, phenobarbital and phenytoin. Consistent with previous studies, our results indicated a similar relationship between stage, time and dose for phenobarbital, but not for phenytoin. Our data are consistent with rapid modulation of GABA(A) receptors during status epilepticus that may result in pharmacoresistance to antiepileptic drugs that enhance GABA(A) receptor-mediated inhibition.

Topics: Animals; Anticonvulsants; Benzodiazepines; Chi-Square Distribution; Diazepam; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Resistance; Electroencephalography; Lithium Chloride; Male; Muscarinic Agonists; Phenobarbital; Phenytoin; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptors, GABA-A; Status Epilepticus; Time Factors

Patients with temporal lobe epilepsy (TLE) usually had an initial precipitating injury in early childhood. However, epilepsy does not develop in all children who have undergone an early insult. As in patients, the consequences of the lithium-pilocarpine-induced status epilepticus (SE) are age dependent, and only a subset of 21-day-old rats will develop epilepsy. Thus with magnetic resonance imaging (MRI), we explored the differences in the evolution of lesions in these two populations of rats.. SE was induced in 21-day-old rats by the injection of lithium and pilocarpine. T2-weighted images and T2 relaxation-time measurements were used for detection of lesions from 6 h to 4 months after SE.. Three populations of rats could be distinguished. The first one had neither MRI anomalies nor modification of the T2 relaxation time, and these rats did not develop epilepsy. In the second one, a hypersignal appeared at the level of the piriform and entorhinal cortices 24 h after SE (increase of 49% of the T2 relaxation time in the piriform cortex) that began to disappear 48-72 h after SE; epilepsy developed in all these animals. The third population of rats showed a more moderate increase of the T2 relaxation time in cortices (14% in the piriform cortex) that could not be seen on T2-weighted images. Epilepsy developed in all these rats. Only in a subpopulation of the 21-day-old rats with epilepsy did hippocampal sclerosis develop.. These results suggest that the injury of the piriform and entorhinal cortices during SE play a critical role for the installation of the epileptic networks and the development of epilepsy.

Topics: Age Factors; Animals; Brain Diseases; Cerebral Cortex; Disease Models, Animal; Entorhinal Cortex; Epilepsy, Temporal Lobe; Female; Lithium Chloride; Magnetic Resonance Imaging; Male; Pilocarpine; Probability; Rats; Rats, Sprague-Dawley; Status Epilepticus

Activation of the tumor suppressor gene, p53, has been strongly implicated in selective neuronal cell death. This study investigated p53 expression in the immature and adult rat brain following status epilepticus induced by the administration of lithium-pilocarpine (LPSE). Both p53 mRNA and protein were examined in relation to neuronal degeneration using in situ hybridization and immunohistochemistry, respectively. Injured cells with eosinophilic cytoplasm with increased p53 mRNA were observed in hippocampal subfields, piriform cortex, amygdala and thalamus. p53 mRNA levels reached a peak by 8 h and returned to baseline by 24 h after the onset of LPSE. The magnitude of p53 mRNA induction was greatest in 21-day-old rats. In contrast to the cellular expression pattern of p53 mRNA, immunohistochemistry demonstrated that p53 protein was increased in all of the eosinophilic cells. Further, double-labeling studies revealed that p53 protein was elevated in neurons that were degenerating. This was supported by colocalization of activated caspase 3 in some cells with damaged DNA. These results provide additional evidence for a critical role for the p53 pathway in excitotoxic neuronal cell death due to status epilepticus.

Topics: Aging; Animals; Animals, Newborn; Antimanic Agents; Brain; Caspase 3; Caspases; Cholinergic Agents; Immunohistochemistry; Lithium Chloride; Nerve Degeneration; Neurons; Pilocarpine; Rats; Rats, Wistar; RNA, Messenger; Status Epilepticus; Tumor Suppressor Protein p53

To characterize electroencephalographic and behavioral effects as well as electrophysiologic and morphologic consequences of a subconvulsive dose of pilocarpine in lithium chloride-pretreated rats.. Pilocarpine (15 mg/kg) was administered intraperitoneally to adult rats pretreated with lithium chloride (3 mEq/kg, i.p.). Behavior was observed for 2 h and videotaped in three consecutive sessions. At the same time, EEG was recorded from the sensorimotor cortex and the dorsal hippocampus. Threshold intensities of currents necessary to elicit hippocampal afterdischarges were determined 24 h and 1 week after the pilocarpine administration. The brains were histologically examined 1 week after pilocarpine administration using Nissl stain.. Pilocarpine induced time-limited nonconvulsive status epilepticus (NCSE). Epileptic EEG activity concurrent with prominent behavioral features was observed both in the neocortex and, predominantly, in the hippocampus. No changes in afterdischarge thresholds were observed in the dorsal hippocampus 24 h and 1 week after NCSE. One week after NCSE, seizure-related brain damage was found mainly in the motor neocortical fields.. Pilocarpine-induced NCSE in rats strongly resembles a short-term human complex partial status epilepticus. Our animal model is suitable for studying the possible adverse effects of prolonged nonconvulsive seizures.

