MOLECULAR REGULATION OF THE PROENKEPHALIN GENE IN VIVO

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The endogenous opioid peptides bind to th e same receptors as morphine and other opiate compounds and they are thought to be involved in pain relief and opiate addiction. Three major groups of opioid peptides exist: enkephalins, endorphins and dynorphins. Each is derived from a distinct gene to produce specific precursor peptides. The subject of this proposal is the regulation of the proenkephalin (Penk) gene and the biosynthesis of enkephalins. The strategy is to determine factors that regulate opioid peptide gene expression and to ultimately develop new approaches for the treatment of pain and opiate drug abuse; The Penk gene is expressed in both the central nervous system and in the adrenal medulla. In the rat adrenal medulla nerve impulse activity both inhibits and stimulates how basal levels of gene expression. This process requires the presence of glucocorticoids. In contrast, Penk gene expression in the hamster adrenals are high, positively regulated by nerve activity like most mammals and appear to be glucocorticoid independent. However, increases in Penk gene expression in the developing hamster adrenal appear to be glucocorticoid dependent. Thus, a combination of nerve impulse activity and hormones regulates Penk gene expression which varies with development, species and treatment. To extend these observations, this proposal will (1) use biochemical, immunocytochemical and in situ hybridization techniques to examine developmental changes in hamster Penk gene expression and evaluate the influence of glucocorticoid and synaptic innervation (2) determine glucocorticoids role int he induction of Penk gene expression in the hamster. [It is hypothesized that basal Penk gene expression in the hamster is glucocorticoid independent and induced Penk gene expression due to treatment or development are glucocorticoids dependent.] (3) use pharmacological manipulation to determine whether multiple transsynaptic and/or receptor mediated mechanisms regulate gene expression. (4) define the cis and/or trans elements that regulate Penk gene expression in the adult and developing hamster using mobility shift analysis, DNase I footprint and/or methylation interference assays and (5) develop conditions, in vitro, that will maintain the hamster adrenal in organ culture and examine under controlled conditions neurotropic and hormonal factors regulating Penk gene expression and peptide release. An examination of the disparity in Penk gene expression between species (hamster and rat), the changes during development and the effects of treatment in vivo will produce new insights on specific issues in Penk gene regulation.