Spine loss in experimental epilepsy: Quantitative light and electron microscopic analysis of intracellularly stained CA3 pyramidal cells in hippocampal slice cultures

The sequence of neuronal alterations resulting from epileptic activity is poorly understood. In the hippocampus of some epileptic patients, there is a loss of certain neuronal types in the hilar region and in CA3. The neuronal alterations preceding this degeneration probably affect synaptic structures. Here we have estimated the number of dendritic spines, major postsynaptic elements of hippocampal neurons, in defined dendritic segments of identified (intracellularly stained) CA3 pyramidal neurons in “epileptic” slice cultures of hippocampus and in control cultures. Slice cultures were prepared from five- or six-day-old rat pups and maintained in vivo for 23 days before epileptic activity was induced by application of the convulsants bicuculline and picrotoxin for three days. Individual CA3 pyramidal neurons were then intracellularly injected with horseradish peroxidase, and the number of dendritic spines was counted in proximodistal dendritic segments by applying the Sholl method. In addition, the total dendritic length was measured and the branching index evaluated. The number of spines on CA3 pyramidal cell dendrites in the “epileptic” cultures was found to be decreased by 40%. This spine loss affected proximal and peripheral dendritic segments of the CA3 pyramidal neurons to a similar extent. No significant differences were observed between control and “epileptic” cultures in dendritic length or in the branching index. Quantitative electron microscopic analysis did not reveal differences between “epileptic” cultures and control cultures in the spine area of the labelled CA3 pyramidal cells, indicating that there was a real spine loss, not just a reduction in the size of the spines. We conclude that epileptic activity causes morphological alterations in defined postsynaptic compartments of hippocampal pyramidal cells surviving under these conditions.