Introduction
The nature of the brain lesions in temporal lobe epilepsy remains largely unknown. Unilateral administration of carbachol (CCh) in the lateral thalamus of Wistar rats triggers secondary generalized convulsive seizures (GCS) associated with bilateral hippocampal cell loss and ipsilateral damage of the piriform/entorhinal cortices and amygdala1, 2 comparable to human mesial temporal lobe sclerosis. The objectives of the study was to establish (1) whether phenomenon of thalamic-evoked lesion-accompanied GCS is strain and species independent; (2) whether regional cerebral blood flow (rCBF) in the piriform cortex is affected by thalamic CCh microinjections and (3) whether apoptotic neuronal death pathway is involved in remote thalamic-evoked brain lesions.
Methods
Experiments were performed in Sprague Dawley (SD) rats and C56B7 mice. Animals were anesthetized and CCh (100 nmol/100 nl in PBS) was stereotaxically injected into thalamic ventroposterolateral/reticular nuclei (VPL/nRT). Animals were allowed to recover and were observed for 2 hours for seizures. 24–72 hours later animals were euthanized; brains were removed and processed for DNA fragmentation (dUTP nick end-labeling, TUNEL); for caspase-3 activation; for changes in levels of major histocompatibility complex 1 (MHC1); and histology. Piriform cortex rCBF (laser Doppler flowmetry) and EEG (monopolar) were recorded before and 4 hours after CCh microinjection in anesthetized rats.
Results
In accord with earlier observations in Wistar rats 2 microinjections of CCh in VPL/nRT triggered multiple (4.6±1.5) episodes of GCS in SD rats (n=4) and mice (n=5) accompanied in 24–72 hours by lesions in hippocampus, piriform/entorhinal cortices, and amygdala as revealed by thionin staining. rCBF in piriform cortex increased by 128.30±90.7 % (p>0.05, n=2) within 2 minutes after ipsilateral thalamic CCh microinjections and gradually returned to the baseline in 45±31.8 minutes. rCBF remained at the baseline level during the rest of observation period (4 h) while EEG demonstrated multiple episodes (6.5±4.5, duration 33.7±23 sec) of spike-wave activity. In mice unilateral intrathalamic CCh administration (n=5) also initiated GCS comparable to those observed in rats. Subsequent histological analysis revealed TUNEL positive staining bilaterally in the ventromedial thalamus, hypothalamus and hippocampus, amygdaloid complex and piriform cortex. TUNEL positivity in the ipsilateral piriform cortex and amygdala was accompanied by lesions, while no lesions were observed in thalamic or hypothalamic areas. Only low levels of caspase-3 activation were seen in the TUNEL positive regions. In the presence of lesions MHC1 immunolabeling increased in ipsilateral piriform/entorhinal cortices, amygdala and bilaterally in the hippocampus However no changes of MHC1 immunoreactivity were observed in animals presenting only GCS.
Conclusions
We conclude that (1) induction of GCS and temporal lobe damages after thalamic stimulation is strain and species independent; (2) brain damages in the piriform/entorhinal cortices and amygdala are not associated with a decrease in rCBF; (3) early TUNEL positivity is not necessarily associated with brain lesions within 72 hours; (4) thalamic-evoked brain lesions seem independent of caspase-3 activation; (4) immune mechanisms may be involved in the thalamic-evoked brain lesions.