The overall objective of this project is to gain an understanding of the molecular mechanisms underlying the development of the mammalian central nervous system. Among the many physiological factors that influence the central nervous system, thyroid hormone is known to be essential for proper development. Thyroid hormone deficiency during a short critical period produces severe neurological defects in humans and in experimental animals. The actions of thyroid hormone are mediated through nuclear receptors, which regulate the expression of specific target genes in response to hormone binding. Although much is known about the function of thyroid hormone receptors, the genes regulated by these receptors in the developing brain are largely unknown. The main objective of the research proposed here is to identify and characterize genes whose expression in the brain is regulated by thyroid hormone during the period of time in which thyroid hormone is known to be critical for proper development. Potential target genes have been identified using subtractive hybridization coupled with polymerase chain reaction amplification to isolate genes that are up-regulated in response to thyroid hormone in developing rat brain. Developmental and tissue specific patterns of expression for the candidate genes will be determined,using Northern analysis and in situ hybridization to identify those most likely to have a role in central nervous system development. The complete nucleotide sequence will be determined for selected genes and the primary amino acid sequence deduced. Antibodies will be produced to determine expression patterns and subcellular localization of the proteins using Western analysis and immunohistochemical methods. An important long term goal of this project is to determine the functional role of thyroid hormone responsive gene products in nervous system development. A detailed understanding the mechanisms by which thyroid hormone effects brain development should be of value in ultimately attempting to treat developmental and degenerative diseases of the central nervous system.
