It is the primary objective of this research to elucidate the mechanisms involved in the control of neurohormone synthesis and release. The complexity of vertebrate neuroendocrine systems has thus far precluded a direct examination of these physiological processes, but since functional and structural homologies exist in the invertebrate neuroendocrine systems fundamental aspects of neuroendocrine physiology can perhaps be better studied in these evolutionarily less complex systems. Of all the invertebrates, insects are the best suited for such research because their neuroendocrine system is fairly simple andis relatively well defined. The insect neurohormone to be used for this study is the prothoracicotropic hormone (PTTH), which is the primary hornworm, Manduca sexta, the PTTH neuroendocrine system consists of a unicellular axis from which two forms of a hormone are differentially released at specific times during neurosecretaroy processes, while its functional complexity will provide information about the modes of release of different forms of a neurohormone as well as about their unique actions on a common target gland. This study proposes: 1) to complete the purification of big and small PTTH; 2) to fully characterize the PTTHs released in situ; 3) to generate monoclonal antibodies to the PTTHs with which to develop a PTTH RIA: 4) to investigate with immunocytochemical techniques the syn thesis and transport of PTTH plus its binding and uptake by its target, the prothoracic glands (PG); 5) to define the nature of the environmental control of PTTH synthesis and release and to demonstrate this control in vitro; and 6) to establish the basic biochemistry of the mechanism(s) involved in the activation of the PG by the two forms of PTTH. Contemporary methods in protein purification, hybridoma technology, RIA, EM immunocytochemistry and autoradiography, in vitro cell culture, enzymology and clock biology etc. will be employed in the accomplishment of these objectives. The information resulting from this proposal should provide use with a basis from which to probe in vitro the mechanisms involved in the regulation of neurosecretory processes in general.
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