Abstracts

Rationale: Kv1.1 knockout (KO) mice model temporal lobe epilepsy with comorbid sleep disorders. We have shown these animals exhibit altered diurnal rhythmicity and disrupted sleep architecture. Our previous studies have also shown that there is a significant increase in the number of cells expressing orexin, a wake-promoting neuropeptide, in the lateral hypothalamus of the KO mice when compared to their wildtype (WT) littermates. Almorexant, a dual orexin receptor antagonist, improves sleep and significantly reduces seizures. Epilepsy onset occurs on approximately postnatal day (P)22 and seizure severity worsens with age. Here, we determined whether there is an ontological development of the sleep disruptions and orexin levels that correlate with epilepsy onset and progression.Methods: Orexin-positive cells in the lateral hypothalamus were labeled with immunohistochemistry and counted using unbiased stereology. Peripheral Orexin A levels were measured in blood samples were collected from KO and WT mice at using a Chemiluminescent EIA Kit. The relative luminescence intensity of each well was measured using a plate reader and the concentration of Orexin A was determined using a standard plot. Behavioral rest-activity cycles were assessed with integrated radio telemetry technology and switch-closure activity monitoring from ~P22 to ~P54. Infrared beam breaks by activity were monitored in 3-min epochs and scored on a scale of 0–150. We calculated 3 actigraphic endpoints during the rest period: 1) total number of rest and wake epochs 2) total actigraphic counts and 3) total transitions between rest and wake epochs.Results: Orexin positive cells increase from 86.46 ± 4.893 at P21 (age of seizure onset) to 168.8 ± 2.552 at P45 (an age of heightened seizure frequency and severity). Peripheral orexin levels were similar in WT and KO mice at P34 (49.853 pg/ml of plasma). In KO mice, orexin increases by approximately 89% with age. In contrast, WT orexin levels diminish with age. Our preliminary actigraphic data indicates that there is no difference in the number of rest or wake epochs during the rest period in young KO and WT mice. However, by P35 the amount of activity is 2-fold higher in KO mice when compared to WT control which becomes 5-fold higher by P54.This drastic difference is due to less time KO mice spend resting and more time spent awake, as indicated by a 91% decrease in the ratio of rest:wake epochs with age (from 1.69 at P23 to 0.14 at P54). When normalized to WT control, the KO mice show a 45% increase in the number of transitions between sleep and wake from age P23 to P54.Conclusions: Our data indicates that the sleep disruptions, brain orexin levels and peripheral orexin levels increase with age in the Kv1.1KO mice and correlate with epilepsy onset and progression.