The transient potassium current, IA, is an important current in determining the firing properties of many types of neurons. In Drosophila and other species, multiple genes code for IA channels, and some of them undergo alternative splicing; this generates multiple mRNA species, each producing an IA channel with different kinetic and pharmacological properties. The physiological role of this genetic complexity is completely unknown. The lobster stomatogastric ganglion provides a unique system in which to study this question. Recent work has shown that IA plays a critical role in determining several aspects of the motor pattern generated by the well- characterized 14-neuron pyloric circuit. IA has different kinetic properties and amplitudes in each of the cell classes, and this heterogeneity is essential in the generation of the motor pattern. This project will study the molecular basis for this heterogeneity. Our hypothesis is that, as in Drosophila, multiple genes code for IA channels in the lobster stomatogastric ganglion, and some of these genes may undergo alternative splicing. Different cells express the IA-related genes and/or alternative splicing differently; this results in different combinations of IA-related mRNA species in each cell, and thus heterogeneity of IA currents in the different cells. Heterogeneity of mRNA thus may play a critical physiological role in determining the different intrinsic electrophysiological properties of different neurons, and helps to shape the pyloric motor rhythm. To test this hypothesis, four specific aims are proposed: 1) Complete a study of IA currents by voltage clamp measurements in all the pyloric neurons. 2) Clone cDNAs for as many different IA-related mRNAs as possible from the lobster nervous system. 3) Using quantitative PCR methods, determine the amounts of each mRNA species in each of the neuron types in the pyloric network. 4) Inject the lobster-specific mRNAs into Xenopus oocytes, both singly and in mixtures approximating those seen in the pyloric neurons, and compare the currents generated by the mRNAs to those measured in the pyloric neurons in Aim 1. This work should allow us to answer WHY there such complexity in expression of IA-related genes. By providing a physiological role for differential gene expression and/or alternative splicing of IA-related genes, our study will also lead towards a better understanding of the roles of different ion channels in the generation of behavior.
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