This proposal, a response to the program announcement for exploratory (R21) grants in pediatric brain disorders, focuses on the issue of epileptogenesis in tuberous sclerosis. Tuberous sclerosis (TS) is an autosomal dominant disorder characterized by the formation of hamartomatous growths in multiple organ systems, including kidney, skin, and brain. Recent studies suggest a TS incidence of 1 in 6,000 live births. The most debilitating of the effects of TS are its nervous system manifestations, including epilepsy and mental retardation. It has been estimated that over 80% of TS patients have epilepsy, often occurring early in development as such difficult-to-control syndromes as infantile spasms. While insights into TS have been greatly advanced by our understanding of the underlying genes (TSC1 and TSC2), the connections between TS gene mutations and brain hamartomas (and particularly cortical tubers), and between tubers and seizure development, remain unclear. The absence of an animal model of TS with a CNS tuber and seizure phenotype has made it difficult to study these relationships. In this proposal, we exploit the Eker rat, a TSC2 +/- """"""""carrier"""""""", to address two aspects of the tuber/epilepsy complex. First, we will examine the hypothesis that cytologically-abnormal cells characteristic of cortical tubers are a result of homozygous mutations at the TSC2 locus (TSC2 -/-); that is, loss of heterozygosity (LOH) is a critical feature of brain tuber formation, just as it is for tumors in other organs (e.g., kidney). TSC2 -/- cells will be obtained from embryonic CNS tissue (from +/- x +/- Eker crosses) and maintained in culture. Morphological and electrophysiological tools will be used to characterize these cells (vs. TSC2 +/- and +/+ cells). Differentiation of cultured TSC2 -/- neuroblasts will be examined under different culture conditions, their response to experimental challenge (e.g., irradiation, excitotoxicity) determined, and their effects on co-cultured tissue assessed. Second, to test the hypothesis that TSC -/- cells give rise to cortical tubers, cultured cells (containing a fluorescent marker gene) will be transplanted into normal rat brain. Morphological features (e.g., tuber formation, circuitry reorganization) of transplanted cortex, as well as its electrical excitability and the animal's seizure susceptibility, will be determined. These studies, while not yet probing the molecular mechanisms through which TSC2 mutations give rise to structural malformations in the developing animal, will address critical phenomenological cause-effect relationships that will serve as the basis for future mechanistic studies.
| Yeung, Raymond S (2003) Multiple roles of the tuberous sclerosis complex genes. Genes Chromosomes Cancer 38:368-75 |
