Abstracts

Rationale: Animal and human studies of temporal lobe epilepsy have long implicated dentate granule cells (DGCs) as mediating many epileptogenic changes. We recently demonstrated that selective deletion of phosphatase and tensin homologue (PTEN) from 10-25% of late-generated DGCs was sufficient to produce a profound epilepsy syndrome (Pun et al., Neuron, 2012). PTEN deletion from granule cells leads to hyperactivation of the mammalian target of rapamycin (mTOR) pathway, and our PTEN knockout (KO) model reproduces abnormalities observed in temporal lobe epilepsy. These findings support the hypothesis that accumulation of abnormal granule cells may mediate temporal lobe epileptogenesis. Although abnormal DGCs are highly implicated, the percentage of abnormal cells in epilepsy is often much less than 25%. To better assess the clinical significance of abnormal cells, therefore, we designed a protocol to delete PTEN from approximately 5% of DGCs. This is closer to the number of abnormal cells present in temporal lobe epilepsy models, and we hypothesized that this reduced number of cells would still be sufficient to cause the disease. Methods: Gli1-CreERT2 hemizygous, PTENfl/fl mice received a subcutaneous injection of 250 mg/kg of tamoxifen at three weeks of age. Beginning at six weeks of age, mice were wirelessly monitored 24/7 by video/EEG using electrodes placed in either neocortex or hippocampus. Seizure onset, frequency, and duration were determined post hoc. Following 12 weeks of continuous EEG monitoring, each mouse was sacrificed for histological studies. Double immunostaining of brain sections was conducted for PTEN and NeuN to determine the percentage of PTEN KO cells. ZnT-3 antibodies were used to reveal sprouted mossy fiber axons.Results: PTEN deletion from approximately 5% of the DGC population was sufficient to cause epilepsy. Animals exhibited spontaneous seizures as early as 7 weeks of age, while no seizures were observed in littermate controls. Initially, animals exhibited localized hippocampal seizures. Around 14 weeks of age cortical seizures were observed, indicating that the epilepsy in these animals progresses from partial to generalized seizures. Interesting, mossy fiber sprouting was absent from these animals, demonstrating that this particular pathology is not required for spontaneous seizures in this model. Conclusions: Animals in which PTEN was deleted from approximately 5% of DGCs exhibit spontaneous seizures. This finding indicates that a load of abnormal granule cells similar to that observed in status epilepticus models of epilepsy is sufficient to drive epileptogenesis. The findings also support the conclusion that initial seizures in the model are restricted to hippocampus, while later seizures recruit hippocampus and cortex. Finally, the disease was less severe in mice with 5% PTEN KO DGC relative to animals in our previous study with 10-25% PTEN KO DGC, with later onset of cortical seizures and reduced mortality. These findings suggest that epilepsy phenotype and disease progression may be dependent upon the number of abnormal granule cells.