Topics: Animals; Behavior, Animal; Disease Models, Animal; Electroencephalography; Hippocampus; Humans; Lithium Chloride; Male; Motor Activity; Neocortex; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Videotape Recording

Status epilepticus (StE) in immature rats causes long-term functional impairment. Whether this is associated with structural alterations remains controversial. The present study was designed to test the hypothesis that StE at an early age results in neuronal loss. StE was induced with lithium-pilocarpine in 12-d-old rats, and the presence of neuronal damage was investigated in the brain from 12 hr up to 1 week later using silver and Fluoro-Jade B staining techniques. Analysis of the sections indicated consistent neuronal damage in the central and lateral segments of the mediodorsal nucleus of the thalamus, which was confirmed using adjacent cresyl violet-stained preparations. The mechanism of thalamic damage (necrosis vs apoptosis) was investigated further using TUNEL, immunohistochemistry for caspase-3 and cytochrome c, and electron microscopy. Activated microglia were detected using OX-42 immunohistochemistry. The presence of silver and Fluoro-Jade B-positive degenerating neurons in the mediodorsal thalamic nucleus was associated with the appearance of OX-42-immunopositive activated microglia but not with the expression of markers of programmed cell death, caspase-3, or cytochrome c. Electron microscopy revealed necrosis of the ultrastructure of damaged neurons, providing further evidence that the mechanism of StE-induced damage in the mediodorsal thalamic nucleus at postnatal day 12 is necrosis rather than apoptosis. Finally, these data together with previously described functions of the medial and lateral segments of the mediodorsal thalamic nucleus suggest that some functions, such as adaptation to novelty, might become compromised after StE early in development.

Topics: Animals; Antigens, CD; Antigens, Neoplasm; Antigens, Surface; Apoptosis; Avian Proteins; Basigin; Blood Proteins; Caspase 3; Caspases; Cytochrome c Group; Disease Models, Animal; Disease Progression; Immunohistochemistry; In Situ Nick-End Labeling; Lithium Chloride; Male; Mediodorsal Thalamic Nucleus; Membrane Glycoproteins; Microglia; Microscopy, Electron; Necrosis; Neurons; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus

Substance P, which modulates synaptic excitability, can be induced by a variety of stimuli. We studied the expression of hippocampal substance P in rats in using lithium-pilocarpine model of status epilepticus during development. Status epilepticus resulted in an age-specific manner of substance P expression that was anatomically distinctive in hippocampal subfields. Maximal induction of substance P immunoreactivity was seen in the CA1 region of the two-week-old rats, and progressively decreased in the three-, four-week-old rats and adults. Meanwhile, the number of substance P-immunoreactive neurons in the CA3 region and dentate granule cell layer was minimal in the two-week-old animals, but approximated the adult level in the three- and four-week-old rats. No substance P-immunoreactive axon terminals were seen in the strata pyramidale and lucidum in the CA3 region of the two-week-old rats, but they were found to progressively increase in the three-, four-week-old rats and adults. To confirm substance P expression after status epilepticus, we studied the expression of preprotachykinin-A mRNA in the hippocampus of the three-week-old rats by in situ hybridization. Two hours following injection of lithium-pilocarpine, preprotachykinin-A mRNA dramatically increased in the granule cells, as well as in the CA3 and CA1 pyramidal cell layers of the hippocampus. To evaluate the relationship between behavioral seizures and substance P induction, we used the NMDA receptor antagonist MK-801. Injection of MK-801 completely blocked lithium-pilocarpine-induced behavioral seizures and SP induction in the two-week-old rats. These results indicate that seizure activity selectively evokes age-dependent and region-selective expression of substance P.

