Instituto Nacional de PediatrÃa, Facultad de Ciencias Biológicas de la Universidad de Morelos .... Zaczek R, Nelson MF & Coyle JT: Eur J Pharmacol 52: 323-327.
Proc. West. Pharmacol. Soc. 43: 35-37 (2000)
Effects of Oxcarbazepine on the Behavioral Response and Neuroanatomical Alterations Following Administration of Kainic Acid A. GONZÁLEZ-MACIEL,* R. REYNOSO-ROBLES, R. M. ROMERO, B. HUERTA, V. GONZÁLEZ, L. VARGAS & F. AYALA-GUERRERO Instituto Nacional de Pediatría, Facultad de Ciencias Biológicas de la Universidad de Morelos, Facultad de Psicología, Universidad Nacional Autónoma de México
Epilepsy is considered to be a disorder intrinsic to the brain, deriving from either a hereditary tendency or a prior insult, in which a portion of the brain is rendered electrically unstable [1]. Temporal lobe epilepsy is the most devastating form of epilepsy commonly encountered in the adult population. The attacks involve loss of consciousness, thus limiting performance of normal functions and exposing the individual to bodily injury. Moreover, long-standing or pharmacologically intractable temporal lobe epilepsy is frequently associated with the loss of neurons from the hippocampus and other brain regions [2-4] Unfortunately, pharmacologically intractable cases are rather common, owing to the relatively low efficacy against this condition of the available anticonvulsants. As ethical considerations rule out use of the modern neuroscientific tools in intact human brain, the study of epilepsy has been dependent upon use of model systems. Much of what is known about the epilepsy is derived from animal models [5]. Basic models of epilepsy have been used to explore questions about seizures and the electrical activity of the brain. These questions are related to the underlying EEG generators of electrical potentials associated with seizures: the nature and identity of neuronal systems able to produce epilepsy; issues of why seizures start, spread and stop, and why seizures occur when they do; what type of pathologies in brain give rise to seizures; whether seizures cause damage to brain; and the mechanisms of action of anticonvulsant drugs [6-8]. In past decades, a variety of animal models of epilepsy have been used to identify and evaluate chemicals with anticonvulsant efficacy. It is of particular interest that systemic administration of kainic acid (KA), an analogue of the excitatory neurotransmitter glutamate, causes persistent seizure activity and a disseminated pattern of seizure-related brain damage. For reasons that are incompletely understood, KA has an especially prominent toxic effect on the hippocampus, even when injected systemically, or at brain sites remotes from hippocampus [9-11]. The accompanying hippocampal lesions may be considered to portray the pattern of limbic cell damage that can occur with clinical status epilepticus [12,13].
It is interesting that the anticonvulsant activity and side effects of several chemicals have been tested in animal models of kainite-induced epilepsy. For example, some benzodiazepines, certain barbiturates and trimethadione are most effective in preventing KA-induced seizures and seizure-induced brain damage [14-17], while chloral hydrate or diethyl ether are less effective in inhibiting the behavioral and neuropathological changes caused by KA [18]. Diphenylhydantoin, carbamazepine, valproate and ethosuximide have not a clear anticonvulsive action on seizures induced by KA [19,20]. On the other hand, oxcarbazepine has been extensively tested in humans [21]. However no large comparative studies on efficacy have been conducted, therefore the aim of this experimental work was to test the possible protective action of the oxcarbazepine against the seizures and brain damage induced by KA administration. METHODS: Experiments were carried out on 16 male Wistar adult rats weighing between 300 and 400 g with food and water continuously available. Rats were divided into 4 similar groups of 4 animals. A control group received a saline injection sc. A further two groups were treated either with KA (10 mg/kg, ip) or oxcarbazepine (100 mg/kg, po) alone, while the final group was pretreated with oxcarbazepine before administration of KA. After substances were administered, animals were kept into their home cage and behavior was observed during 10 continuous hours. After a minimal period of ten days, animals were anesthetized with sodium pentobarbital (55 mg/kg) followed by intracardiac perfusion with 10% formalin in phosphate buffer (pH 7.4). Brains were extracted and kept in the same fixing solution for at least 24 h. Brains were then embedded in paraffin in order to proceed with the histological technique. Serial sections of 5 µm were cut in the coronal plane throughout the rostro-caudal axis with a 70 µm difference between each section. Six slides with four sections each were mounted and stained with Cresyl Violet. Slides were examined under light microscope.
RESULTS: Behavioral findings. Control rats presented alternated period of sleep and wakefulness throughout the observational periods. When awakened they showed apparently normal motor behavior. Animals treated with KA alone displayed an abnormal behavior characterized by general hyperactivity and convulsions. This behavior included eye blinking, masticatory movements, head nodding and myoclonic twitches of the head and fore limbs. Severe motor 35
Figure 1. Nissl-stained section of the hippocampal formation of a untreated rat. Cal: 100 µm.
Figure 2. Nissl-stained section of the hippocampal formation from a rat treated with oxcarbazepine and kainic acid. Noticeable neuronal depopulation occurs in CA1. Box denotes a lesioned portion (inset). Cal: 100µm; inset: 25µm.
seizures involving the whole body were present about 2 h after KA administration. In oxcarbazepine pretreated animals, abnormal motor activity induced by KA was markedly reduced or absent,
while rats treated only with oxcarbazepine adopted a sleep behavior throughout the observational period. Neuropathological findings. Microscopic inspection of brains obtained from animals studied under different 36
experimental conditions revealed significant differences at the hippocampal level. As previously reported [12], neuropathological changes were observed in the hippocampus after KA administration. On the other hand, in spite of the fact that hyperactivity and convulsions decreased significantly or were absent, pretreatment with oxcarbazepine did not protect against the neurodegenerative effect exerted by KA administration. This is evidenced when comparing Nissl stains of the dorsal hippocampus of a control (Fig. 1) and an oxcarbazepine pretreated rat (Fig. 2) Severe damage of pyramidal cells, specially in the CA1 sector, is observed.
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DISCUSSION: Excitatory amino acids, such as glutamate, aspartate and KA have the ability to destroy CNS neurons [22,23]. As structure-activity studies have suggested a relation between neuroexcitatory and neurotoxic actions of these amino acids [24], KA is thought to destroy neuronal cells by exciting them to the point that the cellular energy stores become depleted [25]. Strong support for the central role of excitation in KA-induced neuron death arises from findings that KA does not kill neurons that are insensitive to its excitatory action [26] or which are protected from excitation by deep and prolonged anesthesia [27,28]. Few studies have analyzed effects of anticonvulsant drugs on seizures induced by KA. Results have been contradictory, since these seizures can be attenuated by chlordiazepoxide, trimethadione, phenobarbital and diazepam, but not by phenytoin, ethosuximide or valproic acid [16,29]. The mechanisms whereby these drugs prevent convulsions vary, but they all induce a decrement of neuronal excitation [30-32]. Behavioral seizure activity induced by KA was reduced by oxcarbazepine. However, histological damage was still present in the hippocampus. This evidence suggests that KAinduced brain damage may involve mechanisms that are not directly related to neuroexcitation. Alternatively, oxcarbazepine may exert a neurodegenerative effect even if convulsions disappear or decrease significantly. CONCLUSIONS: Consistent with previous reports, administration of KA produced seizure activity accompanied by histological damage in the hippocampus. KA-induced seizures were moderately inhibited after administering oxcarbazepine. However, this anticonvulsant drug did not protect against hippocampal neuronal degeneration.
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