Seizure-related Brain Damage Research Articles

Pregabalin Attenuates Excitotoxicity in Diabetes

Diabetes can exacerbate seizures and worsen seizure-related brain damage. In the present study, we aimed to determine whether the standard antiepileptic drug pregabalin (PGB) protects against pilocarpine-induced seizures and excitotoxicity in diabetes. Adult male Sprague-Dawley rats were divided into either a streptozotocin (STZ)-induced diabetes group or a normal saline (NS) group. Both groups were further divided into subgroups that were treated intravenously with either PGB (15 mg/kg) or a vehicle; all groups were treated with subcutaneous pilocarpine (60 mg/kg) to induce seizures. To evaluate spontaneous recurrent seizures (SRS), PGB-pretreated rats were fed rat chow containing oral PGB (450 mg) for 28 consecutive days; vehicle-pretreated rats were fed regular chow. SRS frequency was monitored for 2 weeks from post-status epilepticus day 15. We evaluated both acute neuronal loss and chronic mossy fiber sprouting in the CA3 area. In addition, we performed patch clamp recordings to study evoked excitatory postsynaptic currents (eEPSCs) in hippocampal CA1 neurons for both vehicle-treated rats with SRS. Finally, we used an RNA interference knockdown method for Kir6.2 in a hippocampal cell line to evaluate PGB's effects in the presence of high-dose ATP. We found that compared to vehicle-treated rats, PGB-treated rats showed less severe acute seizure activity, reduced acute neuronal loss, and chronic mossy fiber sprouting. In the vehicle-treated STZ rats, eEPSC amplitude was significantly lower after PGB administration, but glibenclamide reversed this effect. The RNA interference study confirmed that PGB could counteract the ATP-sensitive potassium channel (KATP)-closing effect of high-dose ATP. By opening KATP, PGB protects against neuronal excitotoxicity, and is therefore a potential antiepileptogenic in diabetes. These findings might help develop a clinical algorithm for treating patients with epilepsy and comorbid metabolic disorders.

Ketamine combinations for the field treatment of soman-induced self-sustaining status epilepticus. Review of current data and perspectives

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.

Combinations of ketamine and atropine are neuroprotective and reduce neuroinflammation after a toxic status epilepticus in mice

Epileptic seizures and status epilepticus (SE) induced by the poisoning with organophosphorus nerve agents (OP), like soman, are accompanied by neuroinflammation whose role in seizure-related brain damage (SRBD) is not clear. Antagonists of the NMDA glutamate ionotropic receptors are currently among the few compounds able to arrest seizures and provide neuroprotection even during refractory status epilepticus (RSE). Racemic ketamine (KET), in combination with atropine sulfate (AS), was previously shown to counteract seizures and SRBD in soman-poisoned guinea-pigs. In a mouse model of severe soman-induced SE, we assessed the potentials of KET/AS combinations as a treatment for SE/RSE-induced SRBD and neuroinflammation. When starting 30min after soman challenge, a protocol involving six injections of a sub-anesthetic dose of KET (25mg/kg) was evaluated on body weight loss, brain damage, and neuroinflammation whereas during RSE, anesthetic protocols were considered (KET 100mg/kg). After confirming that during RSE, KET injection was to be repeated despite some iatrogenic deaths, we used these proof-of-concept protocols to study the changes in mRNA and related protein contents of some inflammatory cytokines, chemokines and adhesion molecules in cortex and hippocampus 48h post-challenge. In both cases, the KET/AS combinations showed important neuroprotective effects, suppressed neutrophil granulocyte infiltration and partially suppressed glial activation. KET/AS could also reduce the increase in mRNA and related pro-inflammatory proteins provoked by the poisoning. In conclusion, the present study confirms that KET/AS treatment has a strong potential for SE/RSE management following OP poisoning. The mechanisms involved in the reduction of central neuroinflammation remain to be studied.

A new use for an old method: The Woelcke myelin stain for counting degenerating neurons in the brain of mice following status epilepticus

The Woelcke method is classically used for myelin staining. Degenerating neurons can be revealed histologically by hemalun and phloxin (H&P) where they appear “eosinophilic”. In the first 24h following soman-induced status epilepticus, we observed that the Woelcke method also revealed condensed, dark blue/black cells (W+ cells) in the gray matter of brain regions known to be sites of seizure-related brain damage, marked by the presence of eosinophilic cells. In the present study, using adjacent brain sections alternately stained with either the Woelcke or the H&P method, we show that eosinophilic cells and W+ cells are the same degenerating cells. Moreover, we show that semi-automated quantitative evaluation of W+ cells through computerized image analysis is considerably easier and faster than that of eosinophilic cells. It is therefore concluded that the Woelcke technique could be very useful, especially for quantifying acute brain cell damage following status epilepticus.

