An animal model of generalized nonconvulsive status epilepticus: immediate characteristics and long-term effects

Absence seizures are traditionally believed to have no significant long-term neurological consequences, but few basic scientific studies have examined the effects of absence seizures on neuronal function, especially regarding absence status epilepticus. We developed a model of generalized nonconvulsive status epilepticus (GNCSE) in rats to study behavioral, functional, and histological effects of GNCSE. Using repetitive timed injections of low-dose pentylenetetrazol (PTZ), a state of prolonged behavioral arrest and immobility associated with frequent generalized spike-wave discharges on EEG could be induced for hours, consistent with GNCSE. GNCSE occurred reproducibly in adult rats, but surprisingly not in juvenile rats or adult mice. There was no evidence of pathological damage following GNCSE using Fluoro-Jade B and Cresyl Violet histological methods. Although a transient, subtle deficit in place learning occurred in PTZ-treated rats, there were no long-term behavioral effects of GNCSE on spatial learning or sensorimotor function. However, 1 week after a single episode of GNCSE, there was an increase in absence seizures in response to a repeat dose of PTZ compared to controls. These results indicate that an animal model of GNCSE can be generated and that even in the absence of overt neuronal damage, GNCSE may produce functional changes in neurons that alter electrical excitability of neural circuits.     

Neuronal Firing Pattern Following Amygdaloid Kindling in Unrestrained Rats

Chronic recordings of amygdaloid neurons were performed on freely moving rats following kindling. Satisfactory recordings were obtained from 22 amygdaloid neurons of the contralateral amygdala before, during, and after unilateral kindling. Kindling stimulations were given once per hour. Seven cells disappeared during kindling. Seven cells were recorded during the full course of kindling. These units showed (a) an increase in spontaneous firing, (b) a development of high-frequency bursts (the peak interval of the interval histogram decreased from 18 to 2 ms), and (c) high-frequency firings during spontaneous activity that were similar to the firings recording during afterdischarge.     

Strain differences in pentylenetetrazol-kindling development and subsequent potentiation effects

Rats from two different strains, i.e. Wistar rats and Lister hooded rats, were investigated for their ability to acquire the kindling syndrome. After having received 13 kindling stimulations (injection of pentylenetetrazol), the animals were tested for subsequent alterations in induction and maintenance of hippocampal long-term potentiation (LTP) and, moreover in glutamate binding. It was found that rats from both strains did not differ in the response to the initial injection of pentylenetetrazol (PTZ) and the amplitude of the population spike. This suggests that some aspects of basic central excitability are equivalent. Wistar rats acquired the kindling syndrome rapidly whereas seizure outcome was poor in Lister rats. As regards hippocampal LTP, the population spike was only dramatically increased in Wistar rats after kindling completion. Glutamate binding was not altered in animals from the Lister strain. The results suggest that changes in glutamate binding and the increase in the population spike are characteristic consequences of kindling.     

Neuroprotective effect of nimesulide, a preferential COX-2 inhibitor, against pentylenetetrazol (PTZ)-induced chemical kindling and associated biochemical parameters in mice

Brain cyclooxygenases (COX), the rate-limiting enzyme in prostaglandin synthesis, is rapidly and transiently induced by convulsions in hippocampal and cortical neurons. Previous studies have explored the protective effect of naproxen (non-selective COX-inhibitor) or rofecoxib (selective COX-2 inhibitor) against chemical kindling in mice. With this background, the present study was designed to explore the possible effect of nimesulide (a preferential COX-2 inhibitor) against pentylenetetrazol (PTZ)-induced kindling epilepsy in mice. To induce kindling, PTZ was injected in a subconvulsive dose (40 mg/kg, i.p.) every other day for 15 days. Nimesulide (2.5 or 5 mg/kg, p.o.) was administered each day 45 min before either PTZ or vehicle challenge. The intensity of kindling was assessed immediately after PTZ administration according to a prevalidated scoring scale. On 16th day i.e. 24 h after the last dose of PTZ, animals were sacrificed and various biochemical parameters were assessed in the whole brain. Compared with normal control group, PTZ-kindled mice had significantly higher levels of malondialdehyde, nitrite, myeloperoxidase but had lower levels of reduced glutathione in the whole brain homogenate. Chronic treatment with nimesulide (2.5 or 5 mg/kg, p.o.) for 15 days showed significant decrease in kindling score and could play a role in controlling the accompanying biochemical alterations due to PTZ. These results suggested that nimesulide, a preferential COX-2 inhibitor offered neuroprotection against PTZ-induced kindling in mice.     

