Basic and Translational Research on General Anesthetics

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Project Summary. General anesthesia, the reversible pharmacologic inhibition of neural functions underlying consciousness, awareness, memory, and nociceptive motor responses, is an essential medical intervention. There is a clear need for new anesthetic drugs with predictable kinetics and reduced toxicity, which will facilitate medical procedural innovation, efficacy, efficiency, and accessibility. I have contributed to these goals using two rigorous and complementary strategies. First, I have rigorously advanced basic science knowledge of molecular anesthetic mechanisms in established targets such as GABAA receptors. This research has revealed unexpectedly specific interactions between different general anesthetic chemotypes (etomidate derivatives, methyl-phenyl allyl barbiturates or MPABs, neurosteroids like alphaxalone, and benzoyl alcohols) and distinct sets of transmembrane inter-subunit sites in typical synaptic GABAA receptors. I also introduced Monod-Wyman-Changeux (MWC) two-state co-agonist models for quantitative analysis of anesthetic effects in these receptors. Second, recognizing that GABAA receptor-specific drugs have proven unsatisfactory as sole clinical general anesthetic agents, I am among the first to adopt zebrafish as an unbiased pharmacodynamic model to discover new hypnotic chemotypes that may act via multiple molecular targets, and to develop transgenic lines to accelerate mechanisms research. The broad long-term objectives of this R35 (MIRA) grant during the next five years and beyond are to further advance our understanding of anesthetic mechanisms at the molecular level, to discover new chemical families with sedative-hypnotic activity, and to create new transgenic zebrafish to test the roles of specific drug-receptor interactions in anesthetic effects. Molecular knowledge areas that will be addressed by this project include, but are not limited to improving the precision of anesthetic site mapping in GABAA receptors, developing MWC models that account for differential agonist versus GABA modulation effects of the distinct site-selective anesthetic chemotypes, probing the subunit arrangement and structures of other important (e.g. extra-synaptic) GABAA receptor isotypes using subsite-specific anesthetics and mutations that selectively affect their actions, and extending these structure-function approaches to other anesthetic-sensitive pentameric ligand-gated ion channels. I will also apply the platform combining zebrafish larvae and real-time video analysis of up to 96 animals at a time to identify new sedative-hypnotic chemotypes in drug libraries and assess a variety of sedative-hypnotic drug interactions. New hypnotic chemotypes will be characterized for effects in a panel of molecular anesthetic targets using electrophysiology and pharmacologic tools. I will also develop new transgenic zebrafish lines with knock-out or knock-in mutations in anesthetic target proteins as models for testing if these targets mediate the behavioral effects of established or discovered sedative-hypnotics.