Topics: Age Factors; Animals; Dizocilpine Maleate; Female; Hippocampus; Lithium Chloride; Male; Neural Pathways; Neurons; Pilocarpine; Protein Precursors; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Substance P; Tachykinins

We studied the effects of status epilepticus (SE) induced by lithium chloride/pilocarpine treatment on gene expression of neurotrophins of the nerve growth factor (NGF) family and of their high-affinity receptors of the tyrosine protein kinase (trk) family in the forebrain. Using in situ hybridization (ISH), we demonstrated an early (3 h after treatment) increase in brain-derived neurotrophic factor (BDNF) and trkB mRNA expression in the dentate gyrus, amygdala, and piriform cortex, as well as widespread increases in the cerebral cortex. NGF mRNA, but not the mRNA of its receptor trkA, was increased in the dentate gyrus. In contrast, 12 h after treatment, neurotrophin-3 (NT-3) decreased, and its receptor trkC mRNA increased. There was no change in NT-4 mRNA levels. All changes were blocked by pretreatment with scopolamine, a muscarinic antagonist. The noncompetitive N-methyl-D-aspartate (NMDA) antagonist ketamine blocked NGF, BDNF, and trkB mRNA increases in the hippocampus and cerebral cortex, but not in the amygdala and piriform cortex. In contrast, ketamine did not affect NT-3 and trkC changes. These results provide a complete description of changes in mRNA levels of neurotrophins and their receptors in the forebrain after SE and supply additional data supporting the view that neurotrophin gene expression is related to abnormal neuronal activity.

Topics: Animals; Brain-Derived Neurotrophic Factor; Cerebral Cortex; Gene Expression; Hippocampus; In Situ Hybridization; Ketamine; Lithium Chloride; Male; Nerve Growth Factors; Nerve Tissue Proteins; Neurotrophin 3; Pilocarpine; Prosencephalon; Protein-Tyrosine Kinases; Rats; Rats, Wistar; RNA, Messenger; Scopolamine; Status Epilepticus; Tissue Distribution

Administration of subconvulsive dose of pilocarpine (30 mg/kg s.c.) to rats pretreated with lithium chloride (3 meq/kg i.p.) produced a state of status epilepticus in animals. The animals showed characteristic symptoms of generalized convulsions, wet dog shakes (WDS), forelimb clonus and falling back. The symptoms of status epilepticus (SE) developed within 26.8 +/- 3.6 min after administering pilocarpine and these symptoms continued uninterrupted. The phenomenon was totally reproducible, with a consistent latency of onset of seizures and a high mortality rate. The symptoms were blocked by atropine, scopolamine and the GABAergic agents GABA, sodium valproate, (+)-baclofen and clonazepam when given prior to pilocarpine, but not when administered 30 min after pilocarpine administration.

Topics: Animals; Anticonvulsants; Baclofen; Clonazepam; Diazepam; Dizocilpine Maleate; Ethanol; GABA Agents; gamma-Aminobutyric Acid; Lithium Chloride; Male; Neuroprotective Agents; Pentobarbital; Phenobarbital; Phenytoin; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Valproic Acid

Status epilepticus was induced in rats by sequential injections of lithium and pilocarpine. Seizure activity was aborted by a combination of MK-801 and diazepam, with status duration ranging from 3 to 180 min. When the hearts were examined 8-12 days later, rats that had experienced an episode of status epilepticus had significantly heavier hearts than did controls. The nature of the cardiac tissue changes was not examined, and deserves further study.

Topics: Animals; Body Weight; Cardiomegaly; Diazepam; Dizocilpine Maleate; Electroencephalography; Lithium Chloride; Male; Myocardium; Organ Size; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

We investigated the neuroprotective effect of the noncompetitive N-methyl-D-asparatate (NMDA) antagonist ketamine when administered after onset of lithium-pilocarpine-induced status epilepticus (SE). Seizures were induced in Wistar rats with lithium chloride (3 mEq/kg) and pilocarpine (PC) (30-60 mg/kg intraperitoneally, i.p.). Fifteen minutes after SE onset, either ketamine 100 mg/kg or normal saline was injected i.p., and 3 h after SE onset atropine, diazepam (DZP), and phenobarbital (PB) were administered i.p. to terminate the seizures. Twenty-four hours later, rats underwent brain perfusion-fixation, with subsequent brain processing for light-microscopic examination. Rats adminstered saline (n = 5) had neuronal damage in 24 of 25 brain regions examined. Rats administered ketamine (n = 7) had significant neuroprotection in 22 of 24 damaged regions. Ketamine reduced the amplitude of seizure discharges, and in 3 rats EEG seizure activity ceased in 30 min; none of these rats had neuronal damage. In the other 4 rats, EEG seizure discharges persisted > 90 min; in these animals, 21 of 24 damaged regions were protected. In contrast, rats with 1-h high-dose PC-induced SE (400 mg/kg i.p. without lithium chloride preadministration) had 14 damaged regions, of which 7 were significantly different from the undamaged regions of the ketamine subgroup with persistent electrographic seizures. Thus, ketamine is remarkably neuroprotective when administered after onset of SE, whether or not seizure discharges are eliminated.

