Competitive Antagonism of Anesthetic Action at the &ggr;-Aminobutyric Acid Type A Receptor by a Novel Etomidate Analog with Low Intrinsic Efficacy

Background: The authors characterized the &ggr;-aminobutyric acid type A receptor pharmacology of the novel etomidate analog naphthalene–etomidate, a potential lead compound for the development of anesthetic-selective competitive antagonists. Methods: The positive modulatory potencies and efficacies of etomidate and naphthalene–etomidate were defined in oocyte-expressed &agr;1&bgr;3&ggr;2L &ggr;-aminobutyric acid type A receptors using voltage clamp electrophysiology. Using the same technique, the ability of naphthalene–etomidate to reduce currents evoked by &ggr;-aminobutyric acid alone or &ggr;-aminobutyric acid potentiated by etomidate, propofol, pentobarbital, and diazepam was quantified. The binding affinity of naphthalene–etomidate to the transmembrane anesthetic binding sites of the &ggr;-aminobutyric acid type A receptor was determined from its ability to inhibit receptor photoaffinity labeling by the site-selective photolabels [3H]azi-etomidate and R-[3H]5-allyl-1-methyl-5-(m-trifluoromethyl-diazirynylphenyl) barbituric acid. Results: In contrast to etomidate, naphthalene–etomidate only weakly potentiated &ggr;-aminobutyric acid–evoked currents and induced little direct activation even at a near-saturating aqueous concentration. It inhibited labeling of &ggr;-aminobutyric acid type A receptors by [3H]azi-etomidate and R-[3H]5-allyl-1-methyl-5-(m-trifluoromethyl-diazirynylphenyl) barbituric acid with similar half-maximal inhibitory concentrations of 48 &mgr;M (95% CI, 28 to 81 &mgr;M) and 33 &mgr;M (95% CI, 20 to 54 &mgr;M). It also reduced the positive modulatory actions of anesthetics (propofol > etomidate ~ pentobarbital) but not those of &ggr;-aminobutyric acid or diazepam. At 300 &mgr;M, naphthalene–etomidate increased the half-maximal potentiating propofol concentration from 6.0 &mgr;M (95% CI, 4.4 to 8.0 &mgr;M) to 36 &mgr;M (95% CI, 17 to 78 &mgr;M) without affecting the maximal response obtained at high propofol concentrations. Conclusions: Naphthalene–etomidate is a very low-efficacy etomidate analog that exhibits the pharmacology of an anesthetic competitive antagonist at the &ggr;-aminobutyric acid type A receptor.

[1]  S. Brull,et al.  Current Status of Neuromuscular Reversal and Monitoring: Challenges and Opportunities , 2017, Anesthesiology.

[2]  J. Ricco,et al.  Protamine Reduces Bleeding Complications without Increasing the Risk of Stroke after Carotid Endarterectomy: A Meta-analysis. , 2016, European journal of vascular and endovascular surgery : the official journal of the European Society for Vascular Surgery.

[3]  S. Forman,et al.  Correction for Inhibition Leads to an Allosteric Co-Agonist Model for Pentobarbital Modulation and Activation of α1β3γ2L GABAA Receptors , 2016, PloS one.

[4]  D. Raines,et al.  &ggr;-Aminobutyric Acid Type A Receptor Modulation by Etomidate Analogs , 2016, Anesthesiology.

[5]  M. Sivilotti.  Flumazenil, naloxone and the 'coma cocktail'. , 2016, British journal of clinical pharmacology.

[6]  R. Mishra,et al.  Reversal agents in anaesthesia and critical care , 2015, Indian journal of anaesthesia.

[7]  D. C. Chiara,et al.  Positive and Negative Allosteric Modulation of an α1β3γ2 γ-Aminobutyric Acid Type A (GABAA) Receptor by Binding to a Site in the Transmembrane Domain at the γ+-β− Interface* , 2015, The Journal of Biological Chemistry.

[8]  R. Pearce,et al.  Etomidate blocks LTP and impairs learning but does not enhance tonic inhibition in mice carrying the N265M point mutation in the beta3 subunit of the GABAA receptor , 2015, Neuropharmacology.

[9]  J. Krystal,et al.  Role of GABA Deficit in Sensitivity to the Psychotomimetic Effects of Amphetamine , 2015, Neuropsychopharmacology.

