Progress toward Understanding Epileptogenesis in Tuberous Sclerosis Complex: Two Hits, No Outs, and the Eker Rat is up to Bat

Abstract

Abnormal Cortical Cells and Astrocytomas in the Eker Rat Model of Tuberous Sclerosis Complex Takahashi DK, Dinday MT, Barbaro NM, Baraban SC Epilepsia 2004;45:1525–1530 Purpose In patients with tuberous sclerosis complex (TSC), a wide range of neurologic abnormalities develop, including mental retardation and seizures. Brains from TSC patients are characterized by the presence of cortical tubers, large dysmorphic neurons, and abnormal cytomegalic cells. Although analysis of human TSC brain samples led to the identification of these abnormal cell types, very little is known about how these cells function. In an effort to model TSC-associated CNS abnormalities (and ultimately to analyze the electrophysiologic properties of abnormal cells), we examined Eker rats carrying a Tsc2 mutation. Anatomic studies, including standard histologic stains and immunocytochemistry, were performed on young Eker rats exposed to a carcinogen in utero or on aged untreated Eker rats (18–24 months old). Methods Pregnant TSC2+/– females were injected once a day with hydroquinone, and offspring were killed at postnatal day P14 or P28. Coronal tissue sections throughout the CNS were prepared and stained for cresyl violet. In separate studies, brains of old untreated Eker rats were sectioned for anatomic analysis by using standard immunohistochemical techniques. Results Tissue sections stained with cresyl violet did not reveal any gross differences between hydroquinonetreated Eker ( Tsc2 Ek/+) rats and siblings ( Tsc2+/+). However, two classes of abnormal giant cells were observed in brain sections from untreated aged Eker rats: 1) large dysmorphic pyramid-like cells immunoreactive for NeuN, tuberin, and EAAC-1 in layers IV to VI; and 2) abnormal cytomegalic cells immunoreactive for glial fibrillary acidic protein, vimentin, and nestin in deep cortical layers or along the white matter. In addition, large subependymal astrocytomas were observed in four animals. Conclusions Our data suggest that cortical tuber formation in Eker rats is a rare event and that prenatal exposure to a nongenotoxic carcinogen such as hydroquinone is not sufficient to induce tuber formation. However, with advanced age, an increased likelihood of astrocytoma formation and the emergence of dysmorphic neurons and cytomegalic cells in the Eker rat brain might exist; each of these abnormalities mimics those seen clinically and could contribute to neurologic problems associated with TSC. Further analysis of this rodent model may be warranted. Morphology of Cerebral Lesions in the Eker Rat Model of Tuberous Sclerosis Wenzel HJ, Patel LS, Robbins CA, Emmi A, Yeung RS, Schwartzkroin PA Acta Neuropathol (Berl) 2004;108:97–108 Tuberous sclerosis (TSC) is an autosomal dominant disorder, caused by mutations of either the TSC1 or TSC2 gene. Characteristic brain pathologies (including cortical tubers and subependymal hamartomas/giant astrocytomas) are thought to cause epilepsy, as well as other neurologic dysfunction. The Eker rat, which carries a spontaneous germline mutation of the TSC2 gene ( Tsc2+/+), provides a unique animal model in which to study the relation between TSC cortical pathologies and epilepsy. In the present study, we analyzed the seizure propensity and histopathologic features of a modified Eker rat preparation, in which early postnatal irradiation was used as a “second hit” stimulus in an attempt to exacerbate cortical malformations and increase seizure propensity. Irradiated Eker rats had a tendency toward lower seizure thresholds (latencies to flurothyl-induced seizures) than seen in nonirradiated Eker rats (significant difference) or irradiated wild-type rats (nonsignificant difference). The majority of irradiated Eker rats exhibited dysplastic cytomegalic neurons and giant astrocyte-like cells, similar to cytopathologies observed in TSC lesions of patients. The most prominent features in these brains were hamartoma-like lesions involving large eosinophilic cells, similar to giant tuber cells in human TSC. In some cells from these hamartomas, immunocytochemistry revealed features of both neuronal and glial phenotypes, suggesting an undifferentiated or immature cell population. Both normal-appearing and dysmorphic neurons, as well as cells in the hamartomas, exhibited immunopositivity for tuberin, the protein product of the TSC2 gene.