Antagonism of Inhalant and Volatile Anesthetic Enhancement of Glycine Receptor Function

Michael J Beckstead,Rachel Phelan,S John Mihic

doi:10.1074/jbc.m011627200

Michael J Beckstead, Rachel Phelan + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m011627200

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Abstract

Recent studies suggest that alcohols, volatile anesthetics, and inhaled drugs of abuse, which enhance gamma-aminobutyric acid, type A, and glycine receptor-activated ion channel function, may share common or overlapping molecular sites of action on these receptors. To investigate this possibility, these compounds were applied singly and in combination to wild-type glycine alpha(1) receptors expressed in Xenopus laevis oocytes. Data obtained from concentration-response curves of the volatile anesthetic enflurane constructed in the presence and absence of ethanol, chloroform, or toluene were consistent with competition for a common binding pocket on these receptors. A mutant glycine receptor, insensitive to the enhancing effects of ethanol but not anesthetics or inhalants, demonstrated antagonism of anesthetic and inhalant effects on this receptor. Although ethanol (25-200 mm) had no effect on its own in this receptor, it was able to inhibit reversibly the enhancing effect of enflurane, toluene, and chloroform in a concentration-dependent manner. These data suggest the existence of overlapping molecular sites of action for ethanol, inhalants, and volatile anesthetics on glycine receptors and illustrate the feasibility of pharmacological antagonism of the effects of volatile anesthetics.

Highlights

Recent studies suggest that alcohols, volatile anesthetics, and inhaled drugs of abuse, which enhance ␥-aminobutyric acid, type A, and glycine receptor-activated ion channel function, may share common or overlapping molecular sites of action on these receptors
Specific single amino acid mutations alter the actions of allosteric modulators on glycine receptors (Gly-Rs) [6, 7], and chemically diverse compounds, such as n-alcohols, anesthetics, and inhalants, may even share a common binding site on these ligandgated ion channels [3, 7,8]
To determine if CHCl3 could be acting at two distinct sites on glycine receptors, with only one site shared with EtOH, TOL, TCE, and ENF, we replaced an amino acid believed to compose part of the putative alcohol and anesthetic-binding site with the large phenylalanine residue (S267F)

Summary

The abbreviations used are

Gly-R, glycine receptor; GABAA, ␥-aminobutyric acid, type A; EtOH, ethanol; ENF, enflurane; CHCl3, chloroform; TOL, toluene; TCE, 1,1,1-trichloroethane; ECX, the concentration of agonist (glycine) that produces X% of a maximal current. Specific single amino acid mutations alter the actions of allosteric modulators on Gly-Rs [6, 7], and chemically diverse compounds, such as n-alcohols, anesthetics, and inhalants, may even share a common binding site on these ligandgated ion channels [3, 7,8]. We demonstrate a reversible binding site interaction through the simultaneous administration of two positive allosteric modulators on wild-type and mutant ␣1 Gly-Rs. we tested the hypothesis that ethanol (EtOH), the volatile anesthetics enflurane (ENF) and chloroform (CHCl3), and the inhaled drugs of abuse toluene (TOL) and 1,1,1-trichloroethane (TCE) share common or overlapping binding sites on these receptors. Some of this work has been presented previously in abstract form [10]

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION