Abstracts

Rationale: Contactin-associated protein like 2 (Cntnap2) has been linked to autism spectrum disorders and epilepsy. Cntnap2 knockout (KO) mice display deficits in social and repetitive behavior at early age, have alterations in neuronal excitability and develop spontaneous recurrent seizures later in life. However, the mechanisms of Cntnap2 deletion leading to altered neuronal excitability and epilepsy and potential therapeutic targets are unknown. MicroRNAs (miRNAs) regulate post-transcriptional expression of protein-coding mRNAs and are crucial regulators of genes involved in epilepsy. For example, miR-324-5p inhibition provides neuroprotection in a mouse model of epilepsy, whereas miR-218-5p is downregulated in epilepsy and its inhibition may thus be proconvulsive. So far, it is unknown if microRNA-induced silencing plays a role in Cntnap2-associated epilepsy. Here, we addressed this question by investigating the effect of antisense-mediated inhibition of miR-324-5p and miR-218-5p on seizure susceptibility and interictal spike activity in adult and older Cntnap2 KO mice. Methods: Mice were genotyped and used at adult (4-5 months) and older (12-16 months) age points to first investigate spike and seizure development. Cortical EEG recordings using a wireless EEG system were employed to confirm seizures and spike occurrence. Post confirmation of seizures, older mice were injected with miR-324-5p (n=7), miR-218-5p (n=9) or scrambled (n=8) antagomirs, respectively, and were EEG monitored for 1 week. Later mouse brains were perfused and were collected for future histological analyses. Results: Younger mice displayed increased spike activity, but no spontaneous or handling-induced seizures were observed in the EEG. Older mice displayed both epileptiform spikes and spontaneous and handling-induced seizures. EEG analyses showed on average increased numbers of seizures (non-significant) in miR-218 antagomir-injected mice compared to 324-5p or scrambled control (one-way ANOVA, p>0.05) but no neuroprotective effect of miR-324-5p inhibition was observed. No difference in the average spike count or spike trains were observed (one-way ANOVA, p>0.05). An increase in seizure associated mortality post 218-antagomir injection was observed. Conclusions: These results indicate a selective miR-218-dependent mechanism regulating hyperexcitability and seizure susceptibility in Cntnap2 mouse model which could be a potential mechanism to regulate excitability in Cntnap2-associated epilepsy. Notably, in contrast to our previous studies in a mouse model of acquired epilepsy, miR-324-5p inhibition did not reduce seizure frequency in this genetic form of epilepsy. Ongoing work is focused on tissue analysis and investigating antagomir-mediated effects on spikes characteristics in younger KO mice. Funding: NIH grants R01NS092705