instnjctions): The overall aim is to gain a better understanding of the manner in which general anesthetics interact with the activated states of the ligand-gated ion channels of the Cys-loop superfamily. The overall hypothesis is that general anesthetics interact with these channels at allosteric sites whose properties vary with the channel's conformational state. A given binding site's size and affinity for anesthetics varies with time as the receptor changes its conformation following addition of agonist. The approach is structured to encourage a tight integration between kinetic and structural approaches with kinetic studies generating hypotheses about the relative affinity of anesthetics for specific transient conformational states and time-resolved freeze quenched photolabeling being used to detemriine the degree of photoincorporation and to locate anesthetic sites.
Aims 1 and 2 probe the mechanisms and binding sites involved in anesthetic enhancement of agonist- induced ion currents.
Aim 1 addresses this issue for the cationic members of the family where only the smallest anesthetics enhance currents. The specific hypothesis to be tested is that the enhancing site increases in size as the receptor passes from the closed to the open and then desensitized states. The kinetics of this are most easily dissected on the slowest member of the superfamily, the human neuronal 5 HT3AR.
Aim 2 will locate the enhancing site in human neuronal GABAARs in collaboration with Project 3 and using photoactivable anesthetics from the etomidate, propofol, barbiturate and alcohol families. The second part of this aim asks whether the site at which etomidate itself activates GABA currents is the same as that which enhances agonist-induced currents.
Aim 3 explores the role of binding sites at the subunit interface vs. those in the ion channel in inhibition and enhancement using probes specifically designed for the task. This work will use the Torpedo acetylcholine receptor because its known structure will facilitate interpretation. In each aim, the receptor chosen is that best suited to answering the question. Photolabels and purified, heterologously expressed receptors will be provided by the Synthetic Chemistry and Protein Production Cores respectively, and sequencing by the Protein Chemistry Core.

Public Health Relevance

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Program Projects (P01)
Project #
Application #
Study Section
Special Emphasis Panel (ZGM1-PPBC-0)
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Massachusetts General Hospital
United States
Zip Code
Eaton, Megan M; Germann, Allison L; Arora, Ruby et al. (2016) Multiple Non-Equivalent Interfaces Mediate Direct Activation of GABAA Receptors by Propofol. Curr Neuropharmacol 14:772-80
Zhang, Xi (2016) Instant Integrated Ultradeep Quantitative-structural Membrane Proteomics Discovered Post-translational Modification Signatures for Human Cys-loop Receptor Subunit Bias. Mol Cell Proteomics 15:3665-3684
Forman, Stuart A; Miller, Keith W (2016) Mapping General Anesthetic Sites in Heteromeric γ-Aminobutyric Acid Type A Receptors Reveals a Potential For Targeting Receptor Subtypes. Anesth Analg 123:1263-1273
Chiara, David C; Jounaidi, Youssef; Zhou, Xiaojuan et al. (2016) General Anesthetic Binding Sites in Human α4β3δ γ-Aminobutyric Acid Type A Receptors (GABAARs). J Biol Chem 291:26529-26539
Amlong, Corey A; Perkins, Mark G; Houle, Timothy T et al. (2016) Contrasting Effects of the γ-Aminobutyric Acid Type A Receptor β3 Subunit N265M Mutation on Loss of Righting Reflexes Induced by Etomidate and the Novel Anesthetic Barbiturate R-mTFD-MPAB. Anesth Analg 123:1241-1246
Ziemba, Alexis M; Forman, Stuart A (2016) Correction for Inhibition Leads to an Allosteric Co-Agonist Model for Pentobarbital Modulation and Activation of α1β3γ2L GABAA Receptors. PLoS One 11:e0154031
Nourmahnad, Anahita; Stern, Alex T; Hotta, Mayo et al. (2016) Tryptophan and Cysteine Mutations in M1 Helices of α1β3γ2L γ-Aminobutyric Acid Type A Receptors Indicate Distinct Intersubunit Sites for Four Intravenous Anesthetics and One Orphan Site. Anesthesiology 125:1144-1158
Forman, Stuart A; Chiara, David C; Miller, Keith W (2015) Anesthetics target interfacial transmembrane sites in nicotinic acetylcholine receptors. Neuropharmacology 96:169-77
Hamouda, Ayman K; Wang, Ze-Jun; Stewart, Deirdre S et al. (2015) Desformylflustrabromine (dFBr) and [3H]dFBr-Labeled Binding Sites in a Nicotinic Acetylcholine Receptor. Mol Pharmacol 88:1-11
Liu, K; Jounaidi, Y; Forman, S A et al. (2015) Etomidate uniquely modulates the desensitization of recombinant α1β3δ GABA(A) receptors. Neuroscience 300:307-13

Showing the most recent 10 out of 104 publications