Abstracts

Rationale: Status epilepticus (SE) increases the risk for the development of unprovoked recurrent seizures and temporal lobe epilepsy. Neuropathological changes underlying neuronal hyperexcitability following SE include loss of hippocampal dendritic spines and synaptic structures. However, the molecular mechanisms that directly lead to this dendritic and synaptic decline remain elusive. Recent studies indicate that signaling through the C1q-C3 proteins of the classical pathway of the immune complement system mediate synapse elimination in the developing brain. C1q initiates activation of the classical pathway and opsonizes cellular structures for microglia to phagocytize and eliminate. Understanding the potential role that the classical complement pathway may play on the SE-induced dendritic pathology requires information on the extent to which C1q and C3b are altered and their distribution. Therefore, the objective of this study was to quantify SE-induced changes in the level and distribution of C1q and C3 proteins. Methods: SE was induced in with pilocarpine and stopped after 1 hour with diazepam. Hippocampi were collected at various time points after SE (3-, 14-, 21- and 35-days). Immunohistochemistry (IHC) and Western blots (WB) were performed with antibodies against C1q and C3 to identify and quantify subcellular and regional changes. Densitometry analysis along with analysis of variance was used to determine statistical significance. Results: IHC showed that C1q immunoreactivity was localized in the hippocampus of both control and SE rats. In controls, C1q signal was evident within the stratum lacunosum moleculare while in SE samples intense C1q signal was mainly localized to the stratum radiatum. Densitometry analysis revealed a significant increase in the protein levels of C1q in the CA1 region 14 days after SE compared to controls (p < 0.05). WB showed that in SE samples the C3 protein was cleaved into C3ba, C3bߠand iC3b when compared to control samples at all time points investigated. Densitometry analysis revealed a significant increase in the protein levels of the all C3 fragments after SE compared to controls (p < 0.05). Conclusions: C1q is the initiating protein of the classical pathway which leads to the cleavage of C3 into its biologically active forms (C3a, C3ba, C3bߠand iC3b). Therefore, the observed increase in C1q levels that paralleled an increase in C3 cleavage products supports activation of the classical complement pathway in the hippocampus in response to SE. Additional studies are underway to determine the role of this pathway in potentially modulating dendritic and synaptic alterations associated with SE. Funding: AES 411837 (ALB)