Zdeněk Hliňák

Long-term behavioral and morphological consequences of nonconvulsive status epilepticus in rats.

The aims of the present study were to ascertain whether nonconvulsive status epilepticus (NCSE) could give rise to long-term behavioral deficits and permanent brain damage. Two months after NCSE was elicited with pilocarpine (15 mg/kg i.p.) in LiCl-pretreated adult male rats, animals were assigned to either behavioral (spontaneous behavior, social interaction, elevated plus-maze, rotorod, and bar-holding tests) or EEG studies. Another group of animals was sacrificed and their brains were processed for Nissl and Timm staining as well as for parvalbumin and calbindin immunohistochemistry. Behavioral analysis revealed motor deficits (shorter latencies to fall from rotorod as well as from bar) and disturbances in the social behavior of experimental animals (decreased interest in juvenile conspecific). EEGs showed no apparent abnormalities. Quantification of immunohistochemically stained sections revealed decreased amounts of parvalbumin- and calbindin-immunoreactive neurons in the motor cortex and of parvalbumin-positive neurons in the dentate gyrus. Despite relatively inconspicuous manifestations, NCSE may represent a risk for long-term deficits.

Oxiracetam prevents the MK-801 induced amnesia for the elevated plus-maze in mice

We investigated the effect of the nootropic substance oxiracetam on the impairment of memory induced in mice by the non-competitive NMDA antagonist MK-801. Memory capacities of animals having different experience were evaluated using the elevated plus-maze test. Oxiracetam was injected immediately after the acquisition session(s), MK-801 was given 30 min before the retention session which followed 24 h after the acquisition session(s). In slightly experienced animals (Section 3.1), oxiracetam (3 and 30 mg/kg, s.c.) prevented MK-801 (0.15 mg/kg, i.p.) induced memory deficits characterized by a prolongation of the transfer latency. In well-trained animals (Section 3.2), oxiracetam (30 mg/kg, s.c.) attenuated MK-801 (0.15,0. 25 and 0.4 mg/kg, i.p.) induced amnesia for a spatial orientation in the elevated plus-maze. These results show that oxiracetam interacted with the glutamatergic NMDA receptor system and forestalled the impairment of retrieval of long-term memory. The results also justify the usage of the elevated plus-maze method in the evaluation of potential anti-amnesic or nootropic drugs.

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

Summary:  Purpose: To characterize electroencephalographic and behavioral effects as well as electrophysiologic and morphologic consequences of a subconvulsive dose of pilocarpine in lithium chloride–pretreated rats.

Behavioural effects of a subconvulsive dose of kainic acid in rats.

Kainic acid can induce a continuum of non-convulsive seizures characterised by epileptic automatisms and convulsive motor seizures depending on the dose. There are scarce data on the behavioural effects of low doses of kainate inducing only non-convulsive seizures. Therefore, we studied spontaneous behaviour of adult male rats using a method of positive habituation based on a detailed analysis of patterns and attention of animals to a stimulus object. Twenty-three animals were individually tested in the experimental arena on two consecutive days. Comparing the data from the first two exposures, a conspicuous habituation in all animals was observed. On experimental day 3, 12 rats received kainate (6 mg/kg intraperitoneally) and the remaining 11 animals received a physiological saline. After 1 h, animals were put into the arena with an object localised in the centre. It was found that both kainate and saline treated animals exhibited a significant increase in the total number of central area visits, and both the total and mean time spent in the vicinity of the object. However, the mean time spent was significantly shorter in kainate treated rats. Furthermore. kainate rats exhibited a significant decrease in rearing as compared with the controls. In addition, an epileptic automatism (wet dog shakes) was observed in seven out of 12 animals given kainate. The comparison of transition matrices between consecutive behavioural categories showed significant differences between the kainate and control groups. Our results demonstrate that a non-convulsive dose of kainate induced changes in the structure of spontaneous behaviour and impaired the processes related to maintenance of attention.

N-methyl-D-aspartate improved social recognition potency in rats.

Activation of N-methyl-D-aspartate (NMDA) receptors is believed to play an important role in cognitive processes. In the present study we addressed the question of whether systematically administered NMDA can improve social recognition in adult male rats. We utilized the paradigm in which an adult animal is exposed to the same or a novel juvenile during two successive interactions. The adults given NMDA (10, 25 and 50 mg/kg, s.c.) immediately after the initial exposure to a juvenile exhibited significantly reduced social investigation during the second encounter with the same juvenile performed both 120 and 240 min later. Reduction in social investigation was not observed in saline treated animals as well as in those treated with both NMDA and saline but re-exposed to novel juveniles. These results indicate that NMDA improved social recognition potency of adult animals and prolonged retention of previously stored olfactory information.

