Estrogens can be formed and metabolized in their target organs. The critical linkage between estrogen metabolism and action in cells and tissues is, however, poorly understood. During the next 5 year period we propose to continue studies on the role of locally formed estrogens and catechol estrogens, and to compliment our biochemical studies on endocrine control mechanisms with ultrastructural characterization of the sites of estrogen synthesis and action. The latter, long term goal will involve the development of model system utilizing the placenta and hypothalamus. We will address three main questions: 1. What is the physiologic role and mechanism of action of a group of major estrogen metabolites, the catechol estrogens (CE) in vivo and in vitro hypothalamic formation of CE will be studied using radiolabelled estrogens and impeded CE precursors (C-2, C-4, C-11 substituted estrogens). The role of CEs in pregnancy will be examined by RIA measurement of amniotic fluid, fetal blood and maternal blood CE levels and incubation of placental homogenates with estrogens. Effects of CEs on implantation and on rat embryos in culture will be studied. 2. Are estrogens formed, metabolized and active in the same cell, or is more than one cell required. Biochemical techniques and specific antisera to enzymes (aromatase, 17 Beta-estradiol dehydrogenase) and steroid receptors (ER and PR) will be used to determine which cells in cultures of human trophoblast contain these enzymes and receptors, and to localize them in subcellular compartments. 3. Are there specific changes in intracellular organelles and membranes organization which accompany estrogen action Cells from microdissected rat hypothalamic nuclei will be observed by transmission electron microscopy, freeze-fracture, filipin-sterol complexing and lectin binding for perikaryal and membrane organization changes induced during estrogen action. Our overall goal remains the elucidation of the relationship between sex steroids and endocrine sensitive tissues. This research plan will allow studies on estrogen metabolism and action in individuals cells. It is of importance for the understanding and medical management of implantation, fetal development and normal and disordered reproduction.
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