The overall goal of this research is to define how neurochemical information is transmitted by the structure of neuropeptides. Our model is the recently identified hypertrehalosemic hormone (HTH, pGlu-Val-Asn-Phe-Ser-Pro-Gly-Trp-Gly-Thr-NH2). This hormone controls increases in hemolymph sugar concentration and heart beat rate. HTH was isolated from nerve tissue of B. discoidalis, cockroaches, sequenced and synthesized in our laboratory. The specific objectives of the proposed research are to: 1) define the half-life and degradation of HTH, 2) define the properties important for HTH side chain and target tissue receptor interaction and 3) elucidate the conformation of HTH at the activated receptor site. We will approach the 1st objective by monitoring the breakdown and elimination of radiolabeled HTH in vivo and in vitro using a HPLC assay. The HPLC separation will be standardized with synthetic analogs that are potential HTH metabolites. We will use scintillation spectrometry to detect and determine the quantity of HTH metabolites in the appropriate HPLC fractions. The interaction of HTH side chains with tissue receptors and the active HTH conformation will be studied through the use of synthethic sequence substitution and restricted-conformational analogs. We will determine the relative activities of these analogs in several bioassays (e.g. in vivo and in vitro hyperglycemic and semi-isolated heart assays). These studies will provide important information about how intercellular information is transferred by the structure of neurohormones. The stabilized agonists and antagonists that results from these studies will provide powerful tools for defined physiological studies and the identification of different receptor types for the invertebrate peptide family related to HTH. The results of this research will expand a simple model for the study of basic neuroendocrine mechanisms not possible in the more complicated vertebrate nervous and circulatory systems. The use of this model can offer potential insights into the design of pharmaceuticals based on vertebrate peptides with general structural and receptor interactions similar to HTH. Further, this model system could lead to future ways by which insect neuroendocrine systems may be manipulated for pest control and reduce the spread of vector borne disease.
