Damage induced by systemic kainic acid in rats is dependent upon seizure activity--a behavioral and morphological study

Kainic acid (KA) was injected intraperitoneally into rats at a dose (9 mg/kg) which produced status epilepticus in approximately 50% of the animals. Rats were categorized into groups that displayed status epilepticus, partial seizures or no effect in the 4 hr following kainic acid injection. Behavioral and morphological changes were characterized for each group. Rats that were not affected by kainic acid were indistinguishable from a saline-injected control group. When sacrificed 4 hr after treatment, rats displaying partial seizures showed morphological changes similar to, but less severe than, those exhibiting status epilepticus. Additional groups were tested and sacrificed 7 days (d) after treatment. Rats from the limited seizure group showed little behavioral or morphological response, while animals from the status epilepticus group had marked behavioral deficits and severe lesions. The tissue damage and its distribution were similar to lesions observed after seizures induced by other convulsants, and in spontaneously epileptic dogs. These results suggest that the extent of damage resulting from systemic administration of KA is dependent on the extent of seizure activity, which may in turn be related to the influence of kainic acid and other excitatory amino acids on the limbic system.     

Mitochondrial dysfunction and ultrastructural damage in the hippocampus during kainic acid-induced status epilepticus in the rat

PURPOSE: Prolonged and continuous epileptic seizure (status epilepticus) results in cellular changes that lead to neuronal damage. We investigated whether these cellular changes entail mitochondrial dysfunction and ultrastructural damage in the hippocampus, by using a kainic acid (KA)-induced experimental status epilepticus model. METHODS: In Sprague-Dawley rats maintained under chloral hydrate anesthesia, KA (0.5 nmol) was microinjected unilaterally into the CA3 subfield of the hippocampus to induce seizure-like hippocampal EEG activity. The activity of key mitochondrial respiratory chain enzymes in the dentate gyrus (DG), or CA1 or CA3 subfield of the hippocampus was measured 30 or 180 min after application of KA. Ultrastructure of mitochondria in those three hippocampal subfields during KA-induced status epilepticus also was examined with electron microscopy. RESULTS: Microinjection of KA into the CA3 subfield of the hippocampus elicited progressive build-up of seizure-like hippocampal EEG activity. Enzyme assay revealed significant depression of the activity of nicotinamide adenine dinucleotide cytochrome c reductase (marker for Complexes I+III) in the DG, or CA1 or CA3 subfields 180 min after KA-elicited temporal lobe status epilepticus. Conversely, the activities of succinate cytochrome c reductase (marker for Complexes II+III) and cytochrome c oxidase (marker for Complex IV) remained unaltered. Discernible mitochondrial ultrastructural damage, varying from swelling to disruption of membrane integrity, also was observed in the hippocampus 180 min after hippocampal application of KA. CONCLUSIONS: Our results demonstrated that dysfunction of Complex I respiratory chain enzyme and mitochondrial ultrastructural damage in the hippocampus are associated with prolonged seizure during experimental temporal lobe status epilepticus.     

Differential Susceptibility to Seizures Induced by Systemic Kainic Acid Treatment in Mature DBA/2J and C57BLl6J Mice

Summary: Mature DBA/2J (D2) and C57BL/6J (B6) mice aged P.10 weeks were studied to determine susceptibility to behavioral seizures induced by kainic acid (KA) and the possible influence exerted by differences in metabolism and blood-brain barrier (BBB) transport. Mice were observed for 4 h after subcutaneous (s.c.) KA injection. Behavioral seizure parameters included latency to first seizure (clonus), latency to tonic/clonic seizure, and latency to status epilepticus (SE). At a KA dose of 25 mg/kg, 80% of D2 mice exhibited tonicklonic seizures, whereas all B6 mice remained seizure-free. At 30 mg/kg, tonic/ clonic seizures were observed in 100% of D2 mice and 25% of B6 mice. Of D2 mice exhibiting at least one clonic seizure in response to KA at a dose of 25 mg/kg, 50% entered SE and eventually died. Administration of [³H]KA (6.6 × 10⁶dpm) at doses of 25 mg/kg (convulsive) or 11.1 μ/g (nonconvulsive) to mice of both strains resulted in similar levels of radioactivity in cortex, hippocampus, and cerebellum 30 and 60 min after injection. Bioconversion of [³H]KA to a radiolabeled brain metabolite in vivo could not be documented in mice from either strain. Results confirm previously reported differences between D2 and B6 mice in their relative susceptibility to seizures induced by systemic KA administration and suggest that these differences are not related to strain-specific variation in metabolism or BBB transport of KA. Further studies of these two strains of mice may be useful for investigating genetic influences upon seizure susceptibility.     

