A role for the classical complement pathway in hippocampal dendritic injury and hippocampal dependent memory deficits in a model of acquired epilepsy
Status epilepticus (SE) triggers pathological changes to hippocampal dendrites that may promote epileptogenesis. The microtubule associated protein 2 (Map2) helps stabilize microtubules of the dendritic cytoskeleton. Recently, we reported a substantial decline in Map2 that coincided with robust microglia accumulation in the CA1 hippocampal region after an episode of SE. A spatial correlation between Map2 loss and reactive microglia was also reported in human cortex from refractory epilepsy. New evidence supports that microglia are guided by proteins of the classical complement pathway (C1q and C3) to prune dendritic structures. Furthermore, components of complement have been shown to be upregulated in human and experimental epilepsy. Thus, to identify a potential role of the classical complement pathway in SE-induced Map2 and microglial changes, we characterized the spatiotemporal profile of these events. We used immunohistochemistry to determine the distribution of Map2 and the microglia marker IBA1 in the hippocampus after pilocarpine-induced SE from 4 hours to 35 days. We found a decline in Map2 immunoreactivity in the CA1 area that reached minimal levels at 14 days post-SE and partially increased thereafter. In contrast, maximal microglia accumulation occurred in the CA1 area at 14 days post-SE. We then mapped the spatiotemporal profile of C1q using immunohistochemistry at 3-35 days after SE, where substantial Map2 and microglial alterations were observed. We used western blot to determine the levels of C3 and its cleavage products. C1q and C3 were both increased in the hippocampus at 14 days after SE, when Map2 and microglia changes were most profound. Our data indicate that SE-induced Map2 and microglial changes parallel each other’s spatiotemporal profiles. These findings also suggest a potential role for the classical complement pathway in SE-induced Map2-microglial interactions. Cognitive deficits are often associated with epilepsy and experimental models of SE and TLE, particularly because the hippocampus is often vulnerable to injury in TLE. To test the potential role of the classical complement pathway in SE-induced hippocampal injury and hippocampal-dependent cognitive deficits, we used a pharmacological approach to block the classical complement pathway with the C1 esterase inhibitor (C1-Inh). This drug was given at 4-, 24-, and 48-hours after SE. Tissue was collected at 3-, 14-, 25-, and 35-days after SE and a separate cohort of animals were used to test the effects of C1-Inh on cognitive performance using the novel object recognition (NOR) and the Barnes maze (BM) tests. We found that C1-Inh had no effect on hippocampal Map2 loss after SE when compared with hippocampi from the SE groups. Furthermore, microgliosis was exacerbated with C1-Inh treatment, with a significant increase lasting from 3 days to 35 days after SE. We replicated previous studies showing a deficit in the NOR and BM tests after SE. However, C1-Inh treatment did not attenuate these deficits, and in fact exacerbated the effects in the BM. Taken together, these data support the idea that the classical complement pathway may play a role in epileptogenesis, inflammation, and cognitive deficits, but more experiments are needed to understand what the specific proteins and mechanisms are involved. Future experiments will look at alternative targets within the classical pathway after SE.
Brewster, Purdue University.
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