The goal of the proposed research is to understand how the many types of ion channels present in a single neuron work together to produce the firing patterns of that neuron. Ultimately, we hope to use this knowledge to develop ion channel-targeted pharmacological agents that can differentially regulate firing of different types of neurons and thereby treat pathophysiological behavior such as epilepsy and ataxias. Previous grant periods focused on the functional roles of calcium channels, sodium channels, Kv2, and Kv4 potassium channels. The focus in the next grant period is to understand how different potassium channels combine to regulate the firing patterns of specific types of neurons and how inhibitors and enhancers of particular potassium channels differentially modify firing patterns of different neurons. We will follow up preliminary data showing that inhibition of BK and Kv3 potassium channels ? including by the clinically-used drug 4-aminopyridine - often causes paradoxical slowing rather than speeding of firing. We will test two possible mechanisms for this effect: enhanced inactivation of Na channels or recruitment of other potassium channels with slower deactivation. A key element is to compare action potential firing in different types of neurons. We will focus on three examples of major electrophysiological phenotypes: hippocampal CA1 pyramidal neurons, cerebellar Purkinje neurons, and midbrain dopamine neurons. The experimental design will combine current clamp recordings of action potential firing with voltage- clamp analysis of the underlying currents, using both intact neurons in brain slice and acutely dissociated neurons, which allow fast voltage clamp to study gating on the rapid time scale of the action potential. A key feature of the approach will be to study both action potential firing and channel gating kinetics at 37 C. Current clamp and voltage clamp experiments will be directly linked by the action potential clamp technique, in which recordings of natural firing behavior are used as voltage clamp commands to allow pharmacological dissection of the components of current generating patterns of action potentials. The research will include substantial characterization of pharmacological agents that block or enhance potassium channel subtypes, both as experimental tools and as possible leads for clinical drugs.
Understanding the basic mechanisms that control the excitability of central neurons will help in understanding the normal function of the nervous system as well as pathophysiology resulting from epilepsy, ataxia, and other disorders. The work will specifically explore molecular targets and the mechanism of action of 4-aminopyridine, a drug used clinically to treat ataxias. Additionally, the work will characterize and evaluate a variety of investigational agents that inhibit or enhance specific ion channels, providing useful tools for neuroscience research and helping lead to new ways of controlling pathophysiology.
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