Delineation of Hypothalamic Hypophysiotrophic Factors

Yasuda, Naoki

Abstract

Structures of corticotropin-releasing factor (CRF) and growth hormone-releasing factor (GRF) have been elucidated during the past few years. We have developed specific antisera against CRF (sheep, rat/human) and GRF (rat, human) and established respective RIAs. Preliminary results indicated that perifused rat hypothalami and human placental trophoblasts secrete CRF- and/or GRF-like peptides in measurable concentrations. The main proposals in this application relate to CRF and GRF produced at a eutopic site (hypothalamus) and """"""""ectopic"""""""" site (placenta). Because of the multi-factorial nature of the hypothalamic regulation of ACTH and GH secretion, we will study vasopressin (AVP) and oxytocin (OXT) with CRF, and somatostatin with GRF. Whether there are any feedback relationships between the target hormones and these hypophysiotrophic hormones, whether there are any interactions among different hypophysiotrophic hormones, or whether secretion of various hypothalamic or placental hormones are dependent on calcium and/or cyclic AMP will be studied using the perifusion of rat hypothalami and human placental trophoblasts. Various neurotransmitters, their agonists and antagonists, and various neuropeptides such as neurotensin, substance-P or gastrin-releasing peptide will be studied in vitro and in vivo to examine their roles in the regulation of hypothalamic secretion of CRF, GRF and other hypophysiotrophic hormones. A hypothesis that placental CRF is regulating placental ACTH, and placental GRF and somatostatin are regulating human placental lactogen (HPL) in a paracrine manner will be examined using the perifusion technique and immunocytochemistry. Biosynthesis of CRF and GRF by cultured trophoblasts will be studied using pulse-labeling. In some studies, choriocarcinoma cells and normal trophoblasts will be compared. CRF and GRF of extrahypothalamic sources, including cerebral cortex, spinal cord and placenta, will be characterized by chemical, immunological and biological methods. The possible involvement of CRF, AVP and/or OXT in the nycterohemeral variation of ACTH secretion will be examined in perifused hypothalamus and in vivo using hypothalamic paraventricular lesioning and CRF immunoneutralization. Similarly, the potential role of hypothalamic GRF in determining the sex difference in growth rate and the relative contribution of GRF and sex steroids in this phenomenon will be examined. Localization of enzymes involved in degradation of CRF and GRF will be studied by incubating extracts of various tissues with synthetic CRF or GRF.