[10]  S. Forman,et al.  Mutations at Beta N265 in γ-Aminobutyric Acid Type A Receptors Alter Both Binding Affinity and Efficacy of Potent Anesthetics , 2014, PloS one.

[11]  D. Raines,et al.  Analogues of Etomidate: Modifications around Etomidate’s Chiral Carbon and the Impact on In Vitro and In Vivo Pharmacology , 2014, Anesthesiology.

[12]  M. Perouansky.  Coagulation, Flocculation, and Denaturation: A Century of Research into Protoplasmic Theories of Anesthesia , 2014, Anesthesia and analgesia.

[13]  Joseph F. Cotten,et al.  The Pharmacology of Cyclopropyl-Methoxycarbonyl Metomidate: A Comparison with Propofol , 2014, Anesthesia and analgesia.

[14]  J. Changeux.  50 years of allosteric interactions: the twists and turns of the models , 2013, Nature Reviews Molecular Cell Biology.

[15]  D. C. Chiara,et al.  Specificity of Intersubunit General Anesthetic-binding Sites in the Transmembrane Domain of the Human α1β3γ2 γ-Aminobutyric Acid Type A (GABAA) Receptor*♦ , 2013, The Journal of Biological Chemistry.

[16]  Joseph F. Cotten,et al.  In Vivo and In Vitro Pharmacological Studies of Methoxycarbonyl-Carboetomidate , 2012, Anesthesia and analgesia.

[17]  D. Raines,et al.  Pharmacological Studies of Methoxycarbonyl Etomidate's Carboxylic Acid Metabolite , 2012, Anesthesia and analgesia.

[18]  S. Forman.  Monod–Wyman–Changeux allosteric mechanisms of action and the pharmacology of etomidate , 2012, Current opinion in anaesthesiology.

[19]  D. Raines,et al.  Allyl m-trifluoromethyldiazirine mephobarbital: an unusually potent enantioselective and photoreactive barbiturate general anesthetic. , 2012, Journal of medicinal chemistry.

[20]  S. Forman,et al.  Two Etomidate Sites in &agr;1&bgr;2&ggr;2 &ggr;-Aminobutyric Acid Type A Receptors Contribute Equally and Noncooperatively to Modulation of Channel Gating , 2012, Anesthesiology.

[21]  Emery N Brown,et al.  Active Emergence from Propofol General Anesthesia Is Induced by Methylphenidate , 2012, Anesthesiology.

[22]  D. C. Chiara,et al.  Mapping general anesthetic binding site(s) in human α1β3 γ-aminobutyric acid type A receptors with [³H]TDBzl-etomidate, a photoreactive etomidate analogue. , 2012, Biochemistry.

[23]  S. Forman,et al.  An Allosteric Coagonist Model for Propofol Effects on &agr;1&bgr;2&ggr;2L &ggr;-Aminobutyric Acid Type A Receptors , 2012, Anesthesiology.

[24]  K. Miller,et al.  High‐level expression and purification of Cys‐loop ligand‐gated ion channels in a tetracycline‐inducible stable mammalian cell line: GABAA and serotonin receptors , 2010, Protein science : a publication of the Protein Society.

[25]  J. Hunter,et al.  Reversal of neuromuscular block. , 2009, British journal of anaesthesia.

[26]  Ken Solt,et al.  Correlating the clinical actions and molecular mechanisms of general anesthetics , 2007, Current opinion in anaesthesiology.

[27]  R. Olsen,et al.  Identification of a GABAA Receptor Anesthetic Binding Site at Subunit Interfaces by Photolabeling with an Etomidate Analog , 2006, The Journal of Neuroscience.

[28]  N. Franks,et al.  Molecular targets underlying general anaesthesia , 2006, British journal of pharmacology.

[29]  J. Lambert,et al.  Extrasynaptic GABAA Receptors of Thalamocortical Neurons: A Molecular Target for Hypnotics , 2005, The Journal of Neuroscience.

[30]  S. Forman,et al.  Classic Benzodiazepines Modulate the Open–Close Equilibrium in &agr;1&bgr;2&ggr;2L &ggr;-Aminobutyric Acid Type A Receptors , 2005 .

[31]  B. Antkowiak,et al.  Molecular and neuronal substrates for general anaesthetics , 2004, Nature Reviews Neuroscience.

[32]  G. Plourde,et al.  Antagonism of sevoflurane anaesthesia by physostigmine: effects on the auditory steady-state response and bispectral index. , 2003, British journal of anaesthesia.

