Abstracts

Tuberous Sclerosis Complex (TSC) is a multi-organ genetic disease with significant neurological morbidity, including mental retardation, autism, and epilepsy. Recent evidence suggests a possible role of astrocytes in contributing to epileptogenesis in TSC. Conditional inactivation of the [italic]Tsc1[/italic] gene in glia has previously been shown to cause impaired astrocytic glutamate transport and epilepsy in mice ([italic]Tsc1[/italic]^GFAPCKO mice). We hypothesized that defective astrocytic glutamate transport may promote epileptogenesis in [italic]Tsc1[/italic]^GFAPCKO mice by potentiating glutamatergic synaptic transmission and/or by causing excitotoxic neuronal death. Thus we investigated glutamatergic synaptic function and neuronal death in [italic]in vitro[/italic] and [italic]in situ[/italic] preparations from [italic]Tsc1[/italic]^GFAPCKO mice. In astrocyte-neuronal co-cultures, pharmacologically-isolated AMPA- and NMDA-mediated excitatory autaptic currents were examined in wild-type neurons cultured with control or [italic]Tsc1[/italic]^GFAPCKO astrocytes by whole-cell voltage clamp techniques. Glutamatergic excitatory postsynaptic currents were also measured by whole-cell recording in CA1 neurons from hippocampal slices from control and [italic]Tsc1[/italic]^GFAPCKO mice. Long-term potentiation was studied in the CA1 region of hippocampal slices with extracellular recording. Excitotoxic cell death was examined in neocortex and hipocampus of 1 and 3 month old control and [italic]Tsc1[/italic]^GFAPCKO mice by TUNEL, caspase-3, and Fluoro-Jade B immunofluorescent assays. Despite decreased glutamate transport of [italic]Tsc1[/italic]^GFAPCKO astrocytes, no significant difference was detected in glutamatergic synaptic transmission during low frequency stimulation in either culture or slice preparations from [italic]Tsc1[/italic]^GFAPCKO mice. However, long-term potentiation due to high-frequency synaptic stimulation was impaired in hippocampal slices from [italic]Tsc1[/italic]^GFAPCKO mice. In TUNEL, caspase-3, and Fluoro-Jade B assays, neuronal death was detected in both neocortex and hippocampus of [italic]Tsc1[/italic]^GFAPCKO, but not control, mice at both 1 and 3 months of age. These results suggest that impaired astrocyte glutamate transport in [italic]Tsc1[/italic]^GFAPCKO mice can affect glutamatergic synaptic transmission under high-frequency stimulation and stimulate excitotoxic cell death, which may contribute to epileptogenesis in these mice. (Supported by NIH K02NS045583 and U.S. Army Medical Research and Materiel Command grant DAMD17-03-1-0073.)