Kainic acid seizures in the developing brain: status epilepticus and spontaneous recurrent seizures.

Acute and chronic effects of seizures induced by intraperitoneal (i.p.) injection of kainic acid (KA) were studied in developing rats (postnatal days (P) 5, 10, 20, 30, and adult 60). For 3 months following KA-induced status epilepticus, spontaneous recurrent seizure (SRS) occurrence was quantified using intermittent video monitoring. Latency to generalized seizures was then tested using flurothyl, and brains were histologically analyzed for CA3 lesions. In P5-10 rats, KA caused generalized tonic-clonic ('swimming') seizures. SRS did not develop, and there was no significant difference between control and KA-treated rats in latency to flurothyl-induced seizures. In contrast, rats P20 and older exhibited limbic automatisms followed by limbic motor seizures which secondarily generalized. Incidence and frequency of SRS increased with age. P20-30 rats with SRS had shorter latencies to flurothyl seizures than did KA-treated P20-30 rats without SRS or controls. KA-treated P60 rats (with or without SRS) had shorter latencies than controls to flurothyl seizure onset. SRS in P60 rats occurred sooner after KA than in P20-30 rats. CA3 lesions were seen in P20-60 rats with and without SRS, but not in P5-10 rats. These data suggest that there are developmental differences in both acute and chronic responses to KA, with immature animals relatively protected from the long-term deleterious effects of this convulsant.     

Chronic bilateral stimulation of the anterior thalamus of kainate-treated rats increases seizure frequency

PURPOSE: Electrical stimulation of the anterior nucleus of the thalamus (ANT) is receiving increased attention as a novel means of controlling intractable epilepsy, and has entered human clinical trial. Animal data supporting the anticonvulsant benefit of ANT stimulation, however, has been obtained from acute chemoconvulsant models of epilepsy rather than models of chronic epilepsy with spontaneous seizures. It is unknown whether ANT stimulation is effective in models of chronic epilepsy. METHODS: Bilateral ANT stimulation was evaluated in rats with chronic epilepsy following acute status epilepticus (SE) produced by systemic kainic acid (KA) administration. The evolution of epilepsy following KA SE and the effects of ANT stimulation were monitored by continuous video-EEG. RESULTS: Following KA SE, most rats have 2-8 seizures per day, and the average seizure rate increases over time, doubling over the course of 14 weeks. Behavioral seizure severity, after the initial development of epilepsy, remains stable. Seizure frequency during ANT stimulation was 2.5 times the baseline seizure frequency. In some cases stimulation triggered seizures were observed. The effects of stimulation were specific to the ANT. Stimulation applied to electrodes placed outside the ANT did not significantly worsen seizure frequency. CONCLUSIONS: ANT stimulation exacerbated seizure frequency in rats with chronic epilepsy following kainate status epilepticus.     

Resistance of immature hippocampus to morphologic and physiologic alterations following status epilepticus or kindling.

Seizures in adult rats result in long-term deficits in learning and memory, as well as an enhanced susceptibility to further seizures. In contrast, fewer lasting changes have been found following seizures in rats younger than 20 days old. This age-dependency could be due to differing amounts of hippocampal neuronal damage produced by seizures at different ages. To determine if there is an early developmental resistance to seizure-induced hippocampal damage, we compared the effects of kainic acid (KA)-induced status epilepticus and amygdala kindling on hippocampal dentate gyrus anatomy and electrophysiology, in immature (16 day old) and adult rats. In adult rats, KA status epilepticus resulted in numerous silver-stained degenerating dentate hilar neurons, pyramidal cells in fields CA1 and CA3, and marked numerical reductions in CA3c pyramidal neuron counts (-57%) in separate rats. Two weeks following the last kindled seizure, some, but significantly less, CA3c pyramidal cell loss was observed (-26%). Both KA status epilepticus and kindling in duced mossy-fiber sprouting, as evidenced by ectopic Timm staining in supragranular layers of the dentate gyrus. In hippocampal slices from adult rats, paired-pulse stimulation of perforant path axons revealed a persistent enhancement of dentate granule-cell inhibition following KA status epilepticus or kindling. While seizures induced by KA or kindling in 16-day-old rats were typically more severe than in adults, the immature hippocampus exhibited markedly less KA-induced cell loss (-22%), no kindling-induced loss, no detectable synaptic rearrangement, and no change in dentate inhibition. These results demonstrate that, in immature rats, neither severe KA-induced seizures nor repeated kindled seizures produce the kind of hippocampal damage and changes associated with even less severe seizures in adults. The lesser magnitude of seizure-induced hippocampal alterations in immature rats may explain their greater resistance to long-term effects of seizures on neuronal function, as well as future seizure susceptibility. Conversely, hippocampal neuron loss and altered synaptic physiology in adults may contribute to increased sensitivity to epileptogenic stimuli, spontaneous seizures, and behavioral deficits.     

