The development of neuronal circuitry is a complex process constituted by genetic expressions utilizing temporal specified domains to achieve a diverse neural organization. Genetic development produces multiple climbing fibers on each Purkinje cell when only one is optimal. The highest level of refinement, however, can only be achieved through a combination of molecular organization and functional sculpting of connections to optimize the olivo-cerebellar circuitry. The hypothesis is that optimization of the one to one climbing fiber-Purkinje cell connection is based on a functionally driven elimination and synapse stabilization. The first specific aim is to determine how the architecture and fine structural relationships of olivary axons (climbing fibers) and Purkinje cell dendrites change as evidence of the processes of climbing fiber selection and elimination. In the second aim, we will analyze how calcium and calcium channels are related to the elimination and selection process.
The third aim i s to determine how the cerebellar nuclei and the inferior olive are involved in the optimization process. The methods will include in vivo and in vitro rodent preparations using immuno-labeling for light and electron microscopic mapping of structural processes of stabilization and de-stabilization between axons and targets, two photon microscopy for molecular and functional expression of calcium related elimination and anterograde labeling for following changes in axon-target relationships in the cerebellar cortex, cerebellar nuclei and inferior olive.
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