General anesthetics alter brain function via mechanisms that remain unknown. Although much evidence points to the GABAA receptor as a key target, causal relationships between changes in receptor function and behavioral end points have not yet been well established, and molecular mechanisms by which anesthetics alter channel function have not been determined. The goal of the proposed research is to relate molecular actions of anesthetics on GABAA receptors to specific behavioral effects. The strategy is to focus on inhibition in the hippocampus, a brain structure that plays an important role in memory and is particularly susceptible to development of seizure activity. To accomplish this goal, three specific aims are identified: 1) Test the hypothesis that the amnestic and convulsant properties of volatile anesthetics and related agents in vivo result from their actions on separate types of GABAA responses that are observed in vitro; 2) Determine which GABAA receptor channel kinetic parameters are altered by anesthetics, and how these changes cause inhibitory synaptic currents to be prolonged; 3) Determine the structural and kinetic basis of GABAA receptor block by volatile agents. Experiments will be conducted using electrophysiological techniques to record the responses of hippocampal neurons to anesthetics, non-immobilizers, and pro-convulsant agents. Responses of different receptor populations will be compared to test for associations with behaviorally specific actions. Kinetic mechanisms of anesthetic action will be investigated using rapid solution exchange techniques, single channel recordings, and recombinant receptors. Results will be interpreted in terms of standard and newly developed allosteric kinetic models. Structural features associated with channel block will be determined using chimeric receptors created from alpha and gamma subunits, which differ substantially in susceptibility to block. These results will provide new information regarding the cellular and molecular bases of anesthetic action. These drugs are widely used, and understanding the mechanisms by which they produce their beneficial effects and detrimental side-effects will enable the safer use of existing agents and the development of safer new compounds.
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