Neurons communicate specifically with one another and with muscle cells at chemical synapses by secretion of a neurotransmitter substance. Our knowledge of the mechanisms through which neurons and their synapses develop and function is fundamental to our understanding of the nervous system. The dramatic growth and metamorphosis of lepidopteran insects provides attractive research material for studies of neuromuscular development. This research proposal deals with the various developmental stages of an easily-reared, large moth, the tobacco hornworm (Manduca sexta), and has three immediate objectives: (1) to study the sequence of cellular changes in parts of the nervous and muscular systems during development and metamorphosis; (2) to examine the specification and development of selected neurotransmitter systems and correlate these observations with the cellular events; and (3) to study in culture the regulation of transmitter ontogeny and the mutual influences of nerve and muscle during development. Cellular events will be studied with histological, autoradiographic and electrophysiological methods in order to inquire into the origin, specification, growth and maturation of neurons. Sensitive radiochemical procedures will be employed for studies of transmitter system identification and changes during the moth life cycle; both enzymes and substrates will be measured. Techniques of cell and tissue culture will be used to inquire into the influences of hormones, cell-cell interaction, nutrients and other factors on cellulr and biochemical development of nervous and muscular explants from embryos, larvae and pupae.