Prediction of soman-induced cerebral damage by distortion product otoacoustic emissions

The organophosphorus nerve agent soman is an irreversible cholinesterase (ChE) inhibitor that can produce long-lasting seizures and seizure-related brain damage (SRBD) in which acetylcholine and glutamate are involved. Since these neurotransmitters play a key-role in the auditory function, it was hypothesized that a hearing test may be an efficient way for detecting the central effects of soman intoxication. In the present study, distortion product otoacoustic emissions (DPOAEs), a non-invasive audiometric method, were used in rats administered with soman (70 μg/kg). Four hours post-soman, DPOAE intensities were significantly decreased. They returned to baseline one day later. The amplitude of the temporary drop of the DPOAEs was well related to the severity of the intoxication. The greatest change was recorded in the rats that survived long-lasting convulsions, i.e. those that showed the highest ChE inhibition in brain and severe encephalopathy. Furthermore, the administration, immediately after soman, of a three-drug therapy composed of atropine sulfate, HI-6 and avizafone abolished the convulsions, the transient drop of DPOAEs at 4 h and the occurrence of SRBD at 28 h without modifying brain ChE inhibition. This showed that DPOAE change was not directly related to soman-induced inhibition of cerebral ChE but rather to its neuropathological consequences. The present findings strongly suggest that DPOAEs represent a promising non-invasive tool to predict SRBD occurrence in nerve agent poisoning and to control the efficacy of a neuroprotective treatment.

Hypertonic mannitol in mice poisoned by a convulsive dose of soman: Antilethal activity without neuroprotection

A convulsive dose of soman induces seizure-related brain damage (SRBD), including cerebral edema (CE) and cell death. In 1993, an American study demonstrated that hypertonic mannitol (Mann) intravenously (i.v.) administered 1 min and 5 h post-soman was an effective neuroprotectant in intoxicated rats. Using a similar protocol, we recently failed to reproduce this success in intoxicated mice. In the present study, also performed in mice, the persistence or the amplitude of the osmotic load was increased by reducing the time interval between two injections of Mann or by augmenting the number of injections. Mice were pre-treated with the oxime HI-6 and then intoxicated with a convulsive dose of soman (172 μg/kg). Afterward, they were administered a first i.v. bolus of Mann 20% 1 min post-challenge and a second one either 5, or 2, or 1 h after. Additional animals were given either one (1 min post-soman), or two (1 min and 1 h post-soman), or three (1 min, 1 and 2 h post-soman) series of three injections of Mann at 5 min intervals. Non-intoxicated mice treated with Mann (same protocols as above) and intoxicated mice treated with Mann vehicle (saline) served as controls. At 24 h post-intoxication, the survivors were sacrificed and their brains prepared for quantitative histological assessment of cell damage, CE, and ventricle size. Whatever the protocol, Mann had no effect on soman-induced convulsions but did provide considerable antilethal activity. Histologically, Mann did not reduce the cell damage or CE. It even showed a dose-dependent trend toward aggravation of SRBD in some regions and promoted subarachnoid hemorrhages. Conversely, in one of the treatment protocol, it reduced soman-induced enlargement of ventricle size. Although treatment with hypertonic Mann showed some benefit on mortality and ventricle size, it failed to be an effective neuroprotector in soman-intoxicated mice and even increased the detrimental impact of soman at the cerebral level. Therefore, no clear recommendation could be drawn from the present study in view of a possible clinical use of hyperosmolar treatment in the medical management of soman poisoning.

