Quantitative 3-dimensional analysis of functional motor circuit remodelling in holometabolous insects

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The metamorphosis of holometabolous insects, like Manduca sexta and Drosophila melanogaster, has long served as a powerful model to study mechanisms and functions of postembryonic remodelling of neuronal structure. However, this analysis has mostly been restricted to the single-neurone level. The aim of the proposed project is to expand the analysis to the network level, to address mechanisms and functions of dendritic and axonal shape remodeling and synaptic wiring modifications in developing motor circuits with changing behavioural roles. Two parallel approaches will be conducted: First, we will identify descending neurones with their somata in the SEG, which are presynaptic to the motoneurones for which we already have quantitative 3-D databases. SEG neurone populations are good candidates for descending motor control. Identification of such projection neurones will be achieved by a combination of classical tracing techniques and injection or genetic expression of transsynaptic dyes (e.g. Wheat Germ Agglutinin). Both pre- and postsynaptic circuit elements will be implemented into a three-dimensional reference atlas of insect motor circuits. The necessary tools in AMIRA have recently been developed in our group. Second, the localization of various transmitter substances in the terminals of descending neurones will be analyzed quantitatively in relation to the distribution of receptors throughout the dendritic trees of thoracic motoneurones. This approach will expand our analysis of synapse distribution in dendritic trees from the individual identified neurone to the network level and will contribute to the understanding of the development and the function of motor control by higher centres. Third, the internal organization of selected motor neuropiles with respect to their transmitter and receptor equipment and density will be assessed by immuno-labelling of synaptic proteins and receptors on the light microscopy level. The combination of our 3-D atlas of individual network partners with the study of neuropile organization is an important step toward understanding network organization and function. The digital neuroanatomical data will be combined with physiological data to create realistic multi-compartment models of synaptic integration in dendritic trees, and in the long run, of the computational power of dendritic tree networks.