Abstracts

Rationale: The Kcna1-null mouse, a Kv1.1 potassium channel knockout (KO) mouse, is a well-defined model for temporal lobe epilepsy. We have previously shown that KO mice also have altered sleep architecture compared to wild-type (WT) mice, including decreased NREM and REM sleep and increased wakefulness during the rest phase. We have also shown evidence of pathology in the lateral hypothalamus (LH), including seizure-like activity recorded by depth electrode, increased blood-brain barrier (BBB) permeability, and increased incidence of reactive astrocytes. Orexin (also known as hypocretin) neurons, located in the LH, are known to stimulate arousal pathways and contribute to regulation of normal sleep architecture. KO mice show an increased number of orexin neurons at postnatal day 40, a possible explanation for the increased wakefulness and decreased sleep. However, it is unknown if this is a result of increased proliferation of orexin neurons or a decrease in normal pruning, both of which are known to occur in epileptic hippocampus. Here, we use bromodeoxyuridine (BrdU), a synthetic nucleoside used to label actively dividing cells and their progeny, to determine changes in neuronal proliferation in the LH of a chronic epilepsy model.Methods: KO mice and WT littermates were injected with sterile BrdU solution (50mg/kg) twice, six hours apart, on post-natal day 28, 35, or 38. Injected mice were then sacrificed 2 days later, on post-natal day 30, 37, or 40. 30 µm frozen brain sections were then labeled with BrdU antibody and co-labeled with orexin antibody. Sections from separate cohort of animals at post-natal day 12 and 21 were also labeled for orexin to determine orexin-positive neuron number before onset of seizures. BrdU and orexin-positive cell numbers were determined using unbiased stereology.Results: In contrast to previous experiments looking at post-natal day 40, there was no significant difference in the number of orexin-positive neurons between KO and WT animals at post-natal day 12 or 21 (n=6 animals per group). Preliminary data shows a trend to an increase in neurons labeled with BrdU in the LH of KO mice compared to WT control.Conclusions: In this study, we examined proliferation of neurons, including orexin neurons, in the LH of Kv1.1 knockout mice, a chronic epilepsy model. While orexin neurons in the LH are increased in KO mice compared to WT at post-natal day 40, younger mice show no significant difference between genotypes. Increased BrdU labeling in the KO LH suggests an increase in proliferation of new cells, including orexin neurons, between post-natal day 30 and 40. This increase in proliferation of orexin neurons may be responsible for increased wakefulness and decreased sleep in epileptic KO mice. Further studies will need to address the cause of increased proliferation as well as the source of these new neurons.