Effects of Nefiracetam, a novel pyrrolidone-type nootropic agent, on the amygdala-kindled seizures in rats

PURPOSE: Nefiracetam (NEF) is a novel pyrrolidonetype nootropic agent, and it has been reported to possess various pharmacologic effects as well as cognition-enhancing effects. The present study focused on the effects of NEF in amygdala-kindled seizures and its potential for antiepileptic therapy. METHODs: Effects of NEF on fully amygdala-kindled seizures and development of amygdala-kindled seizures were investigated in rats and compared with those of levetiracetam (LEV), a pyrrolidone-type antiepileptic drug (AED). RESULTS: In fully amygdala-kindled rats, NEF (25, 50, and 100 mg/kg, p.o.) decreased afterdischarge induction, afterdischarge duration, seizure stage, and motor seizure duration in a dose-dependent manner. LEV (25, 50, and 100 mg/kg, p.o.) had no effects on afterdischarge induction and slightly decreased afterdischarge duration, whereas it markedly decreased seizure stage and motor seizure duration. In contrast to the results in fully amygdala-kindled rats, NEF (25 and 50 mg/kg/day, p.o.) had few or no effects on the development of amygdala-kindled seizures. As well as fully amygdala-kindled seizures, LEV (50 mg/kg/day, p.o.) markedly inhibited the development of behavioral seizures without reducing daily afterdischarge duration. CONCLUSIONS: Although NEF possesses potent anticonvulsant effects on fully amygdala-kindled seizures, it has few or no effects on the development of amygdala-kindled seizures. LEV shows marked anticonvulsant effects on both phases of kindling. In fully amygdala-kindled rats, NEF inhibits both electroencephalographic and behavioral seizures, whereas LEV inhibits only behavioral seizures. This double dissociation suggests that NEF has a distinct anticonvulsant spectrum and mechanisms from those of LEV.     

戊四氮点燃大鼠模型海马结构内突触体素的变化

目的：探讨癫痫时海马结构的可塑性。方法：利用图像分析分析仪戊四氮诱导的大鼠点燃模型海马CA1、CA3区及齿状回区突触体素的阳性免疫反应产物的光密度值（OD值）。结果：点燃组海马人各部突触体素阳性免疫反应产物光密度值较对照组增加,尤以CA3区苔藓纤维层的齿状回分子层内带为甚。结论：突触体素含量的变化一方面是点燃的结果,另一方面也可能导致点燃维持的分子学基础。     

Specific amygdaloid nuclei are involved in suppression or propagation of epileptiform activity during transition stage between oral automatisms and generalized clonic seizures

Kindling is a model of the neural plasticity that occurs following stimulation to the brain, which can result in epileptogenesis. The amygdala (Am), one of the most sensitive structures from which to induce electrical kindling, is comprised of distinct nuclei that possess differences in threshold for seizure initiation, unique cellular and molecular morphology, and specific neuroanatomical connections within the amygdala and, to other cortical and subcortical brain structures. The aim of this study was to map the spread of epileptiform activity throughout the ipsilateral and contralateral hemispheres during the transition stage between oral automatisms and generalized clonic seizures, by measuring changes in mRNA expression for c-fos, NGFI-A, and BDNF. The stimulating electrode was implanted in either the basolateral (BL) or the lateral (CeL) or medial (CeM) subdivisions of the central nucleus of the amygdala. The rats were kindled once daily using afterdischarge-threshold electrical stimulation until the first forelimb clonic seizure was induced. They were sacrificed 30 min later, and their brains were prepared for in situ hybridization to measure mRNA expression of c-fos, NGFI-A and BDNF. The results demonstrate that: (1) the threshold to elicit an afterdischarge from the BL was lower than that of either the medial (CeM) or lateral (CeL) subdivisions of the Ce, which did not differ from each other; (2) the patterns of mRNA expression for c-fos, NGFI-A and BDNF were highly similar to each other when the stimulation site was the BL or the CeL, and included mainly limbic cortical and subcortical areas ipsilateral to the electrode; (3) c-fos was the only probe to be expressed in the contralateral hemisphere following the first motor seizure, and the pattern of its expression reflected a subset of structures recruited in the ipsilateral hemisphere including the claustrum, insular and perirhinal cortices; (4) unexpectedly, stimulation of the CeM elicited seizures and afterdischarges of shorter duration than those evoked by stimulation of the BL or CeL, and failed to increase mRNA expression for any of the probes in the hippocampus or in the contralateral hemisphere. A neuroanatomical model of Am-induced seizure propagation is proposed suggesting that the Claust–Ins–PRh play a pivotal role during the transition between oral automatisms and generalized clonic convulsions.     