Topics: Amygdala; Animals; Brain; Cell Death; Electroencephalography; Frontal Lobe; Injections, Intraperitoneal; Ketamine; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Status Epilepticus

In the present investigation we address the question of whether one of the responses to increased neuronal activity is a modification of the expression of the different subunits of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA)-selective glutamate receptors (GluR-1, GluR-2, GluR-3). Thus, we used two different models of generalized status epilepticus, as widespread elevated neuronal activity, to study in vivo responses of the AMPA receptor mRNA expression in rat forebrain. By Northern blot analysis and in situ hybridization, we show that one of the delayed responses to LiCl/pilocarpine-induced status epilepticus is a dramatic change in the mRNA level of some subunits of AMPA-selective glutamate receptors. These effects, which appear between 6 and 12 h after the drug treatment, are subunit and brain region specific. The most striking example of differential expression of the three examined GluR mRNAs can be observed in the dentate gyrus of the hippocampus. In this specific brain subregion an increase of GluR-3 mRNA level is induced 12 h after LiCl/pilocarpine treatment, while a clear decrease in GluR-1 mRNA level and no significant change in GluR-2 mRNA level can be observed in the same area under these experimental conditions. Both the GluR-1 decrease and the GluR-3 increase are transient effects and a return to basal level can be observed after 48-72 h. In the CA1 layer of the hippocampus, a parallel decrease of both GluR-1 and GluR-3 expression is found 12-24 h after drug treatment, followed by a recovery of the expression to control values at 48 h. In kainate-induced epilepsy we could reproduce the late increase (12-24 h) in GluR-3 mRNA in the dentate gyrus; however, under this experimental condition, no clear decrease of GluR-1 expression can be observed in this area. A general decrease in mRNA level for the AMPA receptor subunits (GluR-1-3) in the hippocampal layers, in particular in CA3 and CA4 subfields, was also observed. In conclusion the results reported in the present paper reveal a specific regulation of GluR gene expression in the granule cells of the hippocampal dentate gyrus and stimulate further investigation on the functional role of the GluR-3 subunit in the receptor-channel complex.

Topics: alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Base Sequence; Blotting, Northern; Brain; Gene Expression; In Situ Hybridization; Injections, Intraventricular; Kainic Acid; Lithium Chloride; Male; Molecular Sequence Data; Pilocarpine; Rats; Rats, Wistar; Receptors, AMPA; Status Epilepticus

The effects of two protein synthesis inhibitors, cycloheximide and anisomycin, were tested on seizures induced by coadministration of lithium and pilocarpine to rats. Systemic cycloheximide (2 mg/kg, s.c.) and centrally administered anisomycin (300 micrograms/10 microliters, i.c.v.) doubled the latency to initiation of seizures and to status epilepticus, while peripherally administered anisomycin (50 mg/kg, s.c.) completely blocked lithium-pilocarpine seizures. These results indicate that protein synthesis is required for initiation of seizures.

Topics: Animals; Anisomycin; Cerebral Ventricles; Cycloheximide; Electroencephalography; Hippocampus; Injections, Intraventricular; Injections, Subcutaneous; Lithium Chloride; Male; Pilocarpine; Protein Synthesis Inhibitors; Rats; Rats, Sprague-Dawley; Seizures; Status Epilepticus; Time Factors

Core temperature was measured in rats 24 hr. after they had been assigned to one of 8 groups in a 3-way analysis of variance design that involved (1) induction of limbic seizures by a systemic injection of lithium/pilocarpine, (2) physical restraint, and (3) administration of acepromazine. An extraordinarily powerful interaction was noted among seizures, physical restraint, and acepromazine-produced hypothermia (24 degrees C) compared to the other 7 treatments (> 35 degrees C). The putative poikilothermic response is commensurate with the loss of mammalian behaviors that follow these seizures. Implications for survival during the acute stages of brain injury are suggested.

