dizocilpine-maleate and Status-Epilepticus

Organophosphorus nerve agents (NA), potent irreversible cholinesterase inhibitors, could induce severe seizures, status epilepticus (SE), seizure-related brain damage (SRBD) and lethality. Despite the lack of data in the case of NA, clinical evidences suggest that SE survivors could suffer from neurological/cognitive deficits and impairments such as spontaneous recurrent seizures (epilepsy) after a latent period of epileptogenesis. It is beyond doubt that an effective and quick management of the initial seizures and prevention of SRBD are critical to prevent these long-term consequences, explaining why most experimental data are focusing on the 5-40min post-exposure time frame. However, in field conditions, treatment may be delayed and with the exception of NMDA receptor antagonists, currently no drug provides protection (against lethality, seizures, SRBD and neurological consequences) when seizures are left unabated for one hour or more. Ketamine (KET) is the only NMDA antagonist licensed as an injectable drug in different countries and remains an anesthetic of choice in some difficult field conditions. In this short review paper, after a presentation of some of the key points of the pathophysiology of NA-induced SE and a quick survey of the potential therapeutic avenues in the context of delayed treatment of NA-induced SE, we will review the recent data we obtained showing that KET, in combination with atropine sulfate (AS), with or without a benzodiazepine, considerably reduces soman-induced neuroinflammation, provides neuroprotection, histologically and functionally, and also positively modify soman-induced changes in brain metabolism. Finally, we will also mention some results from safety studies including those bringing evidence that, at difference with MK-801, KET does not impair thermoregulation and even seems to reduce AS-induced heat stress. All in all, KET, in combination, appears a good candidate for the out-of-hospital treatment of severe NA-induced SE.

Topics: Animals; Brain; Chemical Warfare Agents; Cholinesterase Inhibitors; Dizocilpine Maleate; Guinea Pigs; Humans; Ketamine; Neuroprotective Agents; Receptors, N-Methyl-D-Aspartate; Soman; Status Epilepticus; Time Factors

The anticonvulsant and antioxidant effects of lamotrigine on status epilepticus (SE) are incompletely understood. We assessed these effects of lamotrigine on pilocarpine (Pilo)-induced SE in mice.. Male C57BL/J6 mice were assigned to three groups: the control group, Pilo (400 mg/kg, s.c.)-induced SE (Pilo group) and lamotrigine (20 mg/kg, i.p.) treated (Pilo/lamotrigine group). The latency to SE of Racine's stage 3 or higher, the mortality rate within 2 h of Pilo administration, and the duration of SE until sacrifice were examined. Nitric oxide (NO), malondialdehyde and glutathione of oxidative stress biomarkers were detected in the hippocampus of the sacrificed animals in the above groups. NO was also detected in the cultured rat hippocampal neurons treated with 4 μM Pilo, Pilo+100 μM lamotrigine (Pilo/lamotrigine) and Pilo/lamotrigine+ N-methyl-D-aspartic acid (NMDA) receptor antagonist (10 μM MK-801, 3 μM ifenprodil) to examine the antioxidant effects of lamotrigine via non-NMDA-related pathways.. lamotrigine prolonged the latency to SE, the SE duration until sacrifice, and decreased the mortality rate in mice with Pilo-induced SE. Lamotrigine also decreased hippocampal concentrations of NO and malondialdehyde and increased the concentrations of glutathione in the SE model. Furthermore, there were significant differences in NO concentrations between groups of cultured rat hippocampal neurons treated with Pilo and Pilo/lamotrigine, and with Pilo/lamotrigine and Pilo/lamotrigine+MK-801.. Our findings suggest that lamotrigine exerts anticonvulsant and antioxidant effects on SE, but its antioxidant activity may not be fully exerted via NMDA-related pathways.

Topics: Animals; Anticonvulsants; Antioxidants; Dizocilpine Maleate; Glutathione; Hippocampus; Lamotrigine; Male; Mice; Mice, Inbred C57BL; Pilocarpine; Rats; Status Epilepticus

We determined the role of the neurosteroid-sensitive δ subunit-containing γ-aminobutyric acid A receptors (δ-GABARs) in epileptogenesis.. Status epilepticus (SE) was induced via lithium pilocarpine in adult rats, and seizures were assessed by continuous video-electroencephalography (EEG) monitoring. Finasteride was administered to inhibit neurosteroid synthesis. The total and surface protein expression of hippocampal δ, α4, and γ2 GABAR subunits was studied using biotinylation assays and Western blotting. Neurosteroid potentiation of the tonic currents of dentate granule cells (DGCs) was measured by whole-cell patch-clamp technique. Finally, the effects of inhibiting N-methyl-d-aspartate receptors (NMDARs) during SE on the long-term plasticity of δ-GABARs, neurosteroid-induced modulation of tonic current, and epileptogenesis were studied.. The inhibition of neurosteroid synthesis 4 days after SE triggered acute seizures and accelerated the onset of chronic recurrent spontaneous seizures (epilepsy). The down-regulation of neurosteroid-sensitive δ-GABARs occurred prior to the onset of epilepsy, whereas an increased expression of the γ2-GABAR subunits occurred after seizure onset. MK801 blockade of NMDARs during SE preserved the expression of neurosteroid-sensitive δ-GABARs. NMDAR blockade during SE also prevented the onset of spontaneous seizures.. Changes in neurosteroid-sensitive δ-GABAR expression correlated temporally with epileptogenesis. These findings raise the possibility that δ-GABAR plasticity may play a role in epileptogenesis.

Topics: Animals; Blotting, Western; Dentate Gyrus; Disease Models, Animal; Dizocilpine Maleate; Down-Regulation; Electroencephalography; Epilepsy, Temporal Lobe; Female; Finasteride; Hippocampus; Lithium Compounds; Male; Neuronal Plasticity; Neurons; Neurotransmitter Agents; Patch-Clamp Techniques; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptors, GABA-A; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Video Recording

The synthetic rodenticide, tetramethylenedisulfotetramine (TMDT), is a persistent and highly lethal GABA-gated Cl(-) channel blocker. TMDT is clandestinely produced, remains popular in mainland China, and causes numerous unintentional and deliberate poisonings worldwide. TMDT is odorless, tasteless, and easy to manufacture, features that make it a potential weapon of terrorism. There is no effective treatment. We previously characterized the effects of TMDT in C57BL/6 mice and surveyed efficacies of GABAergic and glutamatergic anticonvulsant treatments. At 0.4 mg/kg i.p., TMDT produced neurotoxic symptomatology consisting of twitches, clonic and tonic-clonic seizures, often progressing to status epilepticus and death. If administered immediately after the occurrence of the first clonic seizure, the benzodiazepine diazepam (DZP) effectively prevented all subsequent seizure symptoms, whereas the NMDA receptor antagonist dizocilpine (MK-801) primarily prevented tonic-clonic seizures. The latter agent, however, appeared to be more effective at preventing delayed death. The present study further explored these phenomena, and characterized the therapeutic actions of DZP and MK-801 as combinations. Joint treatment with both DZP and MK-801 displayed synergistic protection against tonic-clonic seizures and 24 h lethality as determined by isobolographic analysis. Clonic seizures, however, remained poorly controlled. A modification of the treatment regimen, where DZP was followed 10 min later by MK-801, yielded a reduction in both types of seizures and improved overall outcome. Simultaneous monitoring of subjects via EEG and videography confirmed effectiveness of this sequential regimen. We conclude that TMDT blockage at GABAA receptors involves early activation of NMDA receptors, which contribute to persistent ictogenic activity. Our data predict that a sequential combination treatment with DZP followed by MK-801 will be superior to either individual therapy with, or simultaneous administration of, these two agents in treating TMDT poisoning.

