The mechanistic response of neurotransmitter systems to a variety of long term manipulations, for example in stress or illness, is a central issue in neurobiology. The plasticity of neurotransmitter biosynthesis is much more intricately regulated than previously appreciated. This proposal will study molecular aspects of the regulation of biosynthesis of norepinephrine, a transmitter that plays a crucial role in a surprisingly large number of functions in the CNS and periphery. Tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH), two of the enzymes involved in norepinephrine biosynthesis, are regulated in response to long term physiological, pharmacological and environmental stimuli. A molecular description of their regulated expression will be invaluable both as a model for neurotransmitter regulation in general as well as in the specific elucidation of the mechanisms which regulate the catecholaminergic neuronal system. This project will: I. Delineate the differences between membrane-bound and soluble forms of DBH. Determine what is the mode of membrane attachment of DBH and the nature of the post-translational processing from the membrane-bound to the soluble form. II. Determine the regulation of expression of DBH and TH in vivo in tissues which synthesize norepinephrine, specifically in the adrenal medulla and in the locus coeruleus (LC) of the central nervous system. III. Delineate the regulation of expression of DBH and TH in PC12 cells and primary chromaffin cells. Study the regulation of mRNA levels and transcription of the DBH gene in response to growth factors, cyclic AMP analogs and glucocorticoids and combinations of these treatments. IV. Define upstream regulatory elements in the rat DBH gene. I postulate that there are regulatory elements for cAMP, glucocorticoids and growth factors. Elements for tissue specific expression will be determined. The results of this study will provide a detailed mechanistic interpretation of the long term regulation of norepinephrine biosynthesis, which will have important implications for the responsiveness of the nervous system to prolonged stress and disease, such as hypertension, schizophrenia, depression and dementia of the Alzheimer's type.
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