Limbic seizures without brain damage after injection of low doses of kainic acid into the amygdala of freely moving rats

Kainic acid (KA, 8-15 ng) was injected into the amygdala of conscious freely moving rats via chronically implanted fused silica cannulas. At 15-25 min after the injection, most rats suffered a limbic seizure attack of short duration, consisting of mastication, forelimb clonus, and raising on hind limbs, behaviorally indistinguishable from kindled seizures. Typically, the attack was followed by stereotypies, intense exploration, and by 1 or 2 more attacks. About 60 min after the injection, most rats appeared normal again and histopathological changes in their brains did not exceed those seen in vehicle-injected rats. In 3 cases, however, recurrent seizures culminated in behavioral status epilepticus 60-90 min after the injection. The status epilepticus was stopped by i.p. injection of diazepam (10 mg/kg) after a duration of 10 min (1 case) and 30 min (2 cases), respectively. After 10 min status epilepticus, we observed marginal neuronal damage with slight gliosis in both hippocampi (CA3 and CA1); after 30 min, hippocampal histopathology was more pronounced, with additional necrosis of the ipsilateral piriform cortex. After 0.8 microgram KA, a hundredfold higher dose, the incidence of limbic seizures during the first 40 min was not significantly higher (9/12) than after the lower KA doses (13/19). However, a significantly higher proportion of rats exhibited long-lasting seizure activity, associated with confluent destruction of CA3 pyramidal cells and additional seizure-related brain damage. Our results show that limbic motor seizures do not inevitably lead to histopathological changes in the brain, provided they do not culminate in a state of permanent seizure activity.     

Effects of AT1 receptor antagonism on kainate-induced seizures and concomitant changes in hippocampal extracellular noradrenaline,serotonin, and dopamine levels in Wistar-Kyoto and spontaneously hypertensive rats

In the management of epilepsy, AT₁ receptor antagonists have been suggested as an additional treatment strategy. A hyperactive brain angiotensin (Ang) II system and upregulated AT₁ receptors are implicated in the cerebrovascular alterations in a genetic form of hypertension. Uncontrolled hypertension could also, in turn, be a risk factor for a seizure threshold decrease and development of epileptogenesis. The present study aimed to assess the effects of the selective AT₁ receptor antagonist ZD7155 on kainic acid (KA)-induced status epilepticus (SE) development and accompanying changes in the hippocampal extracellular (EC) neurotransmitter levels of noradrenaline (NAD), serotonin (5-HT), and dopamine (DA) in spontaneously hypertensive rats (SHRs) and their parent strain Wistar-Kyoto (WKY) rats, since monoamines are well-known neurotransmitters involved in mechanisms of both epilepsy and hypertension. Status epilepticus was evoked in freely moving rats by a repetitive intraperitoneal (i.p.) administration of KA in subconvulsant doses. In the treatment group, ZD7155 (5 mg/kg i.p.) was coadministered with the first KA injection. Spontaneously hypertensive rats exhibited higher susceptibility to SE than WKY rats, but the AT₁ receptor antagonist did not alter the development of SE in SHRs or in WKY rats. In vivo microdialysis demonstrated significant KA-induced increases of the hippocampal NAD and DA levels in SHRs and of NAD, 5-HT, and DA in WKY rats. Although SHRs developed more severe seizures while receiving a lower dose of KA compared to WKY rats, AT₁ receptor antagonism completely prevented all KA-induced increases of hippocampal monoamine levels in both rat strains without affecting seizure development per se. These results suggest a lack of direct relationship between KA-induced seizure susceptibility and adaptive changes of hippocampal NAD, 5-HT, and DA levels in the effects of ZD7155 in WKY rats and SHRs.     

