Gamma-aminobutyric acid type A receptors (GABAARs) mediate the majority of synaptic inhibition in the brain and are modulated by a variety of clinically important drugs, such as benzodiazepines, barbiturates, steroids, anesthetics and anti-convulsants. Furthermore, GABAAR mutations have recently been linked to epilepsy. The long-term goal of our research program is to understand the function of the GABAAR in terms of its molecular structure. While recent crystallographic advances have provided valuable structural models of the GABAAR, achieving a full understanding of function also requires knowledge of protein dynamics. GABAARs exist in at least three interconvertible states with distinct functions: inactive/closed, active/open, and desensitized/closed. Very little is known about the protein motions that occur during these structural transitions, which are regulated by neurotransmitter and drug binding. We will use fluorescence recording of site-specific labels in GABAARs expressed in Xenopus oocytes to study the structural rearrangements underlying activation, desensitization, and drug modulation as they occur in real time. We propose to study 1) agonist, partial-agonist and antagonist induced rearrangements, 2) global protein motions, 3) pentobarbital induced structural changes and 4) protein motions during desensitization. The experiments will be interpreted with the aid of recently elucidated atomic- level structures to gain a deeper understanding of the molecular mechanisms underlying the function of GABAARs and their relatives. We cannot hope to predict the actions of a drug or ligand or predict the outcome of a disease-causing mutation in the GABAAR without dissecting the movements in the protein that mediate its function. The research proposed here utilizes an innovative new approach that will enable us to learn how GABAARs function in health and disease states.
The opening and closing of ligand-gated ion channels, which lie in the membranes of nerve cells, regulate information flow throughout the brain. Defects in these channels lead to wide variety of diseases, such as myasthenia, hyperekplexia and epilepsy. These channels are also the targets of a number of clinically used drugs, including muscle relaxants, sedative-hypnotics, anti-convulsants, anxiolytics and anesthetics. We cannot hope to predict the actions of a drug, design safer and more effective drugs, develop better therapeutic strategies or predict the outcome of a disease-causing mutation without knowledge of how these channels work at a molecular level. The research proposed here utilizes an innovative new approach that will increase our understanding of how one type of ion channel, the GABAAR, functions in health and disease and will establish testable hypotheses for elucidating how other related ligand-gated ion channels function.
|Muroi, Yukiko; Theusch, Cassandra M; Czajkowski, Cynthia et al. (2009) Distinct structural changes in the GABAA receptor elicited by pentobarbital and GABA. Biophys J 96:499-509|