Abstracts

Tuberous sclerosis complex (TSC) represents an autosomal dominant disorder frequently manifesting with early life epilepsy and mental retardation. An inactivating mutation in either of the two tumor suppressor genes TSC1 or TSC2 leads to the development of hamartomas in multiple organs, including the brain. We previously demonstrated specific alterations in glutamate receptor (GluR) subunit expression in human cortical tubers, which may contribute to enhanced network excitability and seizures in TSC (Epilepsia; 44 (Suppl.9): 36). To further investigate the role of abnormal glutamatergic transmission in TSC-related seizures we extended these studies to a mutant mouse model in which [italic]Tsc1[/italic] gene was selectively inactivated in neurons. We hypothesized that similar to the human 1) the animal model will exhibit abnormal cortical morphology and 2) GluR subunit expression on the abnormal neurons will be altered. [italic]Tsc1 ^c/[/italic]^c synapsin cre[italic]⁺[/italic] transgenic animals were generated by using the cre recombinase method to convert a conditional ([ldquo]floxed[rdquo]) allele (c) of the Tsc1 to the null allele under the control of the neuron specific promoter synapsin. Brain tissue from mutant (n=7) and wild type (n=6) animals was fixed in 4% paraformaldehyde, cut at 50 [mu]m and immunocytochemically double labeled with neuronal and glial cell markers, in combination with antibodies against c-Fos, AMPAR subunits GluR1-4 and NMDAR subunits NR1, NR2A and NR2B. Immunostaining using neuronal (NeuN, SMI 311) and glial (GFAP, vimentin) markers revealed diffuse alteration of brain architecture and the presence of abnormal large neurons throughout the neocortex, subcortical white matter, hippocampus, thalamus and brain stem The dysplastic neurons displayed intense GluR1, NR1 and NR2B immunoreactivity, while GluR2, GluR3, GluR4 and NR2A subunits were expressed only at low levels. In contrast, normal neurons exhibited predominantly GluR2, GluR3, NR1 and NR2A subunits. Relative to the wild-type, c-Fos immunoreactivity in the mutant mouse brain was significantly increased. 1. In the[italic]Tsc1 ^c/[/italic]^c synapsin cre[italic]⁺ [/italic]mouse, hamartin loss in neurons resulted in abnormally large cells, resembling the dysplastic neurons of human cortical tubers. 2. Dysplastic neurons appeared to express AMPAR subunits suggestive for Ca2+ permeability (high GluR1: GluR2 ratio) and NMDAR subunits consistent with enhanced current flow through the receptor (increased NR2B/NR2A ratio). 3. Elevated c-Fos immunoreactivity in the abnormal neurons suggested active contribution to the high epileptogenicity of the lesion. 4. Our results, showing similarities between human and mouse TSC epileptic brain with respect to GluR expression and cell specific markers, reinforce the validity of the animal model in studying the mechanisms of epileptogenesis in TSC patients. (Supported by Epilepsy Fdn (DMT), TSA (DMT and LR), NS31718 (FEJ).)