Akt Inhibitor Perifosine Prevents Epileptogenesis in a Rat Model of Temporal Lobe Epilepsy

Accumulating data have revealed that abnormal activity of the mTOR (mammalian target of rapamycin) pathway plays an important role in epileptogenesis triggered by various factors. We previously reported that pretreatment with perifosine, an inhibitor of Akt (also called protein kinase B), abolishes the rapamycin-induced paradoxical increase of S6 phosphorylation in a rat model induced by kainic acid (KA). Since Akt is an upstream target in the mTOR signaling pathway, we set out to determine whether perifosine has a preventive effect on epileptogenesis. Here, we explored the effect of perifosine on the model of temporal epilepsy induced by KA in rats and found that pretreatment with perifosine had no effect on the severity or duration of the KA-induced status epilepticus. However, perifosine almost completely inhibited the activation of p-Akt and p-S6 both acutely and chronically following the KA-induced status epilepticus. Perifosine pretreatment suppressed the KA-induced neuronal death and mossy fiber sprouting. The frequency of spontaneous seizures was markedly decreased in rats pretreated with perifosine. Accordingly, rats pretreated with perifosine showed mild impairment in cognitive functions. Collectively, this study provides novel evidence in a KA seizure model that perifosine may be a potential drug for use in anti-epileptogenic therapy.     

Cardiac electrographic and morphological changes following status epilepticus: Effect of clonidine

PurposeStatus epilepticus has been increasingly associated with cardiac injury in both clinical and animal studies. Our group has previously shown that excitotoxic seizure induction results in the formation of ischaemic myocardial infarcts and loss of cardiac haemodynamic function. We hypothesised that attenuation of cardiac sympathetic/parasympathetic balance with a central presynaptic α₂ agonist, clonidine, can reduce the development of interictal ECG and ventricular morphological changes resulting from kainic acid (KA; 10 mg/kg) induced status epilepticus in a conscious rat model.MethodsUsing simultaneous ECG and electrocorticogram (ECoG) radiotelemetry, animals were randomised into saline controls, saline-pretreated KA and clonidine (100 μg/kg, b.i.d.)-pretreated KA groups. Baseline ECG, ECoG and behavioural score recordings were acquired in conscious animals for 2 h post-KA administration.ResultsBradycardia and low level seizure activity occurred immediately following KA administration. As seizure activity (ECoG spiking and high level seizure behavioural scoring) progressively increased, tachycardia developed. Both QTc prolongation and T wave amplitude were transiently but significantly increased. Clonidine treatment attenuated seizure activity, increased the latency to onset of seizure behaviour and reduced seizure-induced changes in heart rate, QTc interval, and T wave amplitude. Histological examination of the ventricular myocardium revealed hypercontraction band necrosis, inflammatory cell infiltration, and oedema at 48 h post-KA. In contrast, clonidine-treatment in seizure animals preserved tissue integrity and structure.ConclusionThese results demonstrate that KA-induced seizures are associated with altered ECG activity and cardiac structural pathology. We suggest that pharmacological modulation of sympathetic/parasympathetic activity in status epilepticus provides a promising therapeutic approach to reduce seizure-induced cardiomyopathy.     

Status epilepticus results in reversible neuronal injury in infant rat hippocampus: novel use of a marker

Despite ready induction of severe limbic status epilepticus by systemic kainic acid (KA) in infant rats, excitotoxic neuronal injury has not been observed. The mechanisms of this resistance of the immature hippocampus to excitotoxicity are unknown. Acid fuchsin stain has been used as a marker of irreversibly injured neurons in the adult brain. We speculated that the dye might map reversibly injured neurons in the infant. Subsequent to KA-induced status epilepticus in 11-day-old rats, acid fuchsin stain was evident in the hippocampal CA3, CA1, dentate gyrus and hilus by 24 h, peaked at 48 h and disappeared by 6 days, without evidence for neuronal loss. Acid fuchsin may be a useful tool for delineating the distribution of reversibly injured immature neurons in experimental seizure paradigms.     