Changes in mouse brain metabolism following a convulsive dose of soman: A proton HRMAS NMR study

Soman, an irreversible organophosphorus cholinesterase inhibitor, induces status epilepticus and, in sensitive brain areas, seizure-related brain damage (e.g. brain edema and neuronal loss). The brain metabolic disturbances associated with these events are ill known. In the present study, we thus evaluated these changes in a murine model of soman-induced status epilepticus up to 7 days after intoxication. Mice, protected by HI-6 and atropine methyl nitrate, were poisoned with soman (172 microg/kg) and then sacrificed at set time points, from 1 h to 7 days. Brain biopsies from the piriform cortex (Pir) and cerebellum (Cer) were analyzed by 1H HRMAS NMR spectroscopy. Spectra were then analyzed using both a supervised multivariate analysis and the QUEST procedure of jMRUI for the quantification of 17 metabolites. The multivariate analysis clearly showed the metabolic differences between a damaged structure (Pir) and a structure with less prominent changes (cerebellum) and helped to globally assess the time course of metabolic changes. Analysis of the individual metabolites showed that the major changes took place in the piriform cortex but that cerebellum was not change-free. The most prominent changes in the former were an early (1-4 h) increase in alanine and acetate, a delayed increase in lactate, glycerophosphocholine and glutamine as well as a delayed decrease in myo-inositol and N-acetylaspartate. A week after poisoning, some metabolic disturbances were still present. Further research will be necessary to clarify what could be the involvement of these metabolites in physiological processes and how they might become useful surrogate markers of brain damage and repair.

Hyperosmolar treatment of soman-induced brain lesions in mice: Evaluation of the effects through diffusion-weighted magnetic resonance imaging and through histology

Purpose A convulsive dose of soman induces seizure-related brain damage (SRBD), including cerebral edema (CE) and neuronal loss. In the present study on soman-intoxicated mice, we applied diffusion-weighted magnetic resonance imaging (DW-MRI) and quantitative histology, and we measured brain water content to investigate the antiedematous and neuroprotective efficacies of two hyperosmolar treatments: mannitol (Mann) and hypertonic saline (HTS). Methods Mice intoxicated with soman (172 μg/kg after a protective pretreatment) were administered 1 min and 5-h post-challenge an i.v. bolus of saline, of Mann or of HTS. 1 day later, mice were examined with DW-MRI and then sacrificed for brain histology. Additional animals were intoxicated and treated similarly for the measurement of the brain water content (dry/wet weight method). Results In intoxicated controls, a significant decrease of the apparent diffusion coefficient (ADC), numerous damaged (eosinophilic) cells, high edema scores, and a significant increase in brain water content were detected 24-h post-challenge in sensitive brain structures. These soman-induced changes were not significantly modified by treatment with Mann or HTS. Conclusions Treatment with hyperosmolar solutions did not reduce the effects of soman on ADC, on cell damage and on CE. Therefore, despite similar treatment protocols, the prominent protection by Mann that was previously demonstrated by others in poisoned rats, was not reproduced in our murine model.

The role of mitochondria in status epilepticus

Reflecting their central roles in cellular metabolism,including energy production and calcium homeostasis, mi-tochondria have been proposed to act as the “stress sensor”of the cell, and in extreme circumstances (e.g., excitotoxi-city) the executioner. A large body of work has establishedmitochondrial dysfunction as the “lowest common denom-inator” in a range of inherited (Kwong et al., 2006) and ac-quired (Cock, 2005) neurodegenerative disease processes.In addition to bioenergetic failure and increased cytosoliccalcium, consequences of impaired mitochondrial func-tion (summarized in Fig. 1) include oxidative stress (ex-cessive free radical production and impaired synthesis ofantioxidants, especially glutathione), and the release ofkey proteins into the cytosol (mitochondrial permeabilitytransition) triggering cell death pathways (caspases). How-ever, it is also increasingly clear that many of the “patho-logical” processes also have key physiological roles inmetabolic control and cell signaling, in which context pro-teins such as uncoupling proteins have attracted particu-lar interest, but are not yet fully understood (Cock, 2005).On this background, interest in the role of mitochondriain seizure-related brain damage has rapidly developed overthe last decade; though only work specifically relating tostatus epilepticus (as opposed to chronic epilepsy) will bediscussed here.

