This is a request for an ADAMHA Research Scientist Development Award, level II. The broad objectives of the research are to characterize the mechanisms which mediate activity-dependent changes in neurotrophic factor gene expression in the adult brain and to evaluate functional consequences of altered expression of the NGF-like neurotrophins following seizure activity. Prior studies by the applicant have demonstrated that seizures alter levels of mRNA for nerve growth factor (NGF) and the structurally related neurotrophins brain derived neurotrophic factor (BDNF) and neurotrophin 3 (NT3) in adult rat brain. In the proposed studies, molecular biological techniques (in situ hybridization, S1 nuclease protection assay, run-on assay) will be used to determine i) if the different mRNAs are colocalized and differentially regulated within individual neurons, ii) if mRNA synthesis is altered following seizure, iii) if growth factor mRNA changes follow, and are dependent upon, increases in c-fos expression, iv) if activity-dependent increases in NGF mRNA and BDNF mRNA become refractory to repeated stimulation, v) if changes in NT3 mRNA are linked to changes in BDNF mRNA and NGF mRNA in a variety experimental paradigms, vi) if seizures stimulate increases in the expression of interleukin-1beta mRNA in forebrain that might play a role in later increases in NGF mRNA expression, and vii) if seizure-induced changes in NGF and BDNF mRNA lead to changes in trophic activity within the forebrain. The training goal for the applicant will be to learn and gain experience in the use of molecular biological techniques needed to support in situ hybridization procedures, to prepare, subclone, and characterize new cDNAs, and to assay levels of mRNA synthesis. This is to be accomplished through increased time working in the laboratory with collaborators on the U.C.I. campus. The long range goals of the research program are i) to further rely on these powerful technical capabilities to independently study the role of activity dependent neuronal gene expression in normal brain function and ii) to identify lines of trophic communication in brain which are necessary for normal function and might be manipulated to promote neuronal viability.
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