Pilocarpine-induced epilepsy is associated with actin cytoskeleton reorganization in the mossy fiber-CA3 synapses

•F-actin density is decreased in the hippocampus of mice that underwent the pilocarpine status epilepticus.•The F-actin changes were consistent with postsynaptic densities in the CA3 region.•F-actin puncta were more highly clustered and enlarged in the epileptic mice.•Synapsin I was not altered in the CA3 region of epileptic mice.•Cytoskeletal changes may be associated with functional changes related to hippocampus sclerosis.

SummaryDramatic structural changes have been demonstrated in the mossy fiber-CA3 synapses in the post status epilepticus (SE) animals, suggesting a potential reorganization of filamentous actin (F-actin) network occurring in the hippocampus. However, until now the long-term effects of SE on the synaptic F-actin have still not been reported. In this study, phalloidin labeling combined with confocal microscopy and protein analyses were adopted to investigate the effects of pilocarpine treatment on the F-actin in the C57BL/6 mice. As compared to the controls, there was ∼ 43% reduction in F-actin density in the post SE mice. Quantitative analysis showed that the labeling density and the puncta number were significantly decreased after pilocarpine treatment (p < 0.01, n = 5 mice per group, Student's t-test). The puncta of F-actin in the post SE group tended to be highly clustered, while those in the controls were generally distributed evenly. The mean puncta size of F-actin puncta was 0.73 ± 0.19 μm2 (n = 1102 puncta from 5 SE mice) in the experimental group, significantly larger than that in the controls (0.51 ± 0.10 μm2, n = 1983 puncta from 5 aged-matched control mice, p < 0.01, Student's t-test). These observations were well consistent with the alterations of postsynaptic densities in the same region, revealed by immunostaining of PSD95, suggesting the reorganization of F-actin occurred mainly postsynaptically. Our results are indicative of important cytoskeletal changes in the mossy fiber-CA3 synapses after pilocarpine treatment, which may contribute to the excessive excitatory output in the hippocampal trisynaptic circuit.