Topics: Animals; Anticonvulsants; Brain Waves; Bridged-Ring Compounds; Central Nervous System; Diazepam; Disease Progression; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Drug Therapy, Combination; Electroencephalography; Epilepsy, Tonic-Clonic; Excitatory Amino Acid Antagonists; GABA Antagonists; gamma-Aminobutyric Acid; Glutamic Acid; Male; Mice, Inbred C57BL; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Time Factors; Video Recording

Identification of new molecular targets as well as the new models recapitulating different aspects of pathophysiology of status epilepticus (SE) in humans might prove essential for the breakthrough in the efforts against pharmacoresistance in epilepsy. Recently, we described a new model of generalized convulsive SE induced with orphenadrine (ORPH) in rats with unique characteristics [5]. The current study was aimed at assessing the efficacy of a new generation antiepileptic drugs (AEDs) and some of the experimental agents in suppressing ORPH-evoked seizures in rats.. ORPH was administered intraperitoneally (ip) in the dose of 80 mg/kg in male Wistar rats. The latency to first seizure, the number of seizure episodes and the duration of overt SE, as well as the incidence of deaths was scored with simultaneous electroencephalographic (EEG) recordings.. ORPH induced seizures in 100% of animals at a dose of 80 mg/kg, associated with low mortality and good behavioural outcome. Among new generation AEDs: felbamate, levetiracetam, topiramate, lamotrigine and progabide did not affect the seizure incidence. Among the experimental drugs, only dizocilpine, the non-competitive NMDA antagonist, dose-dependently affected the occurrence of the SE (p<0.001). However, CGP-39551 competitive NMDA antagonist, the same as scopolamine and mecamylamine (muscarinic and nicotinic receptors antagonists, respectively) showed no effect.. Based on the above findings, one may speculate that NMDA activation is partly involved in the proconvulsant activity of orphenadrine but may not be the primary pathomechanism. ORPH-induced seizures may provide an interesting option for studying novel targets for pharmacological interventions in status epilepticus.

Topics: 2-Amino-5-phosphonovalerate; Animals; Anticonvulsants; Dizocilpine Maleate; Electroencephalography; Hippocampus; Male; N-Methylaspartate; Orphenadrine; Rats; Rats, Wistar; Seizures; Status Epilepticus

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

Organophosphate (OP) compounds are among the most lethal chemical weapons ever developed and are irreversible inhibitors of acetylcholinesterase. Exposure to majority of OP produces status epilepticus (SE) and severe cholinergic symptoms that if left untreated are fatal. Survivors of OP intoxication often suffer from irreversible brain damage and chronic neurological disorders. Although pilocarpine has been used to model SE following OP exposure, there is a need to establish a SE model that uses an OP compound in order to realistically mimic both acute and long-term effects of nerve agent intoxication. Here we describe the development of a rat model of OP-induced SE using diisopropylfluorophosphate (DFP). The mortality, behavioral manifestations, and electroencephalogram (EEG) profile for DFP-induced SE (4 mg/kg, sc) were identical to those reported for nerve agents. However, significantly higher survival rates were achieved with an improved dose regimen of DFP and treatment with pralidoxime chloride (25 mg/kg, im), atropine (2 mg/kg, ip), and diazepam (5 mg/kg, ip) making this model ideal to study chronic effects of OP exposure. Further, DFP treatment produced N-methyl-D-aspartate (NMDA) receptor-mediated significant elevation in hippocampal neuronal [Ca(2+)](i) that lasted for weeks after the initial SE. These results provided direct evidence that DFP-induced SE altered Ca(2+) dynamics that could underlie some of the long-term plasticity changes associated with OP toxicity. This model is ideally suited to test effective countermeasures for OP exposure and study molecular mechanisms underlying neurological disorders following OP intoxication.

Topics: Animals; Calcium; Dizocilpine Maleate; Electroencephalography; Hippocampus; Isoflurophate; Male; Rats; Rats, Sprague-Dawley; Status Epilepticus

Status epilepticus is a clinical emergency that can lead to the development of acquired epilepsy following neuronal injury. Understanding the pathophysiological changes that occur between the injury itself and the expression of epilepsy is important in the development of new therapeutics to prevent epileptogenesis. Currently, no anti-epileptogenic agents exist; thus, the ability to treat an individual immediately after status epilepticus to prevent the ultimate development of epilepsy remains an important clinical challenge. In the Sprague-Dawley rat pilocarpine model of status epilepticus-induced acquired epilepsy, intracellular calcium has been shown to increase in hippocampal neurons during status epilepticus and remain elevated well past the duration of the injury in those animals that develop epilepsy. This study aimed to determine if such changes in calcium dynamics exist in the hippocampal culture model of status epilepticus-induced acquired epilepsy and, if so, to study whether manipulating the calcium plateau after status epilepticus would prevent epileptogenesis. The in vitro status epilepticus model resembled the in vivo model in terms of elevations in neuronal calcium concentrations that were maintained well past the duration of the injury. When used following in vitro status epilepticus, dantrolene, a ryanodine receptor inhibitor, but not the N-methyl-D-aspartic acid channel blocker MK-801 inhibited the elevations in intracellular calcium, decreased neuronal death and prevented the expression of spontaneous recurrent epileptiform discharges, the in vitro correlate of epilepsy. These findings offer potential for a novel treatment to prevent the development of epileptiform discharges following brain injuries.

Topics: Animals; Calcium; Cell Death; Cells, Cultured; Dantrolene; Dizocilpine Maleate; Epilepsy; Excitatory Amino Acid Antagonists; Hippocampus; Muscarinic Agonists; Muscle Relaxants, Central; Neurons; Neuroprotective Agents; Patch-Clamp Techniques; Pilocarpine; Rats; Rats, Sprague-Dawley; Status Epilepticus

Early-life stress has been shown to destabilize the homeostatic synaptic plasticity and compromise the developing brain to the later encountered insults. This study would determine the long-term epileptogenic effect of neonatal isolation (NI) on early-life seizure. There were five groups: normal rearing (NR) rats; NI rats; NR rats suffering from status epilepticus (SE) at P12 (NR-SE); NI-SE rats; NI-SE-MK801 rats. All adult rats were video monitored to detect behavioral seizures, examined with brain magnetic resonance imaging, and assessed for hippocampal NeuN-immunoreactive (NeuN-IR) cells. Behavioral seizures were detected in one of six NR-SE rats, all the NI-SE rats (eight of eight), and none in the NR, NI, or NI-SE-MK801 rats. High hippocampal T2 signal were only found in three of five NR-SE rats, five of six NI-SE rats, and one of five NI-SE-MK801 rats. There was a significant decrease in the number of hippocampal NeuN-IR cells in the NR-SE and NI-SE groups, compared with the NR group, and MK-801 treatment ameliorated the neuronal loss. Our results demonstrated that NI led to an increase in epileptogenesis in rat pups with early-life SE, and treatment with MK-801 could ameliorate brain injuries, indicating a critical role of N-methyl-d-aspartic acid receptor in the epileptogenic process.

Topics: Animals; Animals, Newborn; Behavior, Animal; Dizocilpine Maleate; Female; Hippocampus; Humans; Magnetic Resonance Imaging; Male; Neuroprotective Agents; Pregnancy; Rats; Rats, Sprague-Dawley; Seizures; Social Isolation; Status Epilepticus

About 30% of patients with epilepsy do not respond adequately to drug therapy, making pharmacoresistance a major problem in the treatment of this common brain disorder. Mechanisms of intractability are not well understood, but may include limitation of antiepileptic drug access to the seizure focus by overexpression of the drug efflux transporter P-glycoprotein (Pgp) at the blood-brain barrier. Increased expression of Pgp has been determined both in epileptogenic brain tissue of patients with intractable epilepsy and in rodent models of temporal lobe epilepsy, including the pilocarpine model. The mechanisms underlying the increase of Pgp after seizures are unclear. We have recently suggested that the excitatory neurotransmitter glutamate, which is excessively released by seizures, is involved in the seizure-induced overexpression of Pgp in the brain. This hypothesis was evaluated in the present study in the pilocarpine model in rats. After 90 min of status epilepticus (SE), diazepam was administered, followed by either vehicle or the glutamate receptor antagonist MK-801 (dizocilpine). Following SE in vehicle treated rats, Pgp expression in brain capillary endothelial cells increased about twofold in the hippocampus, which was completely prevented by MK-801. Furthermore, neurodegeneration developing in the hippocampus and parahippocampal regions was reduced by the glutamate antagonist. In contrast, the Pgp inhibitor tariquidar did not affect the SE-induced overexpression of Pgp or neurodegeneration in most regions examined. The data indicate that seizure-induced glutamate release is involved in the regulation of Pgp expression, which can be blocked by MK-801. The finding that MK-801 counteracts both Pgp overexpression and neuronal damage when administered after SE may offer a clinically useful therapeutic option in patients with refractory SE.

Topics: Animals; Apoptosis; ATP Binding Cassette Transporter, Subfamily B, Member 1; Brain Chemistry; Capillaries; Dizocilpine Maleate; Epilepsy, Temporal Lobe; Excitatory Amino Acid Antagonists; Female; Glutamic Acid; Image Processing, Computer-Assisted; Immunohistochemistry; Muscarinic Agonists; Nerve Degeneration; Pilocarpine; Quinolines; Rats; Rats, Wistar; Seizures; 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

By intravenous administration of group I metabotropic glutamate receptor antagonists at 1 or 2h during pilocarpine induced status epilepticus (PISE), we showed that mGluR1 antagonists AIDA or LY367385 (at dosages ranging from 25 to 200mg/kg), mGluR5 antagonists SIB1757 (at dosages ranging from 25 to 200mg/kg), SIB1893 (from 25 to 100mg/kg), MPEP (from 25 to 100mg/kg) injected at 1 or 2h during PISE were ineffective in controlling status epilepticus (SE). However, when administered at 1h during PISE, MPEP at 200mg/kg, combination of MPEP (200mg/kg) with MK801 (0.1mg/kg) or with MK801 (0.1mg/kg) and diazepam (0.5mg/kg), combination of SIB1893 (200mg/kg) with MK801 (0.1mg/kg) could effectively control behavioral SE, and were neuroprotective. In particular, the combination of MPEP with MK801 and diazepam could stop both behavioral SE and electrical SE (under EEG monitoring) within a few minutes after the administration. HPLC study showed that a high level of MPEP was maintained in the blood and its metabolism rate was slow in experimental mice with PISE. We therefore concluded that the combination of MPEP (200mg/kg) with MK801 (0.1mg/kg) and diazepam (0.5mg/kg) could effectively stop SE and its subsequent neuronal loss in the hippocampus when administered 1h during PISE. It may provide a new approach to effectively control intractable SE.

Topics: Animals; Behavior, Animal; Chromatography, High Pressure Liquid; Diazepam; Disease Models, Animal; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Therapy, Combination; Electroencephalography; Excitatory Amino Acid Antagonists; GABA Modulators; Male; Mice; Oxazines; Pilocarpine; Pyridines; Status Epilepticus

This study aimed to determine whether pentylenetetrazol-induced status epilepticus (SE) can induce dentate granule cell neurogenesis in the developing rat and the effect of MK-801, a noncompetitive antagonism of N-methyl-D-aspartate receptor (NMDAR), on neurogenesis.. Two hundred and sixteen postnatal days 7, 14, 21 or 28 Sprague Dawley (SD) rats were involved in this study. Each age group consisted of 54 rats which were randomly assigned into a SE group, a SE + MK-801 group and a Normal control group (n=18 each). SE was induced by intraperitoneal injection of PTZ (80 mg/kg). The SE + MK-801 group was injected intraperitoneally with MK-801 (1 mg/kg) at 1 hr after SE episode. All rats were given 5-bromodeoxyuridene (BrdU) intraperitonealy to label newborn cells at 6, 13 and 27 days after seizures and then were sacrificed 24 hrs after BrdU injection. The immunohistochemistry method was used to measure the expression of BrdU, TuJl (betaIII tubulin), and glial fibrillary acidic protein (GFAP) in the dentate gyrus of hippocampus of rats.. The number of the BrdU positive cells in the SE group was significantly higher than in the age-matched normal controls at 7 and 14 days after SE episode (P <0.05 or 0.01). Approximately 82.5% and 80.3% of BrdU-labeled cells in the SE and the Control groups were co-expressed TuJ1 respectively. MK-801 treatment decreased the BrdU positive cells compared with the SE group at 7 and 14 days after SE seizures (P < 0.01). On the 28th day after SE episode there were no differences among the three groups for the BrdU positive cells.. PTZ-induced SE can increase the dentate granule cell neurogenesis in the developing rat. NMDAR plays an important role in neurogenesis following seizures.

Topics: Animals; Bromodeoxyuridine; Dentate Gyrus; Dizocilpine Maleate; Female; Glial Fibrillary Acidic Protein; Immunohistochemistry; Male; Pentylenetetrazole; Rats; Rats, Sprague-Dawley; Status Epilepticus

Status epilepticus and cerebral ischemia stimulate persistent neurogenesis in the adult brain, but both conditions cause neuronal damage. We determined whether spreading depression, a common epiphenomenon of these conditions, stimulates persistent neurogenesis.. We analyzed the effect of KCl-induced spreading depression on persistent neurogenesis and the spatio-temporal distribution of cells exhibiting immunohistochemical markers for divided and early committed neurons (new neurons) in the adult rat brain.. After induction of spreading depression for 48 hours, the density of mitotic cells, divided cells, and new neurons in the subventricular zone increased at days 1 to 3, days 3 to 6, and day 6, respectively (P<0.05). The divided cell density in the rostral migratory stream and the stream size increased at day 12 (P<0.001). Vehicle (saline) infusion or induction of spreading depression for 4 hours only did not increase the divided cell density, but the latter increased new neuron density in the subventricular zone (P<0.001). Double-labeled new neuron-like cells also appeared in the caudate putamen or cortex in ectopic fashion at day 3, with dramatic increases at days 6 and 12. Administration of the NMDA receptor antagonist, MK-801, which inhibits the propagation of spreading depression, abolished the increase in new neurons in the subventricular zone and the appearance of ectopic new neuron-like cells after 48-hour KCl infusion. There was no neuronal damage, as evidenced by mature neuron density, neurite density, and apoptotic cell appearance after spreading depression for 48 hours.. Spreading depression has the potential to stimulate persistent neurogenesis or to produce ectopic new neuron-like cells.

Topics: Animals; Antimetabolites; Apoptosis; Biomarkers; Brain; Brain Ischemia; Bromodeoxyuridine; Cell Differentiation; Cell Division; Central Nervous System; Cerebral Cortex; Cortical Spreading Depression; Dizocilpine Maleate; Glial Fibrillary Acidic Protein; Immunohistochemistry; In Situ Nick-End Labeling; Membrane Potentials; Microscopy, Confocal; Neurons; Potassium Chloride; Putamen; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Stem Cells; Time Factors

Three different classes of NMDA receptor antagonists were compared for their effectiveness in terminating prolonged status epilepticus (SE), induced by continuous hippocampal stimulation. Animals were treated after 150 min of SE by intraperitoneal administration of increasing doses of 3-((R,S)-2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP), MK-801 (dizocilpine), ifenprodil, or saline. EEG recordings were used to determine seizure termination. The first experiment (n = 57 animals) determined the most effective anticonvulsant dose of each agent by determining its ability to terminate SE within the next 300 min. Five control rats treated with normal saline after 150 min of SE continued to exhibit continuous seizures for the next 300 min. All drugs were administered after 150 min of SE. CPP terminated seizures with an ED(50) of 6.4 mg/kg; the maximal effective dose was 15 mg/kg. MK-801 has an ED(50) of 1.4 mg/kg; the maximal effective dose was 2 mg/kg. Ifenprodil was maximally effective at 30 mg/kg. However, an ED(50) could not be calculated. In a subsequent experiment, the NMDA antagonists were compared for their ability to terminate prolonged SE within 60 min of their administration at the most effective dose. MK-801 (2.0 mg/kg) terminated SE in 6 of 10 animals within 60 min, CPP (15 mg/kg) terminated it in 1 of 9 animals; ifenprodil (30 mg/kg) did not terminate it in any of 9 animals treated. In the 300 min following administration, CPP (6/9) and MK-801 (6/10) were equally efficacious in terminating SE but ifenprodil (2/7) was less effective (P = 0.065, chi-square test). The results indicate that the non-competitive NMDA receptor antagonist MK-801 was superior to the competitive antagonist CPP and the pH-sensitive site antagonist ifenprodil, in terminating prolonged experimental SE.

Topics: Animals; Dizocilpine Maleate; Dose-Response Relationship, Drug; Electric Stimulation; Excitatory Amino Acid Antagonists; Hippocampus; Male; Piperazines; Piperidines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Status Epilepticus

Dendritic targeting of mRNA and local protein synthesis are mechanisms that enable neurons to deliver proteins to specific postsynaptic sites. Here, we demonstrate that epileptogenic stimuli induce a dramatic accumulation of BDNF mRNA and protein in the dendrites of hippocampal neurons in vivo. BDNF mRNA and protein accumulate in dendrites in all hippocampal subfields after pilocarpine seizures and in selected subfields after other epileptogenic stimuli (kainate and kindling). BDNF accumulates selectively in discrete dendritic laminas, suggesting targeting to synapses that are active during seizures. Dendritic targeting of BDNF mRNA occurs during the time when the cellular changes that underlie epilepsy are occurring and is not seen after intense stimuli that are non-epileptogenic, including electroconvulsive seizures and high-frequency stimulation. MK801, an NMDA receptor antagonist that can prevent epileptogenesis but not acute seizures, prevents the dendritic accumulation of BDNF mRNA, indicating that dendritic targeting is mediated via NMDA receptor activation. Together, these results suggest that dendritic accumulation of BDNF mRNA and protein plays a critical role in the cellular changes leading to epilepsy.

Topics: Animals; Biological Transport; Brain-Derived Neurotrophic Factor; Cell Compartmentation; Convulsants; Dendrites; Dizocilpine Maleate; Electroshock; Epilepsy; Hippocampus; Kainic Acid; Kindling, Neurologic; Male; Pilocarpine; Protein Transport; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; RNA, Messenger; Seizures; Status Epilepticus; Synapses

Status epilepticus (SE) can result in acute neuronal injury with subsequent long-term age-dependent behavioral and histologic sequelae. To investigate potential mechanisms that may underlie SE-related neuronal injury, we studied the occurrence of programmed cell death (PCD) in the hippocampus in the kainic acid (KA) model.. In adult rats, KA-induced SE resulted in DNA fragmentation documented at 30 h after KA injection. Ceramide, a known mediator of PCD in multiple neural and nonneural tissues, increased at 2-3 h after KA intraperitoneal injection, and then decreased to control levels before increasing again from 12 to 30 h after injection. MK801 pretreatment prevented KA-induced increases in ceramide levels and DNA fragmentation, whether there was reduction in seizure severity or not (achieved with 5 mg/kg and 1 mg/kg of MK801, respectively).. Both ceramide increases and DNA fragmentation were observed after KA-induced SE in adult and in P35 rats. Ceramide did not increase after KA-induced SE in P7 pups, which also did not manifest any DNA fragmentation. Intrahippocampal injection of the active ceramide analogue C2-ceramide produced widespread DNA fragmentation, whereas the inactive ceramide analogue C2-dihydroceramide did not.. Our data support the hypotheses that (a) N-methyl-d-aspartate-receptor activation results in ceramide increases and in DNA fragmentation; (b) ceramide is a mediator of PCD after SE; and (c) there are age-related differences in PCD and in the ceramide response after SE. Differences in the ceramide response could, potentially, be responsible for observed age-related differences in the response to SE.

Topics: Age Factors; Animals; Apoptosis; Ceramides; Disease Models, Animal; Dizocilpine Maleate; DNA Fragmentation; Dose-Response Relationship, Drug; Hippocampus; Injections, Intraperitoneal; Kainic Acid; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Sphingosine; Status Epilepticus

Most patients with temporal lobe epilepsy (TLE), the most common type of epilepsy, show pronounced loss of neurons in limbic brain regions, including the hippocampus. The massive neurodegeneration in the hippocampus is known as hippocampal sclerosis, and is considered one of the hallmarks of this type of difficult-to-treat epilepsy. There is a long and ongoing debate on whether this sclerosis is the result of an initial pathological event, such as a status epilepticus (S.E.), stroke or head trauma, which often precedes the development of TLE, or is caused by the spontaneous recurrent seizures (SRS) once epilepsy has developed. At present, pharmacological prevention of limbic sclerosis is not available. In a clinical situation, such prevention would only be possible if delayed cell death developing after an initial pathological event is involved. Assuming that sclerotic brain lesions provoke epileptogenesis and that delayed cell death is involved in these lesions, it should be possible to prevent both the lesions and the epilepsy by a prophylactic treatment after an initial insult such as an S.E. In order to test this hypothesis, we used a rat model of TLE in which limbic brain lesions and epilepsy with SRS develop after a kainate-induced S.E. A single low dose of the N-methyl-D-aspartate (NMDA) receptor blocker dizocilpine (MK-801) significantly reduced the damage in limbic regions, including the hippocampus and piriform cortex, and completely protected several rats from such damage when given after an S.E. of 90 min induced by kainate, strongly suggesting that delayed cell death is involved in the damage. This was substantiated by the use of molecular and immunohistochemical markers of delayed active ("programmed") cell death. However, the neuroprotection by dizocilpine did not prevent the development of SRS after the S.E., suggesting that structures not protected by dizocilpine may play a role in the genesis of SRS or that epileptogenesis is not the consequence of structural lesions in the limbic system. The only brain regions that exhibited neuronal damage in all rats with SRS were the hilus of the dentate gyrus and the mediodorsal thalamus, although treatment with dizocilpine reduced the severity of damage in the latter region. The data indicate that NMDA receptor blockade immediately after a prolonged S.E. is an effective means to reduce the damage produced by a sustained S.E. in several brain regions, including the hippocampus, but show that this pa

Topics: Animals; Cell Death; Dentate Gyrus; Disease Models, Animal; Dizocilpine Maleate; DNA Fragmentation; Epilepsy, Temporal Lobe; Excitatory Amino Acid Antagonists; Female; Hippocampus; Kainic Acid; Limbic System; Mediodorsal Thalamic Nucleus; Nerve Degeneration; Neurons; Neuroprotective Agents; Olfactory Pathways; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Status Epilepticus; Treatment Outcome

Hippocampal sclerosis and massive neurodegeneration in other parts of the limbic system are considered hallmarks of temporal lobe epilepsy. Using the rat model of kainate-induced status epilepticus, we sought to determine if limbic sclerosis after an excitotoxic insult follows a delayed type of neurodegeneration and is thus accessible to neuroprotective intervention after the insult. Effective pharmacologic neuroprotection after status epilepticus also addresses the old question of whether degenerative morphologic changes after an epilepsy-inducing event like status epilepticus are the primary cause of epileptogenesis (i.e., the development of recurrent spontaneous seizures) during the following weeks.. Female Wistar rats after 90 min of generalized status epilepticus were used. Molecular biologic and histologic techniques were used to demonstrate markers of delayed cell death (apoptosis) 48 h after the status. The neuroprotective effects of i.c.v. injections of caspase inhibitors and systemic injections of the anticonvulsant drugs (AEDs) dizocilpine and retigabine after the status epilepticus were studied. The effect of neuroprotective intervention on the development of recurrent spontaneous seizures was investigated by behavioral observation of the rats.. After generalized status epilepticus in Wistar rats, massive sclerosis of the hippocampus and the piriform cortex occurred. TUNEL labeling and electron microscopy revealed that apoptosis is involved in the degenerative processes. Immunohistochemical analysis of the time course of the expression of the proapoptotic protein Bax suggested a maximal induction of apoptosis 24-48 h after the status. Application of caspase inhibitors before or after the status did not reduce lesions, although Bax labeling was reduced. Injection of dizocilpine and to a lower extent also of retigabine after the status prevented limbic neurodegeneration and expression of markers of apoptosis. However, the neuroprotection by dizocilpine did not prevent the development of recurrent spontaneous seizures.. Prolonged seizure activity can induce delayed sclerosis in the hippocampus and other parts of the limbic system. This delayed cell loss can be prevented by neuroprotective drugs after a status epilepticus. However, the damage in limbic brain regions is not the main reason for limbic epileptogenesis and the occurrence of recurrent spontaneous seizures.

Topics: Animals; Anticonvulsants; Apoptosis; Biomarkers; Carbamates; Caspase Inhibitors; Dizocilpine Maleate; Enzyme Inhibitors; Epilepsy; Female; Hippocampus; Limbic System; Neuroprotective Agents; Phenylenediamines; Rats; Rats, Wistar; Sclerosis; Status Epilepticus; Time Factors

To examine the effect of therapy on status epilepticus (SE) acutely and on long-term outcome, we compared three drugs with three different mechanisms. Phenobarbital, MK-801, and phenytoin were administered at 1, 2, and 4 hours after initiation of limbic status epilepticus by "continuous" hippocampal stimulation in rats. We evaluated the effects of these drugs on the course of SE and the subsequent development of chronic epilepsy. Phenobarbital and MK-801 were superior to phenytoin in suppressing SE and in preventing chronic epilepsy. There was no benefit if treatment was given 2 hours after the initiation of SE. Phenobarbital was most effective in suppressing electrographic seizure activity, but MK-801 had a slightly wider window for the prevention of chronic epilepsy. Early treatment, rather than electrographic suppression of SE, correlated with prevention of chronic epilepsy. This study shows that the drugs administered, which have different mechanisms of action, have clear differences in altering the outcomes. The findings suggest that studies of SE treatment should examine the effect of therapy on SE itself, as well as the long-term benefits of each treatment. The use of N-methyl-D-aspartate receptor antagonists should be considered early in the treatment of SE.

Topics: Animals; Anticonvulsants; Behavior, Animal; Chronic Disease; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Male; Phenobarbital; Phenytoin; Rats; Rats, Sprague-Dawley; Status Epilepticus

This study focused on the effects of status epilepticus on the activity of calcineurin, a neuronally enriched, calcium-dependent phosphatase. Calcineurin is an important modulator of many neuronal processes, including learning and memory, induction of apoptosis, receptor function and neuronal excitability. Therefore, a status epilepticus-induced alteration of the activity of this important phosphatase would have significant physiological implications. Status epilepticus was induced by pilocarpine injection and allowed to continue for 60 min. Brain region homogenates were then assayed for calcineurin activity by dephosphorylation of p-nitrophenol phosphate. A significant status epilepticus-dependent increase in both basal and Mn(2+)-dependent calcineurin activity was observed in homogenates isolated from the cortex and hippocampus, but not the cerebellum. This increase was resistant to 150 nM okadaic acid, but sensitive to 50 microM okadaic acid. The increase in basal activity was also resistant to 100 microM sodium orthovanadate. Both maximal dephosphorylation rate and substrate affinity were increased following status epilepticus. However, the increase in calcineurin activity was not found to be due to an increase in calcineurin enzyme levels. Finally, increase in calcineurin activity was found to be NMDA-receptor activation dependent. The data demonstrate that status epilepticus resulted in a significant increase in both basal and maximal calcineurin activity.

Topics: Animals; Brain; Calcineurin; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Hippocampus; Kinetics; Male; Nitrophenols; Organ Specificity; Organophosphorus Compounds; Pilocarpine; Protein Tyrosine Phosphatases; Rats; Rats, Sprague-Dawley; Seizures; Status Epilepticus; Substrate Specificity

Prolonged seizures (status epilepticus) induced by kainic acid activate programmed cell death mechanisms, and it is believed that kainic acid-induced status epilepticus induces neuronal apoptosis. In order to test this hypothesis, adult rats were subjected to 3-h kainic acid-induced seizures, with 24- or 72-h recovery periods. Neuronal death was assessed by light microscopy with the Hematoxylin and Eosin stain and with in situ terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL stain), by electron microscopy, and by agarose gel electrophoresis of DNA extracted from five vulnerable brain regions. Spontaneous and MK-801-induced apoptotic neurons from retrosplenial cortex of neonatal rats, evaluated by light and electron microscopy, were used as positive controls for apoptosis. Surprisingly, the large chromatin clumps of apoptotic neurons were TUNEL negative, whereas the cytoplasm showed light-to-moderate TUNEL staining, consistent with a lack of identifiable nuclear membranes ultrastructurally, and with intermingling of nuclear and cytoplasmic contents. Ultrastructurally, the acidophilic neurons produced by kainic acid-induced status epilepticus, identified with Hematoxylin and Eosin stain, were dark, shrunken and necrotic, with pyknotic nuclei containing small, dispersed chromatin clumps, and with cytoplasmic vacuoles, some of which were swollen, disrupted mitochondria. No apoptotic cells were seen. Acidophilic neurons were found in up to 20 of 23 brain regions examined and comprised 10-25% of the total number of neurons examined. A subset of these neurons (<10% of the total number of neurons in five of 23 regions) had TUNEL-positive nuclei 72h but not 24h after status epilepticus. Internucleosomal DNA cleavage (DNA "laddering") occurred in the four most damaged brain regions examined by electron microscopy 24h after SE and the three most damaged regions 72h after status epilepticus. Our results demonstrate that kainic acid-induced status epilepticus produces neuronal necrosis and not apoptosis in adult rats. The necrotic neurons show nuclear pyknosis, chromatin condensation and DNA laddering. Programmed cell death mechanisms activated by kainic acid-induced status epilepticus occur in neurons which become necrotic and could contribute to necrotic, as well as apoptotic, neuronal death.

Topics: Animals; Apoptosis; Dizocilpine Maleate; DNA Fragmentation; Entorhinal Cortex; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Female; Hippocampus; In Situ Nick-End Labeling; Kainic Acid; Male; Microscopy, Electron; Necrosis; Neurons; Nucleosomes; Rats; Rats, Wistar; Seizures; Status Epilepticus

The development of symptomatic epilepsy is a model of long-term plasticity changes in the central nervous system. The rat pilocarpine model of epilepsy was utilized to study persistent alterations in calcium/calmodulin-dependent kinase II (CaM kinase II) activity associated with epileptogenesis. CaM kinase II-dependent substrate phosphorylation and autophosphorylation were significantly inhibited for up to 6 weeks following epileptogenesis in both the cortex and hippocampus, but not in the cerebellum. The net decrease in CaM kinase II autophosphorylation and substrate phosphorylation was shown to be due to decreased kinase activity and not due to increased phosphatase activity. The inhibition in CaM kinase II activity and the development of epilepsy were blocked by pretreating seizure rats with MK-801 indicating that the long-lasting decrease in CaM kinase II activity was dependent on N-methyl-D-aspartate receptor activation. In addition, the inhibition of CaM kinase II activity was associated in time and regional localization with the development of spontaneous recurrent seizure activity. The decrease in enzyme activity was not attributed to a decrease in the alpha or beta kinase subunit protein expression level. Thus, the significant inhibition of the enzyme occurred without changes in kinase protein expression, suggesting a long-lasting, post-translational modification of the enzyme. This is the first published report of a persistent, post-translational alteration of CaM kinase II activity in a model of epilepsy characterized by spontaneous recurrent seizure activity.

Topics: Animals; Brain; Calcium-Calmodulin-Dependent Protein Kinases; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Intercellular Signaling Peptides and Proteins; Isoenzymes; Male; Okadaic Acid; Peptides; Phosphoric Monoester Hydrolases; Phosphorylation; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Secondary Prevention; Seizures; Status Epilepticus; Time Factors

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

Effects of various calcium channel inhibitors have been studied in lithium-pilocarpine model of status epilepticus. Status epilepticus was induced by administration of lithium chloride (3 meq/kg) followed 21 hr later by pilocarpine (30 mg/kg). Diltiazem (5 and 10 mg/kg) was not effective in delaying onset of convulsions. Verapamil (20 mg/kg) showed protection against lithium-pilocarpine-induced convulsions. The dihydropyridine nifidepine (2.5 and 5 mg/kg) did not show any protection in this model. Amlodipine (5 and 10 mg/kg) as partially protective. Flunarizine (10 and 20 mg/kg) delayed the onset of forelimb clonus and rearing and only 60% of the rats underwent status in the 20 mg/kg group. Pre-treatment of MK-801 led to a potentiation of the antiseizure activity of calcium channel inhibitors. The percent increase in amplitude at various time points with amlodipine pretreatment was significant only at the 30th min recording, and at the rest of the time frames was practically similar as the controls. It can be concluded that the anticonvulsant action of MK-801 can be enhanced by centrally acting calcium channel inhibitors.

Topics: Animals; Anticonvulsants; Calcium Channel Blockers; Disease Models, Animal; Dizocilpine Maleate; Drug Synergism; Electroencephalography; Female; Male; Rats; Rats, Wistar; Status Epilepticus

We examined the effects of blockers of N-methyl-D-asparate (NMDA) and +/- -alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)/kainate receptors on the maintenance of self-sustaining status epilepticus (SSSE) induced in rats by brief intermittent electrical stimulation of the perforant path (PPS). Blocking of NMDA receptor at the PCP site by MK-801 (0.5 mg/kg, i.p.) or ketamine (10 mg/kg, i.p.) as well as at the glycine allosteric site by intrahippocampal 5,7-dichlorokynurenic acid (5,7-DCK, 10 nmol), rapidly and irreversibly aborted both behavioral and electrographic manifestation of SSS. Intrahippocampal injection of the AMPA/kainate receptor blocker 6-cyano7-nitroquinixaline-3-dione (CNQX, 10 nmol) transiently suppressed seizures, which reappeared 4-5 h later. We suggest that the maintenance phase of SSSE depends on activation of NMDA receptors and that NMDA receptor blockers may be a promising class of compounds for the treatment of status epilepticus.

Topics: 6-Cyano-7-nitroquinoxaline-2,3-dione; Action Potentials; Animals; Anticonvulsants; Disease Models, Animal; Dizocilpine Maleate; Electric Stimulation; Excitatory Amino Acid Antagonists; Hippocampus; Ketamine; Kynurenic Acid; Male; Perforant Pathway; Rats; Rats, Wistar; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Status Epilepticus

Status epilepticus, prolonged intermittent or continuous seizure activity lasting 30 min or longer, is associated with high morbidity and mortality. The longer a seizure persists, the more refractory to treatment it becomes. The pilocarpine model of status epilepticus in rodents develops refractoriness to many first-line treatments as seizure duration increases, rendering it a good model to study refractory status epilepticus. This study was initiated to study the development of refractoriness of pilocarpine-induced status epilepticus to diazepam. Early pilocarpine-induced status epilepticus responded rapidly to diazepam treatment, whereas status epilepticus of longer duration became increasingly less responsive to treatment. Dizocilpine maleate-pretreated animals responded rapidly to diazepam treatment, even after 60 min of status epilepticus. Animals administered dizocilpine maleate at 15, 30 or 60 min after the onset of status epilepticus also demonstrated a rapid response to diazepam compared to pilocarpine-alone-treated animals. The longer the status epilepticus progressed prior to dizocilpine maleate injection, the longer the status epilepticus lasted after diazepam treatment. However, in all cases where dizocilpine maleate was administered, one injection of diazepam was able to terminate the status epilepticus, in contrast to the animals that did not receive dizocilpine maleate, in which the seizure was only attenuated. The results indicate that N-methyl-D-aspartate receptor activation plays a role in the seizure-induced refractoriness to benzodiazepines in status epilepticus, and blocking N-methyl-D-aspartate receptor activation converts refractory status epilepticus to a seizure responsive to benzodiazepine therapy. These findings offer insights into developing novel therapeutic interventions to improve the treatment of status epilepticus. Understanding the molecular mechanisms that mediate the effects of N-methyl-D-aspartate receptor activation on the development of resistance to treatment in status epilepticus will provide rational insights into more rapid methods to terminate seizure activity in this condition.

Topics: Animals; Anticonvulsants; Diazepam; Dizocilpine Maleate; Drug Resistance; Electroencephalography; Excitatory Amino Acid Antagonists; Muscarinic Agonists; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Status Epilepticus

We have previously shown that NMDA receptor activation during status epilepticus (SE) is required to produce epilepsy in in vitro and in vivo models. As in human symptomatic epilepsy, the epilepsy in these models is permanent, suggesting that the pathological activation of NMDA receptors causes permanent plasticity changes in the brain. Ca(2+) influx through NMDA receptors is known to transiently activate a key transcription factor, serum response factor (SRF). Thus, we investigated whether this factor, in terms of its expression and ability to bind to the consensus serum response element, was altered long term in the pilocarpine model of epilepsy. In hippocampal nuclear extracts, SRF binding to DNA was significantly increased over saline-injected control rats at 24 hr and at 8 weeks after the onset of SE. This increase was shown to be the result of significantly elevated levels of SRF. DNA binding was also persistently increased in the cortical, but not in the cerebellar, extracts. Hippocampal expression of SRF was localized to neurons using immunohistochemistry. NMDA receptor activation during SE was required for these changes to take place, and the spontaneous seizures seen in epileptic rats did not appear to be responsible for the increase in SRF. The results demonstrate that SRF is persistently elevated after SE in the pilocarpine model of epilepsy and support the theory that long-term gene changes in this model occur and are associated with the long-lasting plasticity changes that are initiated during epileptogenesis.

Topics: Animals; Brain; Cell Nucleus; Cerebellum; Cerebral Cortex; Dizocilpine Maleate; DNA-Binding Proteins; Electroshock; Gene Expression Regulation; Hippocampus; Humans; Male; Neuronal Plasticity; Nuclear Proteins; Oligodeoxyribonucleotides; Organ Specificity; Phenytoin; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Seizures; Serum Response Factor; Status Epilepticus; Transcription Factors

NMDA receptor activation has been implicated in modulating seizure activity; however, its complete role in the development of epilepsy is unknown. The pilocarpine model of limbic epilepsy involves inducing status epilepticus (SE) with the subsequent development of spontaneous recurrent seizures (SRSs) and is widely accepted as a model of limbic epilepsy in humans. The pilocarpine model of epilepsy provides a tool for looking at the molecular signals triggered by SE that are responsible for the development of epilepsy. In this study, we wanted to examine the role of NMDA receptor activation on the development of epilepsy using the pilocarpine model. Pretreatment with the NMDA receptor antagonist MK-801 does not block the onset of SE in the pilocarpine model. Thus, we could compare animals that experience similar lengths of SE in the presence or absence of NMDA receptor activation. Animals treated with MK-801 (4 mg/kg) 20 min prior to pilocarpine (350 mg/kg) (MK-Pilo) were compared to the pilocarpine treated epileptic animals 3-8 weeks after the initial episode of SE. The pilocarpine-treated animals displayed both ictal activity and interictal spikes on EEG analysis, whereas MK-801-pilocarpine and control animals only exhibited normal background EEG patterns. In addition, MK-801-pilocarpine animals did not exhibit any SRSs, while pilocarpine-treated animals exhibited 4.8 +/- 1 seizures per 40 h. MK-801-pilocarpine animals did not demonstrate any decrease in pyramidal cell number in the CA1 subfield of the hippocampus, while pilocarpine animals averaged 15% decrease in cell number. In summary, the MK-801-pilocarpine animals exhibited a number of characteristics similar to control animals and were statistically significantly different from pilocarpine-treated animals. Thus, NMDA receptor inhibition by MK-801 prevented the development of epilepsy and interictal activity following SE. These results indicate that NMDA receptor activation is required for epileptogenesis following SE in this model of limbic epilepsy.

Topics: Animals; Cell Count; Dizocilpine Maleate; Electroencephalography; Epilepsy; Excitatory Amino Acid Antagonists; Hippocampus; Male; Neurons; Pilocarpine; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Recurrence; Status Epilepticus; Time Factors

Sequential treatment of rats with low doses of lithium and pilocarpine, a high dose of pilocarpine, or continuous hippocampal stimulation [CHS] (9 epochs, 10 min each) is reported to result in status epilepticus (SE). We report a novel method to establish SE based on continuous ventral hippocampal stimulation (5 epochs) followed by low dose pilocarpine (40 mg/kg) challenge. Motor limbic seizures occured in all the control rats. The latency to spike activity was 15 +/- 1 min after pilocarpine administration. Ventral hippocampal [VHc] and cortical EEG recordings were used to monitor the protective effect of diazepam (5 mg/kg). Except phenobarbital, all the three drugs completely prevented all the phases of seizure activity. Initiation of spikes was significantly prolonged by phenobarbital pretreatment. Further study on the characteristics of these convulsions offers a unique possibility for the recognition of brain regions, pathways, and neurotransmitters engaged in the spread of seizures in this model.

Topics: Animals; Anticonvulsants; Disease Models, Animal; Dizocilpine Maleate; Electroencephalography; Hippocampus; Limbic System; Lithium; Male; Neuroprotective Agents; Phenobarbital; Pilocarpine; Rats; Rats, Wistar; Seizures; Status Epilepticus; Stereotyped Behavior

We investigated whether the non-competitive NMDA receptor antagonist, MK-801, could protect neurons in the immature brain from the excitotoxic affects of perforant path stimulation. A high dose of MK-801 reduced the number of injured hilar interneurons in the stimulated hippocampus from 30.0 +/- 5.2 in unmedicated rats to 12.2 +/- 9.6 in MK-801 treated animals (P < 0.05). MK-801 injection also protected the animals from the scattered dentate granule cell injury observed in non-medicated animals 1 day after stimulation. Other effects of drug injection included exacerbated damage in limbic cortices, retrosplenial cortical damage, and reduced inhibition in a highly epileptogenic region of the dentate gyrus. Our results show that a subpopulation of hilar interneurons is vulnerable to NMDA-induced damage in the immature hippocampus but that non-competitive blockade of the NMDA receptor may be a dangerous therapeutic strategy.

Topics: Age Factors; Animals; Behavior, Animal; Cell Count; Disease Models, Animal; Dizocilpine Maleate; Electric Stimulation; Hippocampus; Neurons; Neuroprotective Agents; Neurotoxins; Rats; Receptors, N-Methyl-D-Aspartate; Status Epilepticus

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

MUSCARINIC, NMDA and metabotropic glutamate receptor antagonists were tested for anticonvulsant effects in a continuous hippocampal stimulation model in rats in order to identify the receptors involved in the initiation of electrically induced status epilepticus (SE). Whereas the muscarinic receptor antagonists scopolamine and atropine and the metabotropic receptor antagonist L(+)-2-amino-3-phosphonopropionic acid (AP3) did not affect SE initiation, the N-methyl-D-aspartate (NMDA) antagonist dizocilpine (MK-801) (1 mg kg-1 i.p.) and the non-NMDA ionotropic receptor antagonist 2,3-dihydroxy-6-nitro-7-sulphamoylbenzo(F)-quinoxaline (NBQX) (two doses of 50 micrograms i.c.v.) prevented the induction of SE. It has been shown in a previous study that non-NMDA ionotropic receptors are involved in SE maintenance and it is now suggested that activation of NMDA receptors may principally initiate electrically induced SE, although non-NMDA ionotropic receptors may also be involved.

Topics: Animals; Atropine; Cerebral Ventricles; Dizocilpine Maleate; Electric Stimulation; Hippocampus; Injections, Intraperitoneal; Injections, Intraventricular; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, AMPA; Receptors, N-Methyl-D-Aspartate; Scopolamine; Status Epilepticus

The role of N-methyl-D-aspartate (NMDA), non-NMDA glutamate, metabotropic and muscarinic receptors in the maintenance of status epilepticus (SE) was investigated. SE induced in rat brain by continuous electrical stimulation to the hippocampus was terminated by intracerebroventricular (i.c.v.) injection of the non-NMDA antagonists DNQX and NBQX, but not by the muscarinic antagonists scopolamine or atropine, or the metabotropic antagonist AP3. The NMDA antagonist, MK-801 suppressed motor seizure activity but did not terminate electrographic seizures when generalized SE was induced, suggesting that both non-NMDA and NMDA receptors maintain generalized convulsive SE. However, when limbic SE was induced, MK-801 also had an anticonvulsant effect suggesting differences in the mechanisms maintaining limbic SE and generalized SE.

Topics: Animals; Anticonvulsants; Atropine; Brain; Cerebral Ventricles; Dizocilpine Maleate; Electric Stimulation; Electroencephalography; Hippocampus; Injections, Intraventricular; Male; Quinoxalines; Rats; Rats, Wistar; Receptors, Glutamate; Receptors, Muscarinic; Scopolamine; Status Epilepticus

Neurons undergoing delayed neuronal death produced by hypoxia-ischaemia (HI) or status epilepticus (SE) showed a massive expression of c-Jun in their nuclei 24 h after the insult. With SE there was also a weaker induction of c-Fos and Jun B in dying neurons. SE induced in the presence of the NMDA antagonist MK-801 produced no delayed c-Jun expression in the hippocampus and nerve cell death did not occur in this region, although there was a delayed c-jun expression in the amygdala/piriform region, and cell death occurred in this area. Activation of central muscarinic receptors with pilocarpine, or block of D2 dopamine receptors with haloperidol, treatments which do not cause neuronal damage, strongly induced Fos and Jun B in hippocampal and striatal neurons, but only induced c-Jun very weakly. Thus, c-Jun may participate in the genetic cascade of events that produce programmed cell death in neurons.

Topics: Animals; Apoptosis; Brain Ischemia; Dizocilpine Maleate; Genes, fos; Genes, jun; Haloperidol; Hypoxia, Brain; Male; Neurons; Pilocarpine; Rats; Rats, Wistar; Status Epilepticus; Transcription, Genetic

MK801 is a noncompetitive blocker of N-methyl-D-aspartate receptors which has antiepileptic properties. To evaluate whether MK801 pretreatment in immature rats affects the future spontaneous recurrent seizure (SRS) rate or seizure susceptibility in a model of limbic epilepsy, MK801 (0.2 or 1.0 mg/kg, i.p.) or saline was administered to prepubescent rats 30 min prior to kainic acid (KA; 10 mg/kg, i.p.). With or without MK801 pretreatment, KA caused prolonged status epilepticus. SRS rate over the next 4 weeks, as assessed by intermittent video monitoring, was significantly lower in MK801 treated rats than in those which received KA alone. In addition, fewer MK801 treated rats (43%) developed SRS than those which got KA alone (88%). Susceptibility to generalized seizures was then tested using the volatile convulsant flurothyl; at both doses of MK801, flurothyl seizure latency was significantly greater in pretreated animals. These results show that MK801 pretreatment prior to KA induced status epilepticus reduces subsequent SRS frequency and flurothyl seizure susceptibility, while not substantially altering the acute epileptogenic effects of KA.

