Abstracts

Rationale: Pilocarpine-induced status epilepticus (SE) results in the formation of abnormalities localized to the dentate gyrus. These include sprouting of mossy fiber axons, formation of hilar basal dendrites, ectopic migration of newborn granule cells and mossy cell death. Hyperactivation of the PTEN, AKT, mTOR pathway has been implicated in driving mossy fiber sprouting, but the role of this pathway in other dentate abnormalities is not known. Here, we administered the mTOR inhibitor rapamycin to pilocarpine-treated mice to assess whether treatment would reduce the extent of these other abnormalities.Methods: Gli1CreERT2::GFP and Gli1CreERT2::tdTomato mice were used in this study. Tamoxifen was administered once-weekly for five weeks beginning when the mice were three-weeks-old. Tamoxifen induces GFP or tdTomato expression in newborn dentate granule cells. When the mice were eight-weeks-old, SE was induced with pilocarpine (controls were given saline). Beginning the day after pilocarpine treatment, 6mg/kg rapamycin i.p. (or vehicle) was given once-daily for 6 weeks. This study design resulted in 4 treatment groups: 1) SE/rapamycin, 2) SE/vehicle, 3) control/rapamycin, and 4) control/vehicle. Mice were sacrificed at fourteen weeks-of-age, the day following the final rapamycin treatment. Brain sections were immunostained for phospho-S6, ZnT3, Prox1 and GluR2/3.Results: SE resulted in an 80% increase in phospho-S6 immunoreactivity within the granule cell body layer relative to controls (p=0.002). Rapamycin treatment reduced phospho-S6 expression by 89% in control mice (p<0.001) and 78% in SE mice (p<0.001). Rapamycin reduced mossy fiber sprouting by 50% relative to epileptic mice receiving vehicle (p=0.042). By contrast, rapamycin treatment produced only a trend towards reduced numbers of newborn cells with basal dendrites (SE/vehicle 55.8±5.6% newborn cells exhibited basal dendrites; SE/rapamycin, 39.2±5.6%, p=0.066) and had no effect on the total number of ectopic cells (SE/vehicle 21.4±2.1 cells/mm3 (x1000), SE/rapamycin 23.1±2.2 cells/mm3 (x1000), p=0.566) or surviving mossy cells (SE/vehicle 10.9±6.1 cells/mm3 (x1000), SE/rapamycin 13.1±6.3 cells/mm3 (x1000), p=0.252).Conclusions: These data indicate that, aside from the established effect on mossy fiber sprouting, rapamycin has limited efficacy in preventing aberrant granule cell integration and mossy cell death. The anti-epileptogenic effect of rapamycin in rodent models of epilepsy, therefore, is likely mediated by other mechanisms.