Oxiracetam prevented the scopolamine but not the diazepam induced memory deficits in mice

In mice, the elevated plus-maze paradigm was used to investigate the effect of scopolamine hydrobromide and diazepam and their interaction with oxiracetam on the retrieval of spatial memory trace. This paradigm measures (using the transfer latency) an animals capacity to escape from the open arm to the enclosed one. The retention session followed 24 h after the acquisition one. Experiment 1: Scopolamine (0.25 and 0.5 mg/kg) and diazepam (0.5 and 1.0 mg/kg) given 30 min before the retention session significantly prolonged the transfer latency as compared with the saline treated mice and those given the lowest dose of scopolamine (0.125 mg/kg) and diazepam (0.25 mg/kg). Experiment 2: Oxiracetam administered at doses of 3, 10 and 30 mg/kg immediately after the acquisition session prevented the scopolamine induced prolongation of the transfer latency. Thus, oxiracetam forestalled the impairment of retrieval of memory trace: the animals were able to remember the spatial configuration of the plus-maze. On the contrary, oxiracetam was not effective in the diazepam treated mice. We suggest that beneficial effect of oxiracetam might be confounded or blocked by the anxiolytic effect of diazepam.

Spontaneous alternation behaviour in rats: kynurenic acid attenuated deficits induced by MK-801.

The present study was undertaken to investigate the effects of pharmacological modulation of the NMDA receptors on spontaneous alternation behaviour. The performance of rats treated with MK-801 and kynurenic acid (KYNA) was assessed in the cross-arm-maze. We evaluated: (a) the total number of arm entries representing locomotor activity, (b) spontaneous variation of different arms thought to reflect alternation performance. In the first experiment, MK-801 (0.01, 0.025, 0.05, 0.1 and 0.2 mg/kg, i.p.) was given 30 min prior to the testing. Beginning the dose of 0.05 mg/kg the drug increased locomotion and impaired alternation performance. An ability of animals to enter subsequently three or four different arms was reduced significantly. In the second experiment, the dose of 0.05 mg/kg was chosen as the lowest possible dose of MK-801 producing marked behavioural impairment. KYNA (0.3, 3 and 30 mg/kg, s.c.) was administered 60 min prior to the MK-801. While all KYNA doses prevented hyperlocomotion, only the highest dose (30 mg/kg) maintained alternation score at the control levels, i.e. the KYNA plus MK-801 treated animals alternated regularly three or four different arms. The results suggest different sensitivity of the two behavioural systems, i.e. locomotion and space orientation, towards pharmacological insult. In conclusion, the study confirmed protective behavioural effects of KYNA given in sufficient amounts and sufficiently long prior MK-801.

Nonconvulsive status epilepticus in rats: impaired responsiveness to exteroceptive stimuli.

An animal model of human complex partial status epilepticus induced by lithium chloride and pilocarpine administration was developed in our laboratory. The objective of the study was to provide a detailed analysis of both ictal and postictal behavior and to quantify seizure-related morphological damage. In order to determine the animals responsiveness to either visual or olfactory stimuli, adult male rats were submitted to the following behavioral paradigms: the object response test, the social interaction test, and the elevated plus-maze test. The rotorod test was used to evaluate motor performance. Two weeks after status epilepticus, brains were morphologically examined and quantification of the brain damage was performed. Profound impairment of behavior as well as responsiveness to exteroceptive stimuli correlated with the occurrence of epileptic EEG activity. When the epileptic EEG activity ceased, responsiveness of the pilocarpine-treated animals was renewed. However, remarkable morphological damage persisted in the cortical regions two weeks later. This experimental study provides support for the clinical evidence that even nonconvulsive epileptic activity may cause brain damage. We suggest that the model can be used for the study of both functional and morphological consequences of prolonged nonconvulsive seizures.

Oxiracetam pre- but not post-treatment prevented social recognition deficits produced with trimethyltin in rats

The social recognition paradigm was used to investigate the effect of trimethyltin (TMT) in adult male rats. Consequently, the effect of chronic oxiracetam (OXI) treatment in TMT impaired animals was evaluated. In all experiments, a behavioural testing was performed 3 weeks after TMT administration. Experiment 1: A single TMT oral dose, 5 and 7.5 but not 2.5mg/kg, impaired the natural ability of the adults to recognize a juvenile conspecific that they encountered 30 min before. The dose of 5mg/kg TMT was chosen to be used in subsequent experiments. Experiment 2: Chronic OXI pre-+post-treatment, daily 3 or 30 mg/kg sc for 7 days before and 7 days after the insult, protected the adults against recognition deficit produced by TMT. Experiment 3: OXI pre- but not post-treatment (always 3 and 30 mg/kg) had beneficial effects on the social recognition. The findings suggest that social recognition ability of adult male rats pre-treated sufficiently long with OXI is resistant to the neurotoxicity effect of TMT.