The proposed experiments will address a fundamental hypothesis concerning the mechanisms of stroke-induced synaptic reorganization and the potential for epileptogenesis. A modification of the four vessel occlusion model will be used in the proposed experiments; this treatment leads to global cerebral ischemia and will serve as a model for stroke. This model has not been used previously to study lesion-induced epilepsy, and thus seizures have not been reported for this model. The hypothesis to be tested is that synaptic reorganization in the dentate gyrus, following a stroke, leads to new abnormal recurrent excitatory circuits. The subsequent sprouting of mossy fibers may not be sufficient to induce hippocampal epileptogenesis, unless the ischemic accident is directly associated with seizures. Bilateral in vivo and in vitro electrophysiological experiments, as well as unbiased quantitative stereological morphometric techniques, will be utilized to test this hypothesis. In vivo recording from the hippocampus of freely-behaving animals in combination with video monitoring will be used to evaluate the subsequent development of spontaneous seizure activity and interictal spikes, and to detect focal and generalized seizures. In vitro studies will employ extracellular and whole-cell path-clamp recording in the hippocampal slice preparation to evaluate recurrent inhibition and excitation. Glutamate micro-stimulation with flash photolysis of caged glutamate will be used to analyze the properties of local synaptic circuits between granule cells. These slices, and the contralateral hippocampus, will then be processed for anatomical studies using cresyl violet and Timm staining to quantify neuronal loss and mossy fiber sprouting. These experiments should provide a detailed analysis of the formation of new local synaptic circuits after ischemia-induced neuronal death and subsequent axonal sprouting.