Antiepileptic and prophylactic effects of tetrahydrocannabinols in amygdaloid kindled rats

The antiepileptic and prophylactic effects of delta9-tetrahydrocannabinol (delta9-THC) and delta8-THC were examined in rats that developed generalized seizures in response to intermittent electrical stimulation of the amygdala (kindling). Both isomers of the THC were able to acutely suppress kindled seizures, but consistent antiepileptic effects were obtained only with high, toxic dosages. Tolerance to the antiepileptic effects of THC developed very rapidly when the drugs were give repeatedly, and there was evidence that the repeated administration of a high dosage of delta9-THC resulted in a state of acute physical dependence. Administration of the isomers of THC during seizure development resulted in a suppression of kindling, suggestive of a prophylactic effect. The rate of rekindling after withdrawal of the drugs was not significantly different from that of vehicle-treated control rats, however, indicating that a genuine prophylactic effect was not obtained.     

Effect of Unilateral Dentate Nucleus Lesions on Amygdaloid Kindling in Rats

Amygdala kindling is used to investigate cerebellar dentate nucleus participation in the neuronal plasticity of the cerebrum. We studied the behavioral change, the occurrence of epileptiform events, and the duration of afterdischarge during amygdaloid kindling under control conditions and after a dentate lesion of the contralateral cerebellum. Our results indicate that the dentate lesion induced the facilitation of the behavioral development of the amygdaloid kindling, whereas there was only a slight increase in the duration of afterdischarge and the spontaneous epileptiform potential of the amygdala. There was a discrepancy between the behavioral kindling and the electroencephalographic epileptic activity. Therefore the dentate lesion must play a role in controlling the generalization mechanism of epilepsy. We also found that on a right amygdala that had been influenced by a left dentate lesion there were few spikes, while on the left amygdala there were frequent spikes. These results suggest that the dentate nucleus plays an important role in amygdaloid kindling as well as in the neuronal plasticity of the amygdala.     

Anticonvulsant Activity of Felbamate in Amygdala Kindling Model of Temporal Lobe Epilepsy in Rats

Summary: Purpose : Previous studies have demonstrated that felbamate (FBM, 2-phenyl-1,3-propanediol dicarbamate) at nontoxic doses exerts potent anticonvulsant activity in a variety of animal epilepsy or seizure models. We further characterized the anticonvulsant activity of FBM by using the kindling model of temporal lobe epilepsy (TLE).
Methods : The experiments were performed in fully kindled rats. The anticonvulsant effect of FBM was assessed by determining seizure severity, after discharge (AD) duration and seizure duration either at the focal seizure threshold, or after supra threshold stimulation. In addition, the neurological performance of kindled rats after FBM administration was evaluated in the open field and by the rotorod test.
Results : FBM at doses of 12.5–50 mg/kg, given intraperitoneally (i.p.) 60 min before testing, dose-dependently increased the AD threshold (ADT). The maximal effect was achieved after the highest dose tested and reached almost 600% of the control ADT. This dose of FBM significantly diminished other seizure parameters, e.g., seizure severity, seizure duration, and AD duration. When the rats were stimulated with suprathreshold current (500 μA) seizure severity was moderately but significantly reduced. No behavioral abnormalities were noted in kindled rats after administration of either of the doses.
Conclusions : FBM potently increases the threshold for focal seizures and reduces seizure severity, seizure duration, and AD duration at doses that produce no adverse behavioral effects in amygdala-kindled rats. These data are thus compatible with clinical experience with FBM in TLE and substantiate that kindling is a good predictor of anticonvulsant activity against TLE.     

