This research project is directed at understanding how interactions between functionally-linked nerve cells influence development of the specialized neuronal forms and functions found in the brainstem auditory system of mammals and birds. The first group of experiments will investigate how the forms of the highly-specialized axon terminals and synapses found in the cochlear nucleus are influenced by interaction of developing axons with their target neurons. Previous findings from this project have suggested that some neurons (e.g., in the cochlear ganglion) have more potential than others (e.g., in the cochlear nucleus) to form a diversity of axon terminal types under the control of different target neurons. This hypothesis will be evaluated experimentally in chick embryos by pairing abnormal synaptic partners through in vivo transplantations and through in vitro co-cultures of brainstem neurons with various ganglion neurons; the structures of the resulting axon terminals and synapses would be studied by light and electron microscopy. During development, the neurotransmitter amino acids glutamate and gamma- aminobutyric acid (GABA) can produce large changes in cytosolic free calcium concentrations ([Ca2+]i) in neurons; these changes in turn can affect a variety of developmental processes, including growth of axons and dendrites, synapse formation, and cell death. Glutamate is the principal excitatory neurotransmitter in the brainstem auditory system of birds and mammals and GABAergic endings are also prevalent there. In the developing brain stem auditory nuclei of the chick, changes in glutamate and GABA receptors have recently been found to occur in parallel with morphological transformations that give these neurons characteristic shapes related to their functions in hearing; this has suggested that amino acid receptor-induced changes in [Ca2+]i could underlie these events. A second group of proposed experiments is therefore directed at understanding how 1) normal maturation of receptor structure and function affects changes in [Ca2+]i evoked by glutamate and GABA and 2) how normal maturation of these receptors is affected by altered levels of stimulation, including those that occur after early deafening. These studies would make use of immunohistochemical and in situ hybridization histochemical localization of receptor proteins and messenger RNAs, cobalt sulfide histochemical localization of agonist- induced calcium accumulation, quantitative fluorescence microscopy of evoked changes in [Ca2+]i, and whole-cell patch-clamp electrophysiological analysis of the permeability of various receptor- associated channels to Ca2+. These studies will 1) contribute to further development of an animal model of congenital hearing loss that has been widely used for studies of development and regeneration in the auditory system and 2) explore the ability of various sensory ganglion neurons to successfully innervate central auditory neurons.
