Abstracts

Tuberous Sclerosis (TSC) -related seizures are refractory to conventional antiepileptic drugs requiring surgical excision. As levels of expression and subunit composition of AMPA (AMPAR) and NMDA (NMDAR) subtypes of glutamate receptors affects neuronal excitability, we hypothesized that 1) AMPARs and NMDARs are expressed in dysplastic neurons, and 2) subunit composition is altered compared to cells in perituberal tissue.
Cortical tubers from 5 patients ages 4-7 years were obtained during epilepsy surgery, in accordance with the Clinical Research Committee at Children[rsquo]s Hospital, Boston. All patients met clinical criteria for TSC with neuropathologic confirmation of the diagnosis. Tissue was fixed in 4% paraformaldehyde, cut at 20 [mu]m and immunocytochemically double labeled with the neuronal cell markers NeuN, non-phosphorylated neurofilament (SMI 311), Snap 25 and GABA, and the glial markers vimentin and GFAP in combination with antibodies to AMPAR subunits GluR1, GluR2 and GluR4 and NMDAR subunits NR2A and NR2B.
Immunolabeled dysplastic neurons were distinguished from normal neurons and giant cells using morphologic criteria (cell size, orientation, and process morphology) and reactivity for cell-specific markers. The majority of the abnormal cells in tissue specimens examined were dysplastic neurons. Dysplastic neurons expressed GABA and Snap 25 indicative of predominantly excitatory differentiated neurons with synaptic contacts. Dysplastic neurons showed strong GluR1, GluR2 and GluR4 immunoreactivity. However, GluR1 and GluR4 were present in the cell body and processes, whereas GluR2 appeared to be restricted to the cell body. NMDARs were also expressed in dysplastic neurons and giant cells. Notably was the strong immunopositivity for NR2B observed exclusively in cytomegalic, as well as normo-sized, disoriented dysplastic neurons. Giant cells were characterized by the presence of multiple nuclei and immunoreactivity for vimentin but not Snap 25. In contrast to dysplastic neurons, giant cells homogenously expressed GluR1, GluR2, and GluR4.
1.There is differential expression of the specific AMPAR and NMDAR subunits between dysplastic neurons and giant cells, suggesting functional differences. 2. GluR1 and GluR4 are strongly expressed in dysplastic neurons in both the cell body and processes, while GluR2 immunoreactivity is limited to the cell body, suggesting selective presence of calcium permeable AMPARs on the processes. Activation of these AMPARs could enhance epileptogenesis via calcium activated pathways.3. The NMDAR subunit NR2B is highly expressed in dysplastic neurons and may contribute to their hyperexcitability, since this subunit is associated with enhancement of current flow through the receptor, depolarization and increase in intracellular calcium.
[Supported by: NS31718 (FEJ); Boston Neurosurgical Fnd (PMB)]