Anticonvulsant Efficacy of Gabapentin on Kindling in the Immature Brain

Summary: The anticonvulsant and motor effects of gabapentin (GBP) were evaluated in rat pups aged 16–17 days. Fourteen‐day‐old rat pups received an implanted stimulating electrode in the amygdala unilaterally. Kindled seizures were produced on day 16 of life by repeatedly applying an electrical current stimulus to the amygdala electrode. Animals received kindling stimulation until they achieved three consecutive generalized convulsions. On day 17, rat pups received one of four doses of GBP 10, 25, 50, or 100 mg/kg. After receiving GBP, rat pups again received electrical stimulation to the amygdala electrode to determine the extent to which GBP prevented the kindled seizure. Anticonvulsant effects were found at doses as low as 10 mg/kg. A separate group of naïve rats received GBP to determine the motor effects of each treatment dose. Impaired motor performance, quantified as time on a balance beam, occurred at doses of ≥50 mg/kg. In summary, our data indicate that in immature rats, GBP exerts an anticonvulsant effect against kindled seizures at doses that do not significantly impair motor performance.     

Reversal of a selective decrease in hippocampal acetylcholine release, but not of the persistence of kindling, after discontinuation of long-term pentylenetetrazol administration in rats

The time course of the effect of pentylenetetrazol (PTZ)-induced kindling on acetylcholine release in the hippocampus of freely moving rats was investigated with the transversal microdialysis technique. The basal extracellular concentration of acetylcholine in the hippocampus was reduced significantly (−29%, P<0.05) after 3 weeks, and the effect was maximal (−52%, P<0.01) after 4 weeks and remained essentially unchanged during the remaining 4 weeks of PTZ treatment (30 mg/kg, i.p., 3 times/week), relative to vehicle-treated rats. The basal release of acetylcholine in the prefrontal cortex and in the striatum of kindled rats was unchanged compared with that of vehicle-treated rats. The specific binding of [³H]quinuclidinyl benzilate, a non-selective ligand of muscarinic receptors, was significantly increased (+29%, P<0.01) in hippocampal membranes, but not in membranes prepared from the prefrontal cortex or striatum, of PTZ-kindled rats. Thirty days after discontinuation of PTZ treatment, both hippocampal acetylcholine output and the density of muscarinic receptors had returned to values characteristic of vehicle-treated rats, whereas seizure susceptibility did not differ significantly from that apparent 4 days after PTZ administration. These results suggest that the selective and transient decrease in acetylcholine output and the parallel increase in the density of postsynaptic muscarinic receptors in the hippocampus may play a role in facilitating the development of kindling rather than in the maintenance of the kindled state. © 1997 Elsevier Science B.V. All rights reserved.     

Antiepileptogenic and anticonvulsant effects of NBQX, a selective AMPA receptor antagonist, in the rat kindling model of epilepsy

To investigate the role of non-NMDA receptors in epileptic seizures, we examined the antiepileptogenic and anticonvulsant effects of NBQX (2,3-dihydroxy-6-nitro-7-sulfamoyl-benzo(F)-quinoxaline), a potent and selective AMPA receptor antagonist, in the rat kindling model. Systemic administration of 10–40 mg/kg NBQX significantly and dose dependently suppressed previously kindled seizures from the amygdala (AM), assessed in terms of the motor seizure stage and afterdischarge (AD) duration. The maximal effects were observed at 0.5–1 h after drug injection. When the intensity of electrical stimulation was increased to twice the generalized seizure-triggering threshold (GST), the anticonvulsant effects of NBQX on AM-kindled seizures were not reversed, suggesting that the effects were not due to non-specific elevation of the GST. In contrast to AM-kindled seizures, 20–40 mg/kg NBQX significantly suppressed only the motor seizure stage without reducing the AD duration of previously hippocampal-kindled seizures. Daily administration of 15 or 30 mg/kg NBQX prior to each electrical stimulation of the AM markedly and significantly suppressed the development of kindling. During drug sessions, the growth of the AD duration was blocked almost completely, while the waveform of ADs became more complex. These results indicate that NBQX has potent antiepileptogenic and anticonvulsant actions on kindling, at least more from the AM and that non-NMDA receptors have an important role in seizure propagation.     

