Elsevier

Brain Research

Volume 1066, Issues 1–2, 20 December 2005, Pages 37-48
Brain Research

Research Report
Behavioral changes resulting from the administration of cycloheximide in the pilocarpine model of epilepsy

https://doi.org/10.1016/j.brainres.2005.09.037Get rights and content

Abstract

Cycloheximide influences synaptic reorganization resulting from pilocarpine-induced status epilepticus (SE). To investigate the possible behavioral consequences of this effect, we subjected animals to pilocarpine-induced SE either in the absence (Pilo group) or presence of cycloheximide (Chx group). Animals were further divided regarding the occurrence of spontaneous recurrent seizures (SRS). Two months after SE induction animals were exposed to different behavioral tests. Age-matched naïve animals were used as controls. All epileptic groups showed a significantly diminished freezing time in contextual and tone fear conditioning, performed poorly in the Morris water maze and present less seconds in immobility position as compared to controls. Only Pilo animals explored more extensively the open arms of the elevated plus maze and showed increased in horizontal exploratory activity in the open field as compared to controls. With the exception of Pilo animals without recorded SRS, all other groups had extensive tissue shrinkage in central nucleus of the amygdala as compared to controls. Cycloheximide-treated animals differed from Pilo animals in the extent of hilar loss and supragranular mossy fiber sprouting as well as tissue shrinkage in the dorsal hippocampus. Despite the histological differences seen in the dorsal hippocampus between experimental groups, no differences were encountered in the cognitive tests used to evaluate dorsal hippocampal function. The encountered histological differences between Chx and Pilo animals, however, might underlie the different emotional responses between the two groups.

Introduction

“Neuroplasticity” subsumes diverse processes of vital importance by which the brain perceives, adapts to and responds to a variety of internal and external stimuli. The manifestations of neuroplasticity in the adult central nervous system have been characterized as including alterations of dendritic function, synaptic remodeling, axonal sprouting, neurite extension, synaptogenesis and even neurogenesis [46]. Plasticity plays an important role in response to injury. Lesions of the entorhino-hippocampal pathway induce intact neurons to sprout collateral branches so that they can occupy the denuded synaptic targets in the molecular layer of the dentate gyrus [12], [46].

Histological analysis in human or experimentally-induced animal models of temporal lobe epilepsy (TLE) has revealed pathological alterations in the hippocampus [23], [44], [55], a region that is thought to be involved in cognition, and hence such anatomical alterations are assumed to cause cognitive deterioration [63]. In one of the most studied model of TLE, when animals are given pilocarpine, a muscarinic cholinergic agonist, they present status epilepticus (SE), and after an average latent period of 2–3 weeks, they show spontaneous recurrent epileptic seizures (SRS) [45], [62]. The brain damage induced by SE may be considered an equivalent of the initial precipitating injury event, such as a prolonged febrile convulsion, which is commonly found in patients with TLE [23], [44]. Mossy fiber sprouting (MFS) has been described in human mesial temporal sclerosis [4], [26], [62] as well as in several animal models of temporal lobe epilepsy [9], [45]. MFS involves the formation of new asymmetrical synaptic contacts between mossy fiber terminals and dendrites of granule cells and inhibitory interneurons in the inner molecular layer [8], [29], [49]. The precise mechanisms and functional consequences of seizure-induced MFS remain to be defined. Ultrastructural and electrophysiological studies of MFS into the inner molecular layer suggest that this synaptic reorganization may result in recurrent excitatory circuits and subsequent hippocampal hyperexcitability [7], [64]. A recent study, however, has suggested that hilar cell loss rather than other plastic consequences of status epilepticus (SE), such as MFS, is responsible for the altered function of this tissue [68]. We have shown that the administration of cycloheximide, prior to the induction of SE prevents the subsequent MFS without affecting epileptogenesis [41], [42], [43]. The data from these studies suggest that the MFS is neither necessary, nor sufficient, for seizures generation in temporal lobe epilepsy.

It has been suggested that the MFS seen in epileptic tissue might be an adaptive phenomenon that develops after brain injury [56], [59]. Accordingly, if this hypothesis is correct, MFS could potentially diminish the cognitive deficits seen in patients with temporal lobe epilepsy. An alternative suggestion is that the abnormal circuit remodeling characterized by MFS might cause additional hippocampal malfunction, thus leading to further cognitive deficits [28], [63]. To investigate the possible functional consequence of MFS at cognitive aspects, we sought to study this issue by using the reported ability of cycloheximide to block MFS [41], [42], [43] that would otherwise ensue after pilocarpine-induced SE. The choice of the pilocarpine model of TLE was based namely by the fact that MFS in this model is amongst the most exuberant found in animal models of TLE [41], [42].

Section snippets

Materials and methods

This study was conducted under protocols approved by the Animal Care and Use Ethics Committee at Universidade Federal de São Paulo and in accordance with the Guide for Care and Use of Laboratory Animals adopted by the National Institutes of Health.

Spontaneous recurrent seizures

Spontaneous seizures were seen in a similar 61% of the animals of the Pilo group and 75% of the animals in the Chx group. Spilo and Schx did not differ from each other with regard to the frequency of spontaneous recurrent seizures.

MWM

In the MWM test, all groups of Pilo and Chx animals demonstrated a significantly worse performance as compared to controls, without differing from each other (Fig. 1, Fig. 2). In the acquisition phase (Fig. 1), the two-way ANOVA for repeated measures revealed

Discussion

In all cognitive tasks (contextual and tone fear conditioning, as well as in spatial reference memory version of the MWM) and in despair swim (emotional aspects), the performance of all Pilo and Chx groups was impaired as compared to controls, with no difference from each other. However, Pilo animals showed a diminished emotional arousal in the elevated plus maze test and presented an augmented horizontal exploratory activity in the open-field test. At the histological level, only Pilo animals

Acknowledgments

We thank Ivone de Paulo for excellent technical assistance. This research was supported by CNPq-Institutos do Milenio, FAPESP-CEPID and PRONEX. J.G.S.J. is a FAPESP fellow (01/07455-6).

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