Topics: Animals; Cerebral Cortex; Dizocilpine Maleate; Electroencephalography; Epilepsy, Tonic-Clonic; Flurothyl; Kainic Acid; Male; Rats; Rats, Sprague-Dawley; Seizures; Status Epilepticus

High doses of pilocarpine are known to produce prolonged seizure activity in rats. Previous studies show that pretreatment with LiCl will sensitize rats to the convulsant effects of pilocarpine. Here we report that the anti-epileptogenic and anticonvulsant drug MK-801 (dizocilpine) paradoxically also sensitizes rats to pilocarpine induced limbic seizures and limbic status epilipticus (LSE). If rats were pretreated with MK-801, normally subconvulsive doses of pilocarpine rapidly produced LSE. LSE could be prevented by co-administration of either atropine or diazepam with MK-801.

Topics: Animals; Atropine; Diazepam; Dizocilpine Maleate; Drug Synergism; Electroencephalography; Limbic System; Male; N-Methylaspartate; Pilocarpine; Rats; Rats, Wistar; Seizures; Status Epilepticus

Topics: Animals; Cerebral Cortex; Disease Models, Animal; Dizocilpine Maleate; Electric Stimulation; Electroencephalography; Hippocampus; Male; Phenobarbital; Rats; Rats, Sprague-Dawley; Status Epilepticus

The involvement of the NMDA receptor in the neurotoxicity induced by soman, an organophosphorus compound which irreversibly inhibits cholinesterase, was studied in guinea pigs. The drug MK-801 (0.5, 1 or 5 mg/kg, i.p.) was given as a pretreatment before a convulsant dose of soman or as a posttreatment (30, 100 or 300 micrograms/kg, i.m.) 5 min after the development of soman-induced status epilepticus. Pyridostigmine, atropine and pralidoxime chloride were also given to each subject to counteract the lethality of soman. All subjects that were challenged with soman and given the vehicle for MK-801 (saline) exhibited severe convulsions and electrographic seizure activity. Neuronal necrosis was found in the hippocampus, amygdala, thalamus and the pyriform and cerebral cortices of those subjects surviving for 48 hr. Pretreatment with 0.5 or 1 mg/kg doses of MK-801 did not prevent nor delay the onset of seizure activity but did diminish its intensity and led to its early arrest. At the largest dose (5 mg/kg), MK-801 completely prevented the development of seizure activity and brain damage. Posttreatment with MK-801 prevented, arrested or reduced seizure activity, convulsions and neuronal necrosis in a dose-dependent manner. The NMDA receptor may play a more critical role in the spread and maintenance, rather than the initiation of cholinergically-induced seizure activity.

Topics: Amygdala; Animals; Brain; Cerebral Cortex; Dizocilpine Maleate; Electroencephalography; Evoked Potentials; Guinea Pigs; Hippocampus; Necrosis; Neurons; Organ Specificity; Soman; Status Epilepticus; Thalamus; Time Factors

Pharmacologic control of refractory status epilepticus has been little-studied in experimental models. In this experiment, rats in status epilepticus induced by lithium and pilocarpine were treated with MK-801 alone, diazepam alone or MK-801 plus diazepam, with treatment begun at a time when this model of status is refractory to anticonvulsant drugs. EEG samples were digitized before and for two hours after treatment, and the digitized samples subjected to computerized frequency analysis. MK-801 plus diazepam halted all manifestations of status epilepticus. Although neither MK-801 alone nor diazepam alone stopped the ongoing electrographic status epilepticus, both drugs diminished motor seizures and total EEG power. MK-801 treatment prevented the progression of changes in EEG pattern which normally occurs in this model of status epilepticus, while diazepam did not. MK-801, with and without diazepam, allowed the rats to survive the episode of status epilepticus, but rats treated with MK-801 alone required several days to recover completely, while the MK-801 plus diazepam rats appeared normal the next day. MK-801 may be a useful agent for treatment of human refractory status epilepticus, because of its neuroprotective action as well as its ability to potentiate GABAergic drugs.

Topics: Animals; Behavior, Animal; Diazepam; Dizocilpine Maleate; Drug Interactions; Electroencephalography; Lithium; Male; Pilocarpine; Rats; Rats, Inbred Strains; Status Epilepticus

Status epilepticus (SE) evolves through several stages when untreated. The later stages of SE are less responsive to standard anticonvulsants and may require general anesthesia to suppress seizures. Antagonists acting at the N-methyl-D-aspartate (NMDA) subclass of glutamate (excitatory) receptors have been demonstrated to exert antiepileptic activity in some seizure models. We report experiments performed to determine if NMDA receptor antagonists are effective in stopping seizures in the late stages of SE. A model of limbic SE induced by 90 min of 'continuous' electrical stimulation of the hippocampus in rats was employed. Three NMDA receptor antagonists, one 'competitive' (CPP) and two 'non-competitive' (ketamine and MK-801), were compared to 3 standard antiepileptic drugs (diazepam, phenobarbital, and phenytoin) for their ability to suppress seizures at a physiologically defined stage of SE. All NMDA receptor antagonists, diazepam and phenobarbital were effective in suppressing behavioral and electrographic seizures for varying periods of time. Phenytoin had no effect on SE. Ketamine and MK-801 induced a paradoxical enhancement of electrographic seizures that preceded SE suppression. We believe that NMDA-receptor antagonists offer a novel approach for treating the late stages of SE.

Topics: Animals; Anticonvulsants; Dibenzocycloheptenes; Dizocilpine Maleate; Electric Stimulation; Hippocampus; Male; Piperazines; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Status Epilepticus

MK-801, a noncompetitive N-methyl-D-aspartate (NMDA) receptor antagonist, was tested for anticonvulsant effects in rats using two seizure models, coadministration of lithium and pilocarpine and administration of a high dose of pilocarpine alone. Three major results are reported. First, pretreatment with MK-801 produced an effective and dose-dependent anticonvulsant action with the lithium-pilocarpine model but not with rats treated with pilocarpine alone, suggesting that different biochemical mechanisms control seizures in these two models. Second, the anticonvulsant effect of MK-801 in the lithium-pilocarpine model only occurred after initial periods of seizure activity. This observation is suggested to be an in vivo demonstration of the conclusion derived from in vitro experiments that MK-801 binding requires agonist-induced opening of the channel sites of the NMDA receptor. Third, although it is relatively easy to block seizures induced by lithium and pilocarpine by administration of anticonvulsants prior to pilocarpine, it is more difficult to terminate ongoing status epilepticus and block the lethality of the seizures. Administration of MK-801 30 or 60 min after pilocarpine, i.e., during status epilepticus, gradually reduced electrical and behavioral seizure activity and greatly enhanced the survival rate. These results suggest that activation of NMDA receptors plays an important role in status epilepticus and brain damage in the lithium-pilocarpine model. This was further supported by results showing that nonconvulsive doses of NMDA and pilocarpine were synergistic, resulting in status epilepticus and subsequent mortality.

Topics: Animals; Anticonvulsants; Aspartic Acid; Cerebral Cortex; Dibenzocycloheptenes; Dizocilpine Maleate; Dose-Response Relationship, Drug; Drug Synergism; Electroencephalography; Hippocampus; Lithium; Male; N-Methylaspartate; Pilocarpine; Rats; Rats, Inbred Strains; Receptors, N-Methyl-D-Aspartate; Receptors, Neurotransmitter; Status Epilepticus