戊四氮点燃癫痫大鼠空间学习记忆障碍及海马突触素表达减少的研究

目的:探讨戊四氮点燃癫痫对大鼠空间学习记忆的影响及可能的分子机制。方法:戊四氮(pentylenetet-razol,PTZ)点燃建立慢性癫痫(chronic epileptic,CEP)模型,Morris水迷宫进行行为学检测,western blot方法观察大鼠海马突触素(synaptophysin,P38)蛋白的表达变化。结果:水迷宫试验检测癫痫组大鼠空间学习记忆能力受损(P<0.05,P<0.01);western blot结果表明海马P38蛋白表达较对照组明显减少(P<0.05)。结论:戊四氮点燃癫痫大鼠伴有学习记忆功能减退,其海马P38蛋白的表达减少可能参与了空间学习记忆受损。     

Endurance of the kindling effect is independent of the degree of generalization

The endurance of the effect of amygdaloid kindling was studied in rats kindled to stages 1–4, then left unstimulated for 45 days prior to resumption of kindling. Afterdischarge duration and behavioral stages upon resumption of kindling revealed retention of the full kindling effect. Endurance of the localized effect of kindling appears to be independent of the degree of generalization prior to interruption of kindling.     

The usefulness of olfactory bulb kindling as a model for evaluation of antiepileptics

Purpose: The present study was undertaken to clarify the behavioral and electroencephalographic characteristics of olfactory bulb (OB) kindling in rats, in comparison with those of amygdala (AMG) kindling. In addition, the usefulness of OB kindling as a model to evaluate antiepileptics was studied. Methods: Bipolar electrical stimulation was applied to the OB or AMG every day until generalized seizure was achieved. Antiepileptics (carbamazepine, sodium valproate, zonisamide, clobazam, and topiramate), which are used for complex partial epilepsy or secondary generalized epilepsy in clinical practice, were orally administrated to kindled rats. Results: The afterdischarge (AD) threshold of OB kindling is not different from that of AMG kindling. OB‐kindled rats showed more rapid development of the seizure stage and AD duration than AMG‐kindled rats; however, fully kindled AD duration did not differ between groups. In AMG kindled rats, AD on day 1 was localized only at the stimulation site, whereas in OB‐kindled rats, AD on day 1 was observed at not only the stimulation site (OB) but also in the frontal cortex, hippocampus, and AMG. All five antiepileptics significantly inhibited both the seizure stage and AD duration in OB‐kindled rats. In addition, carbamazepine, zonisamide, and topiramate were more effective in suppressing OB‐kindled seizures. Zonisamide was not effective at any dose tested in AMG‐kindled rats. Discussion: OB kindling can be used as a new valuable model to evaluate antiepileptic drugs, with the advantage of its rapid development and the efficacy of antiepileptics.     

Deep Prepiriform Cortex Lesion and Development of Amygdala Kindling

The effects of a unilateral thermocoagulating lesion of the deep prepiriform cortex on the development of amygdala kindling were studied in adult rats. The lesion did not alter the local excitability of the ipsilateral amygdala. Furthermore, the number of stimulations required to produce generalized seizures did not significantly differ between lesioned and control animals. The results suggest that the deep prepiriform cortex may not be important in the development of amygdala kindling.     

Effects of Unilateral Lesion in the Midbrain Reticular Formation on Kindled Amygdaloid Convulsion in Cats

Daily stimulation of the amygdala in cats resulted in progressive development of electroclinical seizures culminating in a generalized convulsion (kindling). An electrolytic lesion in the midbrain reticular formation, ipsilateral to the stimulated amygdala, markedly elevated the generalized seizure-triggering threshold and reduced susceptibility to pentylentetrazol challenge. In contrast, globus pallidus lesions had no appreciable effect upon the generalized seizure-triggering threshold and appeared to enhance susceptibility to pentylentetrazol. The results support the hypotheses that (1) the midbrain reticular formation participates significantly in the kindled amygdaloid seizures and (2) the effects of lesions in the midbrain reticular formation do not depend upon the presence of forebrain bisection.