Experimental limbic seizure status epilepticus and focus resection in cats

Status epilepticus is a neurological emergency and refractory one often resulted in neurological damage or death. Since the basic mechanisms of status epilepticus was not fully understood, a surgical treatment was not attempted until now. In the present study, a surgical resection of the epileptic focus was made in experimentally induced limbic status epilepticus and influences of the surgery upon status epilepticus was discussed. Limbic status epilepticus was induced by means of kainic acid (KA) microinjection into unilateral amygdala in cats and effects of focus resection upon limbic seizure status were studied. Ten adult cats were stereotaxically operated on under pentobarbital anesthesia. Bipolar electrodes were placed in bilateral amygdala and hippocampus. An injection cannula, designed for kainic acid injection, was placed in the left amygdala. The cats were then divided into two groups. Group A (5 cats) received 0.5 microgram of KA injection into the amygdala resulted in mild limbic status. Two of them were controls and 3 of them received amygdalotomy after induction of the limbic seizure status. Group B (5 cats) received 2.0 micrograms KA injection resulted in severe limbic status. Moreover, independent spontaneous seizure activities were observed in the ipsilateral hippocampus. Two of them were controls and 3 of them were operated on. After amygdalotomy, limbic seizure stopped in the operated cats of Group A. In the operated cats of Group B, repeated seizures in the epileptogenic focus (amygdala) was completely suppressed, however, spontaneous seizures of the ipsilateral hippocampus persisted even after the surgery.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kainic acid-induced substantia nigra seizure in rats: behavior, EEG and metabolism

RATIONALE: In order to clarify the role of substantia nigra pars reticulata (SNr) upon the development of epileptic seizure, kainic acid (KA) was injected into a unilateral SNr. MATERIALS AND METHODS: Wistar rats weighing 250-350 g were used. A stainless-steel cannula and depth electrode were inserted stereotaxically into the left substantia nigra pars reticulata (SNr). At 7 days after surgery, 1.0 microg of KA was injected into the left SNr. Experiment 1: In eight rats, behavior and electroencephalograms (EEG) were continuously recorded for about 30 h, and intermittently monitored following 1 month. Experiment 2: Two hours after KA injection into SNr, rats demonstrated status epilepticus. Then, 100 microCi/kg of [(14)C]2-deoxyglucose (2-DG) was intravenously injected in seven rats, and the rats were processed for autoradiographic study. RESULTS: Changes in behavior and EEG: On EEG, a secondary generalized seizure status was observed at about 70 min after KA injection. In video, limbic seizure manifestations such as salivation were observed as a initial symptom and followed by rolling and generalized tonic seizures. [(14)C]deoxyglucose autoradiographic study demonstrated increased local cerebral glucose metabolism in the medial and lateral septal nucleus, substantia nigra, hippocampus, parietal cortex, piriform cortex, medial and lateral geniculate nucleus, anterodorsal, lateral and ventral nucleus of the thalamus, amygdala and midbrain reticular formation. SUMMARY: The result suggested that the substantia nigra played an important role in the secondary generalization in the substantia nigra seizure model due to the decreased function of the GABAergic projection system induced by an excessive epileptic excitation of SNr.     

A short episode of seizure activity protects from status epilepticus-induced neuronal damage in rat brain

Kainic acid (KA)-induced status epilepticus (SE) in adult rats results in extensive neuronal damage throughout the limbic system and the loss of selectively vulnerable neuronal populations, particularly CA3 neurons. We investigated the effects of a short episode of seizure activity on neuronal death elicited by a subsequent prolonged SE episode. A short episode of seizure activity was produced by sub-cutaneous (s.c.) injection of KA followed after 1 h by pentobarbital administration. Twenty-four hours later, KA was administered again, and animals were sacrificed 3 days later. Neuronal damage was estimated by visual analysis of neuronal density. Our results show that a short episode of seizure activity did not produce neuronal damage but almost completely protected vulnerable neurons from KA-induced neuronal damage. These results extend to epileptic tolerance the notion of tolerance previously described in the case of ischemia.     

