Estradiol- and Progesterone-Associated Changes in microRNA-Induced Silencing and Reduced Antiseizure Efficacy of an Antagomir in Female Mice.

About one-third of individuals living with epilepsy have treatment-resistant seizures. Alternative therapeutic strategies are thus urgently needed. One potential novel treatment target is miRNA-induced silencing, which is differentially regulated in epilepsy. Inhibitors (antagomirs) of specific microRNAs (miRNAs) have shown therapeutic promise in preclinical epilepsy studies; however, these studies were mainly conducted in male rodent models, and research into miRNA regulation in females and by female hormones in epilepsy is scarce. This is problematic because female sex and the menstrual cycle can affect the disease course of epilepsy and may, therefore, also alter the efficacy of potential miRNA-targeted treatments. Here, we used the proconvulsant miRNA miR-324-5p and its target, the potassium channel Kv4.2, as an example to test how miRNA-induced silencing and the efficacy of antagomirs in epilepsy are altered in female mice. We showed that Kv4.2 protein is reduced after seizures in female mice similar to male mice; however, in contrast to male mice, miRNA-induced silencing of Kv4.2 is unchanged, and miR-324-5p activity, as measured by the association with the RNA-induced silencing complex, is reduced in females after seizure. Moreover, an miR-324-5p antagomir does not consistently reduce seizure frequency or increase Kv4.2 in female mice. As a possible underlying mechanism, we found that miR-324-5p activity and the silencing of Kv4.2 in the brain were differentially correlated with plasma levels of 17ß-estradiol and progesterone. Our results suggest that hormonal fluctuations in sexually mature female mice influence miRNA-induced silencing and could alter the efficacy of potential future miRNA-based treatments for epilepsy in females.

MiR-324-5p inhibition after intrahippocampal kainic acid-induced status epilepticus does not prevent epileptogenesis in mice.

Background: Acquired epilepsies are caused by an initial brain insult that is followed by epileptogenesis and finally the development of spontaneous recurrent seizures. The mechanisms underlying epileptogenesis are not fully understood. MicroRNAs regulate mRNA translation and stability and are frequently implicated in epilepsy. For example, antagonism of a specific microRNA, miR-324-5p, before brain insult and in a model of chronic epilepsy decreases seizure susceptibility and frequency, respectively. Here, we tested whether antagonism of miR-324-5p during epileptogenesis inhibits the development of epilepsy. Methods: We used the intrahippocampal kainic acid (IHpKa) model to initiate epileptogenesis in male wild type C57BL/6 J mice aged 6-8 weeks. Twenty-four hours after IHpKa, we administered a miR-324-5p or scrambled control antagomir intracerebroventricularly and implanted cortical surface electrodes for EEG monitoring. EEG data was collected for 28 days and analyzed for seizure frequency and duration, interictal spike activity, and EEG power. Brains were collected for histological analysis. Results: Histological analysis of brain tissue showed that IHpKa caused characteristic hippocampal damage in most mice regardless of treatment. Antagomir treatment did not affect latency to, frequency, or duration of spontaneous recurrent seizures or interictal spike activity but did alter the temporal development of frequency band-specific EEG power. Conclusion: These results suggest that miR-324-5p inhibition during epileptogenesis induced by status epilepticus does not convey anti-epileptogenic effects despite having subtle effects on EEG frequency bands. Our results highlight the importance of timing of intervention across epilepsy development and suggest that miR-324-5p may act primarily as a proconvulsant rather than a pro-epileptogenic regulator.