Elemental anomalies in the hippocampal formation after repetitive electrical stimulation: an X-ray fluorescence microscopy study.

Our previous studies carried out on the pilocarpine model of seizures showed that highly resolved elemental analysis might be very helpful in the investigation of processes involved in the pathogenesis of epilepsy, such as excitotoxicity or mossy fiber sprouting. In this study, the changes in elemental composition that occurred in the hippocampal formation in the electrical kindling model of seizures were examined to determine the mechanisms responsible for the phenomenon of kindling and spontaneous seizure activity that may occur in this animal model. X-ray fluorescence microscopy was applied for topographic and quantitative analysis of selected elements in tissues taken from rats subjected to repetitive transauricular electroshocks (ES) and controls (N). The detailed comparisons were carried out for sectors 1 and 3 of the Ammon's horn (CA1 and CA3, respectively), the dentate gyrus (DG) and hilus of DG. The obtained results showed only one statistically significant difference between ES and N groups, namely a higher level of Fe was noticed in CA3 region in the kindled animals. However, further analysis of correlations between the elemental levels and quantitative parameters describing electroshock-induced tonic and clonic seizures showed that the areal densities of some elements (Ca, Cu, Zn) strongly depended on the progress of kindling process. The areal density of Cu in CA1 decreased with the cumulative (totaled over 21 stimulation days) intensity and duration of electroshock-induced tonic seizures while Zn level in the hilus of DG was positively correlated with the duration and intensity of both tonic and clonic seizures.

Differences in the hippocampal frequency of creatine inclusions between the acute and latent phases of pilocarpine model defined using synchrotron radiation-based FTIR microspectroscopy.

Temporal lobe epilepsy (TLE) is the most common type of epilepsy in adults. Of the animal models developed to investigate the pathogenesis of TLE, the one with pilocarpine-induced seizures is most often used. After pilocarpine administration in animals, three distinct periods--acute, latent, and chronic--can be distinguished according to their behavior. The present paper is the continuation of our previous study which has shown an increased occurrence of creatine inclusions in rat hippocampal formations from the acute phase of pilocarpine-induced status epilepticus (SE) and positive correlation between their quantity and the total time of seizure activity within the observation period. In this paper, we tried to verify if anomalies in hippocampal creatine accumulation were the temporary or permanent effect of pilocarpine-evoked seizures. To realize this purpose, male Wistar rats in the latent phase (3 days after pilocarpine administration) were examined. The results obtained for the period when stabilization of animal behavior and EEG occurs were afterwards compared with ones obtained for the acute phase of pilocarpine-induced SE and for naive controls. To investigate the frequency of creatine inclusions within the hippocampal formation as well as in its selected areas (sectors 1-3 of Ammon's horn (CA1-CA3), dentate gyrus (DG), and hilus of DG) and cellular layers (pyramidal, molecular, multiform, and granular cell layers), synchrotron radiation-based Fourier-transform infrared microspectroscopy was used. The applied technique, being a combination of light microscopy and infrared spectroscopy, allowed us to localize microscopic details in the analyzed samples and provided information concerning their chemical composition. Moreover, the use of a synchrotron source of IR radiation allowed us to carry out the research at the diffraction-limited spatial resolution which, because of the typical size of creatine inclusions (from a few to dozens of micrometers), was necessary for our study. The comparison of epileptic animals in the latent phase with controls showed statistically significant increase in the number of creatine inclusions for most of the analyzed hippocampal regions, all examined cellular layers, as well as the whole hippocampal formation. Moreover, for the hilus of the DG and CA3 area, the number of creatine deposits was higher in the latent than in the acute phase after pilocarpine injection. In light of the obtained results, an anomaly in the hippocampal accumulation of creatine is the long-term effect of pilocarpine-evoked seizures, and the intensity of this phenomenon may increase with time passing from the primary injury.

Progress of elemental anomalies of hippocampal formation in the pilocarpine model of temporal lobe epilepsy--an X-ray fluorescence microscopy study.

In the present paper, X-ray fluorescence microscopy was applied to follow the processes occurring in rat hippocampal formation during the post-seizure period. In the study, one of the status epilepticus animal models of epilepsy was used, namely the model of temporal lobe epilepsy with pilocarpine-induced seizures. In order to analyze the dynamics of seizure-induced elemental changes, the samples taken from seizure-experiencing animals 3 h and 1, 4, and 7 days after proconvulsive agent administration were analyzed. The obtained results confirmed the utility of X-ray fluorescence microscopy in the research of mechanisms involved in the pathogenesis and progress of epilepsy. The topographic and quantitative elemental analysis of hippocampal formations from different periods of epileptogenesis showed that excitotoxicity, mossy fibers sprouting, and iron-induced oxidative stress may be the processes responsible for seizure-induced neurodegenerative changes and spontaneous recurrent seizures occurring in the chronic phase of the pilocarpine model. The analysis of correlations between the recorded elemental anomalies and quantitative parameters describing animal behavior in the acute period of pilocarpine-induced status epilepticus showed that the areal densities of selected elements measured in the latent period strongly depend on the progress of the acute phase. Especially important seem to be the observations done for Ca and Zn levels which suggest that the intensity of the pathological processes such as excitotoxicity and mossy fibers sprouting depend on the total time of seizure activity. These results as well as dependencies found between the levels of S, K, and Cu and the intensity of maximal seizures clearly confirm how important it is to control the duration and intensity of seizures in clinical practice.