Topics: Acepromazine; Animals; Body Temperature Regulation; Brain Injuries; Chlorides; Hypothermia; Limbic System; Lithium; Lithium Chloride; Male; Neurons; Pilocarpine; Rats; Restraint, Physical; Seizures; Status Epilepticus

Lithium is known to potentiate the ability of pilocarpine to induce status epilepticus in rats. The goal of this study was to determine whether lithium could potentiate pilocarpine-induced seizures in developing animals. Behavioral, electroencephalographic (EEG), and histopathological changes induced by systemic administration of lithium (3 meq/kg) followed 20 h later by pilocarpine (3, 10, 30, 60 mg/kg) were studied in 3-30-day-old rats. Lithium followed by pilocarpine (30 and 60 mg/kg) induced hyperactivity, tremor, loss of postural control and scratching but no electrographic seizures in 3-8-day-old rats. In the 7-10-day-old animals pretreatment with lithium and pilocarpine 60 mg/kg induced status epilepticus with sustained myoclonus and continuous bilateral synchronous spike and sharp wave, but doses of pilocarpine lower than 60 mg/kg had no effect. The susceptibility to lithium-pilocarpine-induced status epilepticus increased markedly during the third postnatal week of life. During this time period, rats treated with lithium (3 meq/kg) plus pilocarpine 10 mg/kg exhibited behavioral and EEG manifestations of status epilepticus. The same combination of lithium and pilocarpine failed to induce status epilepticus either before or after the third week of life. Histopathological analysis of the brains of the animals used in these studies failed to demonstrate the widespread damage reported in adult rats that have undergone lithium-pilocarpine-induced status epilepticus.

Topics: Aging; Animals; Animals, Newborn; Cerebral Cortex; Chlorides; Drug Administration Schedule; Electroencephalography; Hippocampus; Lithium; Lithium Chloride; Motor Activity; Pilocarpine; Rats; Rats, Inbred Strains; Reference Values; Status Epilepticus; Stereotyped Behavior; Time Factors

The specific binding of [3H]hemicholinium-3 ([3H]HCh-3) and high-affinity [3H]choline uptake were measured in rats with status epilepticus induced by lithium and pilocarpine. The specific binding of [3H]HCh-3 in cortex and hippocampus from rats with status epilepticus increased to 2- to 3-fold of control while the striatal [3H]HCh-3 binding increased minimally. Scatchard analyses revealed that the observed changes resulted from an increase in Bmax of [3H]HCh-3 binding. High-affinity [3H]choline uptake remained unchanged. These results further implicate phospholipase A2 in the regulation of [3H]HCh-3 binding sites.

Topics: Acetylcholine; Animals; Cerebral Cortex; Chlorides; Choline; Hemicholinium 3; Hippocampus; Kinetics; Lithium; Lithium Chloride; Male; Pilocarpine; Rats; Rats, Inbred Strains; Seizures; Status Epilepticus

Electroencephalographic techniques were used to study generalized convulsive status epilepticus induced by administration of subconvulsive doses of cholinomimetics (e.g., pilocarpine) to rats pretreated with lithium chloride. Status epilepticus induced by this treatment was compared with status epilepticus induced by kainic acid. Lithium/pilocarpine-induced status epilepticus developed within 10 min of initial paroxysmal spike activity, 24 +/- 1 min (N = 20) after administration of pilocarpine, and continued uninterrupted for more than 3 h. Kainic acid (10 mg/kg)-induced status epilepticus developed approximately 60 min after initial spike activity, 96 +/- 3 min (N = 7) after kainate administration, and continued for 0.5 h. Thus, the interval of intermittent seizure activity and the duration of status epilepticus differed markedly between these two models. The potentiation by lithium (3 meq/kg) of the convulsant effect of cholinergic agonists was found to be 10 to 13-fold for two direct-acting cholinomimetics, pilocarpine and arecoline, whereas the convulsant effect of the indirect-acting agonist, physostigmine, was potentiated by 50%. The full proconvulsant effect of lithium lasted from 2 to 24 h after a single acute treatment (3 meq/kg). The dose response of the proconvulsant effect of lithium was determined and the EC50 of lithium was approximately 1.5 meq/kg when pilocarpine (30 mg/kg) was administered 20 h later. Chronic treatment with lithium for 4 weeks potentiated the convulsant effect of pilocarpine by more than 26-fold. These results demonstrated that both acute and chronic administration of lithium enhance cholinergic function in vivo. Potentiation of cholinergic function by lithium may play a role in the therapeutic action of lithium in affective disorders.

Topics: Animals; Arecoline; Chlorides; Dose-Response Relationship, Drug; Drug Synergism; Electroencephalography; Kainic Acid; Lithium; Lithium Chloride; Male; Physostigmine; Pilocarpine; Rats; Rats, Inbred Strains; Status Epilepticus