Prolonged inflammatory gene response following soman-induced seizures in mice

Following exposure to the organophosphorus nerve agent soman, the development of long-lasting seizures and build-up of irreversible seizure-related brain damage (SRBD) still represent a therapeutic challenge. A neuro-inflammatory reaction takes place in the brain after poisoning but its characteristics and potential role in SRBD and post-status epilepticus epileptogenesis is not well understood. In the present study we have analyzed by quantitative RT-PCR the time course of changes in mRNA levels of IL-1β, TNFα, IL-6, ICAM-1 and SOCS3 in hippocampus, whole cortex and cerebellum in a mouse model of severe seizures and neuropathy up to 7 days after poisoning. Mice received an injection of the oxime HI-6 (50 mg/kg) 5 min prior to the administration of a convulsive dose of soman (172 μg/kg). An important and highly significant increase of the five mRNA levels was recorded in cortex and hippocampus. In the cortex, the activation was generally detected as early as 1 h post-intoxication with a peak response recorded between 6 and 24 h. In the hippocampus, the gene up-regulation was delayed to 6 h post-soman and the peak response observed between 24 and 48 h. After peaking, the response declined (except for ICAM in the hippocampus) but remained elevated, some of them significantly, at day 7. Interestingly, in the cerebellum, some changes were also observed but were several fold smaller. In conclusion, the present study indicates a quick neuro-inflammatory gene response that does not subside over 7 days suggesting a potential role in the neurological consequences of soman-induced status epilepticus.

Protective effects of S(+) ketamine and atropine against lethality and brain damage during soman-induced status epilepticus in guinea-pigs

Soman poisoning is known to induce full-blown tonic–clonic seizures, status epilepticus (SE), seizure-related brain damage (SRBD) and lethality. Previous studies in guinea-pigs have shown that racemic ketamine (KET), with atropine sulfate (AS), is very effective in preventing death, stopping seizures and protecting sensitive brain areas when given up to 1 h after a supra-lethal challenge of soman. The active ketamine isomer, S(+) ketamine (S-KET), is more potent than the racemic mixture and it also induces less side-effects. To confirm the efficacy of KET and to evaluate the potential of S-KET for delayed medical treatment of soman-induced SE, we studied different S-KET dose regimens using the same paradigm used with KET. Guinea-pigs received pyridostigmine (26 μg/kg, IM) 30 min before soman (62 μg/kg, 2 LD 50, IM), followed by therapy consisting of atropine methyl nitrate (AMN) (4 mg/kg, IM) 1 min following soman exposure. S-KET, with AS (10 mg/kg), was then administered IM at different times after the onset of seizures, starting at 1 h post-soman exposure. The protective efficacy of S-KET proved to be comparable to KET against lethality and SRBD, but at doses two to three times lower. As with KET, delaying treatment by 2 h post-poisoning greatly reduced efficacy. Conditions that may have led to an increased S-KET brain concentration (increased doses or number of injections, adjunct treatment with the oxime HI-6) did not prove to be beneficial. In summary, these observations confirm that ketamine, either racemic or S-KET, in association with AS and possibly other drugs, could be highly effective in the delayed treatment of severe soman intoxication.

Efficacy of the ketamine–atropine combination in the delayed treatment of soman-induced status epilepticus

Nerve agent poisoning is known to induce full-blown seizures, seizure-related brain damage (SRBD), and lethality. Effective and quick management of these seizures is critical. In conditions of delayed treatment, presently available measures are inadequate calling for optimization of therapeutic approaches. The effects of ketamine/atropine sulfate (KET/AS) combinations were thus assessed as potential valuable delayed therapy in soman-poisoned male guinea pigs. Animals received pyridostigmine (26 μg/kg, i.m.) 30 min before soman (62 μg/kg, i.m.) followed by therapy consisting of atropine methyl nitrate (4 mg/kg) 1 min later. KET was then administered i.m. at different times after the onset of seizures, starting at 30 min post-poisoning. KET was always injected with atropine sulfate, itself given at a dose that was unable to modify seizures (2 to 10 mg/kg). Different treatment schemes (dose and time of injection) were evaluated. Sub-anesthetic doses of KET (10 mg/kg) could prevent lethality and stop ongoing seizures only when administered 30 min after challenge. An extended delay before treatment (up to 2 h) called for an increase in KET dose (up to 60 mg/kg three times), thus reaching anesthetic levels but without the need of any ventilation support. KET proved effective in stopping seizures, highly reducing SRBD and allowing survival with a progressive loss of efficacy when treatment was delayed beyond 1 h post-challenge. Preliminary results suggest that association with the benzodiazepine midazolam (1 mg/kg) might be interesting when treatment is initiated 2 h after poisoning, i.e., when KET efficacy is dramatically reduced. All in all, these observations suggest that KET, in association with atropine sulfate and possibly other drugs, may be highly effective in the delayed treatment of severe soman intoxication.

The recurrent mossy fiber pathway of the epileptic brain.