Hypothermia reduces brain edema, spontaneous recurrent seizure attack, and learning memory deficits in the kainic acid treated rats

Aims: It is unknown whether hypothermia can disrupt the progress of epileptogenesis. The present study aimed to determine the effect of hypothermia on brain edema and epileptogenesis and to establish whether brain edema is associated with epileptogenesis after severe status epilepticus (SE). Methodology: Rats were injected with a single dose of Kainic acid (KA) to produce either chronic epileptic rats (rats with spontaneous recurrent seizure, SRS) or rats without spontaneous recurrent seizure (no‐SRS rats). A second KA injection was used to induce SE in SRS rats and in no‐SRS rats. The number of SRS was counted and the brain edema induced by SE was assessed by brain water content measurement. The cognitive function was assessed by the radial‐arm maze (RAM) test. Results: A second KA injection resulted in brain edema that was more severe in SRS rats than in no‐SRS rats. After second injection of KA, hypothermia treatment attenuated the KA induced brain edema and reduced the SRS attack in SRS rats. Additionally cognitive function was better in hypothermia‐treated SRS rats than in nomothermia treated SRS rats 1 month after the second KA injection. Conclusions: Hypothermia treatment immediately after SE not only exhibited protective effects against the chronic spontaneous recurrent convulsant seizures but also improved cognitive function. These antiepileptogenic properties of hypothermia may be related to its attenuating effect on brain edema induced by SE. They therefore suggest that brain edema may be involved in the progress of epileptogenesis.     

Seizures in the intrahippocampal kainic acid epilepsy model: characterization using long‐term video‐EEG monitoring in the rat

Objective – Intrahippocampal injection of kainic acid (KA) in rats evokes a status epilepticus (SE) and leads to spontaneous seizures. However to date, precise electroencephalographic (EEG) and clinical characterization of spontaneous seizures in this epilepsy model using long‐term video‐EEG monitoring has not been performed. Materials and Methods – Rats were implanted with bipolar hippocampal depth electrodes and a cannula for the injection of KA (0.4 μg/0.2 μl) in the right hippocampus. Video‐EEG monitoring was used to determine habitual parameters of spontaneous seizures such as seizure frequency, severity, progression and day–night rhythms. Results – Spontaneous seizures were detected in all rats with 13 out of 15 animals displaying seizures during the first eight weeks after SE. A considerable fraction (35%) of the spontaneous seizures did not generalize secondarily. Seizure frequency was quite variable and the majority of the KA‐treated animals had less than one seizure per day. A circadian rhythm was observed in all rats that showed sufficient seizures per day. Conclusions – This study shows that the characteristics of spontaneous seizures in the intrahippocampal KA model display many similarities to other SE models and human temporal lobe epilepsy.     

Seizure-induced memory impairment is reduced by choline supplementation before or after status epilepticus

Prenatal choline supplementation can protect rats against cognitive deficits induced by status epilepticus induced by the cholinergic agent pilocarpine [J. Neurosci. 20 (2000) 1]. In the present day, we have extended this novel finding by investigating the effects of pre- and postnatal choline supplementation in memory deficits associated with status epilepticus induced with kainic acid (KA). In the first experiment pregnant rats received a normal, choline-supplemented, or choline deficient diet starting on the 11th day of gestation and continuing until postnatal (P) 7. At P42, rats were given a convulsant dosage of KA. Two weeks following the KA-induced status epilepticus rats underwent testing of visual-spatial memory using the Morris water maze test. Rats receiving supplemental choline performed better in the water maze than the deficient and control groups. Moreover, the activity of hippocampal choline acetyltransferase was 18% lower in the choline deficient animals as compared with the other two groups. In the second experiment we administered KA to P35 rats that had been given a normal diet. Following the status epilepticus the rats were given a choline-supplemented or control diet for 4 weeks and then tested in the water maze. Rats receiving choline supplementation performed far better than rats receiving a regular diet. This study demonstrates that choline supplementation prior to or following KA-induced status epilepticus can protect rats from memory deficits induced by status epilepticus.     