[33]  K. Sillar,et al.  The in vitro and in vivo enantioselectivity of etomidate implicates the GABAA receptor in general anaesthesia , 2003, Neuropharmacology.

[34]  R. Olsen,et al.  2-(3-Methyl-3H-diaziren-3-yl)ethyl 1-(1-phenylethyl)-1H-imidazole-5-carboxylate: a derivative of the stereoselective general anesthetic etomidate for photolabeling ligand-gated ion channels. , 2003, Journal of Medicinal Chemistry.

[35]  B. Antkowiak,et al.  General anesthetic actions in vivo strongly attenuated by a point mutation in the GABAA receptor β3 subunit , 2003, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[36]  J. Amin,et al.  Benzodiazepines act on GABAA receptors via two distinct and separable mechanisms , 2000, Nature Neuroscience.

[37]  W. R. Lieb,et al.  Which molecular targets are most relevant to general anaesthesia? , 1998, Toxicology letters.

[38]  N L Harrison,et al.  Propofol and other intravenous anesthetics have sites of action on the gamma-aminobutyric acid type A receptor distinct from that for isoflurane. , 1998, Molecular pharmacology.

[39]  E. Sigel,et al.  The benzodiazepine binding site of GABAA receptors. , 1997, Trends in pharmacological sciences.

[40]  E. Sigel,et al.  ReviewThe benzodiazepine binding site of GABAA receptors , 1997 .

[41]  D. Cafiso,et al.  Contrasting membrane localization and behavior of halogenated cyclobutanes that follow or violate the Meyer-Overton hypothesis of general anesthetic potency. , 1997, Biophysical journal.

[42]  J. A. Peters,et al.  Subunit‐dependent interaction of the general anaesthetic etomidate with the γ‐aminobutyric acid type A receptor , 1997, British journal of pharmacology.

[43]  R. Olsen,et al.  Functional domains of GABAA receptors. , 1995, Trends in pharmacological sciences.

[44]  I. Ueda,et al.  Is membrane expansion relevant to anesthesia? Mean excess volume. , 1983, Anesthesiology.

[45]  J. Richards,et al.  Agonist and antagonist benzodiazepine receptor interaction in vitro , 1981, Nature.

[46]  R. Kaufman.  Biophysical Mechanisms of Anesthetic Action: Historical Perspective and Review of Current Concepts , 1977, Anesthesiology.

[47]  K. Miller,et al.  General anaesthetics can selectively perturb lipid bilayer membranes , 1976, Nature.

[48]  S. Snyder,et al.  Properties of opiate-receptor binding in rat brain. , 1973, Proceedings of the National Academy of Sciences of the United States of America.

[49]  E. J. Simon,et al.  Stereospecific binding of the potent narcotic analgesic (3H) Etorphine to rat-brain homogenate. , 1973, Proceedings of the National Academy of Sciences of the United States of America.

[50]  K. Miller,et al.  Site of Action of General Anaesthetics , 1965, Nature.

[51]  A. S. Gale.  The effect of methylphenidate (ritalin) on thiopental recovery. , 1958, Anesthesiology.

[52]  T. E. Waine,et al.  Thiopentone anæsthesia terminated by bemegride , 1958 .

[53]  H. Grant-Whyte.  Thiopentone , 1956 .

[54]  Loren J. Martin,et al.  Cellular / Molecular 5 GABAA Receptors Mediate the Amnestic But Not Sedative-Hypnotic Effects of the General Anesthetic Etomidate , 2006 .

[55]  S. Forman,et al.  Classic benzodiazepines modulate the open-close equilibrium in alpha1beta2gamma2L gamma-aminobutyric acid type A receptors. , 2005, Anesthesiology.

[56]  S. Forman,et al.  Gating allosterism at a single class of etomidate sites on alpha1beta2gamma2L GABA A receptors accounts for both direct activation and agonist modulation. , 2004, The Journal of biological chemistry.

[57]  D. S. Weiss,et al.  Allosteric activation mechanism of the alpha 1 beta 2 gamma 2 gamma-aminobutyric acid type A receptor revealed by mutation of the conserved M2 leucine. , 1999, Biophysical journal.

[58]  D. Raines,et al.  The enhancement of proton/hydroxyl flow across lipid vesicles by inhalation anesthetics. , 1989, Anesthesiology.