The dentate gyrus is believed to play a key role in the pathogenesis of temporal lobe epilepsy. In normal brain the dentate granule cells serve as a high-resistance gate or filter, inhibiting the propagation of seizures from the entorhinal cortex to the hippocampus. The filtering function of the dentate gyrus depends in part on the near absence of monosynaptic connections among granule cells. In humans with temporal lobe epilepsy and in animal models of temporal lobe epilepsy, dentate granule cells form an interconnected synaptic network associated with loss of hilar interneurons. This recurrent mossy fiber pathway mediates reverberating excitation that can reduce the threshold for granule cell synchronization. Factors that augment activity in this pathway include modest increases in [K+]o; loss of GABA inhibition; short-term, frequency-dependent facilitation (frequencies of 1-2 Hz); feedback activation of kainate autoreceptors; and release of zinc from recurrent mossy fiber boutons. Factors that diminish activity include short-term, frequency-dependent depression (frequencies < 1 Hz); feedback activation of type II metabotropic glutamate receptors; and the potential release of GABA, neuropeptide Y, adenosine, and dynorphin from recurrent mossy fiber boutons. The axon sprouting and reactive synaptogenesis that follow seizure-related brain damage can also create or strengthen recurrent excitation in other brain regions. These changes are expected to facilitate participation of these regions in seizures. Thus, reactive processes that are often considered important for recovery of function after most brain injuries probably contribute to neurological dysfunction in epilepsy.

Local cerebral blood flow during lithium-pilocarpine seizures in the developing and adult rat: role of coupling between blood flow and metabolism in the genesis of neuronal damage.

Coupling between local cerebral blood flow and local cerebral metabolic rate for glucose is involved in the pathogenesis of epilepsy-related neuronal damage in the adult brain; however, its role in the immature brain is unknown. Lithium-pilocarpine-induced status epilepticus is associated with extended damage in adult rats, mostly in the forebrain limbic areas and thalamus, whereas damage was moderate in 21-day-old rats (P21) or absent in P10 rats. The quantitative autoradiographic [14C]iodoantipyrine technique was applied to measure the consequences of lithium-pilocarpine status epilepticus on local cerebral blood flow. In adult and P21 rats, local cerebral blood flow rates increased by 50% to 400%; the highest increases were recorded in regions showing damage in adults. At P10, local cerebral blood flow rates decreased by 40% to 60% in most areas, except in some forebrain regions showing no change during status epilepticus. In areas injured when status epilepticus was induced in adults, a strong hypermetabolism (Fernandes et al., 1999) not matched by comparable local cerebral blood flow increases was present in rats of all ages, whereas in damage-resistant areas, local cerebral metabolic rate for glucose and local cerebral blood flow remained coupled in the three age groups. Thus, the level of coupling between blood flow supply and metabolism is not involved in seizure-related brain damage in the developing brain, which appears to be resistant to the consequences of such a mismatch.

An Animal Model of Nonconvulsive Status Epilepticus: A Contribution to Clinical Controversies

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.

GM1 monosialoganglioside pretreatment protects against soman-induced seizure-related brain damage.

The effects of GM1 monosialoganglioside pretreatment on brain damage resulting from soman-induced seizure activity were examined in this study. Male Sprague-Dawley rats were infused with GM1 via an osmotic minipump connected through a permanent cannula implanted intracerebroventricularly and challenged with soman (83 micrograms/kg, i.e., 1.25 x LD50) 4 d after initiation of GM1 infusion. Electrocorticographic recordings were monitored via indwelling cortical electrodes. Twenty-seven hours after soman administration, anesthetized rats were euthanized via transcardial perfusion with buffered paraformaldehyde. Brains were processed for hematoxylin and eosin (H&E), cresyl violet (CV), and acetylcholinesterase (AChE) histochemistry, and glial fibrillary acidic protein (GFAP) and microtubule-associated protein 2 (MAP2) immunohistochemistry. All soman-challenged rats not infused with GM1 (n = 14) developed status epilepticus (SE).

Induction of the dual specificity phosphatase PAC1 in rat brain following seizure activity.