7-Nitroindazole, a selective inhibitor of nNOS, increases hippocampal extracellular glutamate concentration in status epilepticus induced by kainic acid in rats

The glutamate extracellular concentration is controlled by metabolic and neuronal pathways via release and uptake mechanisms. Stimulation of glutamate receptors induces neuronal nitric oxide (NO) release, which in turn modulates glutamate transmission. In this study, the influence of neuronally derived NO on hippocampal glutamate extracellular concentration was investigated in conditions of intense metabolic activation, i.e., during status epilepticus induced by systemic kainic acid (KA). Glutamate, arginine and citrulline concentrations were measured by microdialysis coupled to HPLC. Experiments were performed in conscious rats implanted with a microdialysis probe within the hippocampal CA3 area. Three groups were used: (1) rats treated with KA i.p. (12 mg/kg) and vehicle locally, via the microdialysis probe (n = 9); (2) rats given KA i.p. and a selective inhibitor of neuronal NO synthase, 7-nitroindazole (7-NI, 1.25 mM) locally (n = 13); (3) rats treated with saline i.p. and 7-NI locally (n = 7). Infusion of 7-NI or vehicle was performed throughout the second hour of status epilepticus. In groups 1 and 3, no significant modifications of extracellular glutamate, arginine and citrulline concentrations were measured. In group 2, the local application of 7-NI in the hippocampus during status epilepticus significantly increased extracellular glutamate and arginine concentrations, whereas citrulline concentration remained constant. The concomitant increases of extracellular glutamate and arginine concentrations under local 7-NI perfusion in seizure conditions, suggest that glutamate and arginine are linked in a common metabolic pathway and/or that glutamate is involved in the cross-talk between glia and neurons. A cerebrovascular effect of 7-NI which triggers glutamate release may also occur.     

Effects of drugs on the initiation and maintenance of status epilepticus induced by administration of pilocarpine to lithium-pretreated rats

The ability of various drugs to prevent the onset of status epilepticus induced by administration of the muscarinic agonist, pilocarpine, to lithium-pretreated rats was determined. Motor limbic seizures and status epilepticus occurred in 100% of rats administered pilocarpine (30 mg/kg, s.c.) 20 h after pretreatment with lithium (3 meq/kg, i.p.). The latency to spike activity and to status epilepticus was 20 +/- 1 min and 24 +/- 1 min, respectively. Atropine, diazepam, phenytoin, carbamazepine, phenobarbital, paraldehyde, and L-phenylisopropyladenosine (L-PIA) prevented all phases of seizure activity induced by lithium/pilocarpine treatment. The initiation of status epilepticus was significantly prolonged by pretreatment with sodium valproate. These findings indicate that the seizures induced by administration of lithium and pilocarpine accurately model generalized tonic-clonic epilepsy. The anticonvulsant activity of L-PIA was prevented by prior treatment with the adenosine antagonist, theophylline. The latency to spike and seizure activity was decreased by theophylline, indicating that endogenous adenosine may have a tonic inhibitory influence on cholinergic neurons. Atropine, diazepam, phenobarbital, phenytoin, sodium valproate, L-PIA, and carbamazepine did not interrupt seizure activity when administered 60 min after pilocarpine (approximately 35 min after initiation of status epilepticus). When rats were administered paraldehyde at this time, status epilepticus was rapidly terminated and all rats survived. Thus, status epilepticus induced by lithium and pilocarpine provides a seizure model that is not responsive to conventional anticonvulsants.     

Zonisamide: Electrophysiological and Metabolic Changes in Kainic Acid-Induced Limbic Seizures in Rats

Summary: We studied the pharmacological mechanism of zonisamide (ZNS) using an electrophysiological and autoradiographical method in a limbic seizure model in rats. Limbic seizure status epilepticus was induced by a unilateral microinjection of kainic acid (KA) into the amygdala. Initially, observed seizures were limited to the side of the injected amygdala and then propagated to bilateral sensorimotor cortex. Eighty minutes after injection, secondarily generalized seizure status epilepticus was induced, with each seizure lasting ∼30 s and recurring every 5 min. ZNS 100 mg/kg was administered intravenously (i.v.) during the generalized seizure. Forty minutes after ZNS administration, epileptic activity was observed only at the KA-injected amygdalar site and spikes were not observed in the bilateral sensorimotor cortex. We studied local cerebral glucose utilization (LCGU) after ZNS or saline administration using an autoradiographical method in the same limbic seizure preparation. In the ZNS group, LCGU decreased in the ipsilateral sensorimotor cortex and hippocampus, whereas in the controls LCGU increased in these structures. On the other hand, ZNS did not suppress the epileptic activity of the primary focus and no decrease in LCGU was observed in the KA-injected amygdala. ZNS inhibited seizure propagation from the epileptogenic focus but did not suppress the epileptic activity of the focus. Our results suggest that ZNS is effective for the treatment of secondarily generalized seizure.