The objectives of the research are to isolate and characterize an insect neurohormone gene and investigate regulation of its expression. The neurohormone-gene model for investigation is the hypertrehalosemic hormone (HTH) of the neotropical cockroach Blaberus discoidalis. HTH is a hyperglycemic hormone that stimulates fat body glycogenolysis to produce precursors for the synthesis of hemolymph trehalose, the principal insect blood sugar. HTH is a representative peptide in the adipokinetic hormone/red-pigment concentrating hormone family of insects and crustaceans. This neuropeptide family appears ubiquitous among all insect species examined and is important for regulation of an array of metabolic processes related to energy production/consumption. Blaberus HTH is a useful model for examining the regulation of neurohormone gene expression in insects because of the breadth of knowledge concerning both the specific importance of HTH in Blaberus physiology and this hormone family, in general. Development of biotechnology to disrupt the insect neuroendocrine balance can be applied to interfere with vital physiological processes of insects as a new approach to the management of insect disease-vectors. Interference with specific neurohormone synthesis and secretion is one avenue for disturbing the insect neuroendocrine balance. Objective 1 will isolate mRNA from the corpora cardiaca and generate HTH-cDNA by polymerase chain reaction methods. Objective 2 will make use of the resulting HTH-cDNA to isolate and characterize the HTH structural gene. The sequence of the structural gene will be examined to identify potential cis-acting regulatory sequences upstream of the transcription start site that might provide important clues concerning how the gene is regulated. Objective 3 will use antisense HTH-mRNA as a probe for use in dot/slot blots to quantitate HTH mRNA and determine if hemolymph concentrations of trehalose other carbohydrates exert feedback regulations at the level of HTH gene transcription. The research will examine whether regulations occur through direct effects on HTH-neurosecretory cells or indirectly through aminergic neural intermediates. Results from this research will define the organization of the hypertrehalosemic hormone gene and provide the first evidence in insects concerning how the physiological state of the animal affects neurohormone synthesis-secretion.