Recurrent seizure activity leads to delayed neuronal death as well as to inflammatory responses involving microglia in hippocampal subfields CA1, CA3 and CA4. Since mitogen activated protein (MAP) kinases control neuronal apoptosis and trigger generation of inflammatory cytokines, their activation state could determine seizure-related brain damage. PAC1 is a dual specificity protein phosphatase inactivating MAP kinases which we have found to be undetectable in normal brain. Despite this, kainic acid-induced seizure activity lead to rapid (approximately 3 h) but transient appearance of PAC1 mRNA in granule cells of the dentate gyrus as well as in pyramidal CA1 neurons. This pattern changed with time and after 2-3 days PAC1 was induced in dying CA1 and CA3 neurons. At this time PAC1 mRNA was also expressed in white matter microglia as well as in microglia invading the damaged hippocampus. PAC1 may play an important role controlling MAP kinase involvement in both neuronal death and neuro-inflammation following excitotoxic damage.

MEMANTINE EFFECTS ON SOMAN-INDUCED SEIZURES AND SEIZURE-RELATED BRAIN DAMAGE

Excitatory am ino acids have been implicated in the seizures and seizure-related brain damage (SRBD) resulting from soman intoxication. Memantine (Mem) is an open channel blocker of the glutaminergic NMDA receptor subtype reported to have anticonvulsant activity against soman. The aim of the present experiments was to evaluate the effect of Mem on electrographic seizure activity (EGSA) and SRBD produced by soman. Male rats were implanted with cortical screw electrodes for recording bipolar electrocorticogram s and allowed to recover for 5-7 days. EGSA was precipitated by injecting soman (180 mug/ kg, SC) 30 m in after HI6 (125 mg/ kg, IP) pretreatm ent. Saline, Mem (18 mg/ kg), atropine (A;10 mg/ kg), or Mem + A was adm inistered IP either 5 or 40 m in after the onset of EGSA. Saline-treated anim als displayed limbic convulsive behaviors and EGSA lasting for 4 or m ore hours. Mem or Mem + A suppressed behavioral signs of soman intoxication, but did not terminate EGSA or prevent SRBD. These results indicate that soman-induced seizures are not ameliorated by Mem or Mem + A, and that the presence or absence of som an-induced seizures cannot be verified reliably through behavioral observations alone.

Microtubule-associated protein 2 (MAP-2): a sensitive marker of seizure-related brain damage

We have assessed the efficacy of MAP-2 immunohistochemistry as a marker of seizure-related brain damage and its suitability for quantitation of the damage using densitometric and morphometric image analysis. Seizures were produced in rats by administration of 1.5 LD 50 soman, an irreversible AChE inhibitor. Our results demonstrate that neuronal damage, assessed using hematoxylin and eosin, and cresyl violet staining, was colocalized on adjacent serial sections with clearly demarcated reductions in MAP-2 staining. The most severely damaged brain regions were devoid of MAP-2 staining. Reductions in MAP-2 immunostaining were found to be exceptionally well suited for quantitation using densitometric and morphometric image analysis. This study represents the first demonstration of seizure-induced excitotoxic alterations in MAP-2.

Quisqualate Injection into the Nucleus Basalis Magnocellularis Produces Seizure-Related Brain Damage That Is Prevented by MK-801

Quisqualate (Quis) and other excitotoxins such as ibotenate and N-methyl aspartate, have been used to destroy neurons in the area of the nucleus basalis magnocellularis (NBM) in order to study the relationship between loss of cholinergic neurons in the basal forebrain and various behavioral deficits, including learning and memory impairments. The results of several studies suggest that although Quis NBM lesions may produce greater depletions in cortical choline acetyltransferase levels than ibotenate lesions, the learning/memory deficits tend to be milder following Quis lesions. In these studies, it was often assumed that the lesions induced by Quis were restricted to the local vicinity of the injection. However, in the present study, we found that an injection of Quis into the NBM/substantia inominata (SI) region often induces limbic seizures and disseminated brain damage. Specifically, we found that an injection of Quis into the NBM/SI area of female rats at a dose (120 umol) used by others in previous behavioral studies produced massive damage in areas distant from the lesion site, particularly in the amygdala and piriform cortex. This disseminated damage occurred in 50% of the rats treated with Quis, was typically more severe than damage at the injection site, and was often accompanied by equally severe "mirror" lesions in the contralateral amygdala and piriform cortex. Injecting rats with MK-801 (1 mg/kg) 30 min before the Quis injection protected against the disseminated damage. These data underscore the need for careful histological evaluation of excitotoxic lesions and for caution in interpreting the relationship between altered transmitter markers and learning/memory impairment seen following these lesions.