Development of synapses in the antennal lobes of the moth Manduca sexta during metamorphosis.

During the metamorphosis of holometabolous insects, the larval nervous system is restructured to provide the circuitry needed by the developing adult. Prominent new centers in the brain, the antennal lobes, arise to receive olfactory afferent axons from the developing adult antennae and provide an excellent system in which to study the development of synapses in a central nervous system. We have examined the anatomy and physiology of developing synapses in the antennal lobes of the moth Manduca sexta during the 18 days of metamorphic adult development. On day 5, the neuropil of the newly emerging antennal lobe condenses into distinct glomeruli, in which intercellular junctional complexes have already begun to form. Although some junctions have associated synaptic vesicles, most complexes are desmosome-like until day 9, when the number of synaptic complexes begins to increase. Early synapses are characterized by membrane-associated densities in at least two abutting cellular processes and a small number of synaptic vesicles clustered near the membrane of one process. As adult development proceeds, the membrane-associated densities become denser and more extensive, and the number of synaptic vesicles in the clusters increases. At day 14 synapses appear ultrastructurally mature, and almost all junctions in the neuropil can be identified as synaptic. Not until day 9 do antennal lobe neurons begin to respond postsynaptically when the antennal nerve is stimulated electrically, suggesting that the earliest synapses observed in the electron microscope may not be made by antennal nerve axons. At first the postsynaptic responses are graded and fatigue rapidly. By day 11, the antennal lobe neurons respond with action potentials, but the fatigability does not decline to adult levels until day 13. Filling of antennal lobe neurons with cobalt reveals that the arborizations of both local interneurons and output neurons continue to mature morphologically until about day 13. Previous work (Schweitzer, E. S., J. R. Sanes, and J. G. Hildebrand (1976) J. Insect Physiol. 22: 955-960) showed that antennal sensilla do not begin to be responsive to odors until day 14. Thus the establishme lt of the synaptic network in the antennal lobe apparently occurs in the absence of functional olfactory input to the system. Neurons in vertebrates often depend upon establishment of contact with their targets for their survival and ’ This work was supported by National Science Foundation Grants BNS-77-13281 and BNS-80-13511 and by National Institutes of Health Grant AI-17711. L. P. T. and S. G. M. were supported by National Institutes of Health postdoctoral fellowships and National Institutes of Health Training Grant NS-07112. We thank M. Imperato and R. Montague for excellent technical assistance, H. Harrow and J. Torrisi for secretarial help, A. M. Schneiderman for collaboration on deafferentation experiments, Drs. A. H. Baumhover, J. A. Svoboda, and C. M. Williams for supplying Munduca eggs, and Dr. A. FrGhlich for helpful comments on the manuscript. ’ Present address: Department of Anatomy, Georgetown University School of Medicine, 3900 Reservoir Road, Washington, DC 20007. 3 To whom correspondence should be addressed at Department of Biological Sciences, Fairchild 913, Columbia University, New York, NY 10027. normal maturation (e.g., Cowan and Wenger, 1967; Black et al., 1972; Landmesser and Pilar, 1972) and may strongly influence the development of their target cells (e.g., Guth, 1957; Black et al., 1971). Invertebrate neurons, such as the lamina monopolar cells in the firstorder optic neuropil of several arthropods (e.g., Kop&, 1922; Macagno, 1977; Anderson, 1978; Maxwell and Hildebrand, 1981), likewise may depend upon appropriate cell-cell contacts for their normal development. In the olfactory pathway of the moth Manduca sexta, however, the effects of preor postsynaptic deprivation seem to be less drastic. The sensory neurons in the antennal flagellum follow the normal schedule of morphological, biochemical, and electrophysiological differentiation even when deprived of their target cells in the brain (Sanes et al., 1976). Moreover, the antennal lobe, the primary olfactory center in the brain, when deprived