Neurons in the auditory system of vertebrates encode the intensity, frequency and location of sounds through the time of their firing. Their ability to encode timing precisely results from adaptations at the cellular and molecular level of individual neurons and their interconnections. Preliminary evidence suggests that glutamate receptors of the AMPA (-amino-3-hydroxy-5-methyl-4-isoxazole-propionate) class, which mediate excitatory transmission at many stages of the auditory pathway, are also specialized for audition. The expression of unique receptor subtypes may be critical to processing auditory input; defects, either direct or indirect, in the control of receptor expression could significantly impair normal function. I propose to examine the biophysical properties of AMPA receptor subtypes in identified neurons in auditory brain stem nuclei of mice. In these same cell types, I will then correlate the kinetic and permeation characteristics of receptors with the behavior of functioning synapses. Recordings will be made from cells in brain slices maintained in vitro. Whole-cell and excised membrane patch recordings will be made of synaptic activity and of responses to agonists of glutamate receptors. Neurons will be identified by dye injection and subsequent morphological reconstruction. In addition, I will apply these techniques to genetically engineered mice with knockouts of the subunits of specific glutamate receptors, to assess the role of each subunit in the function of the native, subsynaptic AMPA receptor in auditory neurons. Finally, in a separate study, we will test the hypothesis that specializations in receptor expression arise from neuronal interactions within a given pathway using a cell culture model of the cochlear nucleus. Neurons from auditory and non-auditory sources will be co-cultured. We will determine the kinetic properties of glutamate receptors expressed under different culture conditions using whole-cell and excised patch techniques. Together, the results from these experiments will provide new information regarding how neurons which perform particular tasks in the brain develop specialized biophysical and molecular properties.
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