Reversal of Ethanol-induced Intoxication by a Novel Modulator of Gβγ Protein Potentiation of the Glycine Receptor

Loreto San Martin,Fabian Cerda,Chunyang Jin,Veronica Jimenez,Gonzalo E Yevenes,Tania Hernandez,Daniela Nova,Jorge Fuentealba,Luis G Aguayo,Leonardo Guzman

doi:10.1074/jbc.m116.740555

Abstract

The acute intoxicating effects of ethanol in the central nervous system result from the modulation of several molecular targets. It is widely accepted that ethanol enhances the activity of the glycine receptor (GlyR), thus enhancing inhibitory neurotransmission, leading to motor effects, sedation, and respiratory depression. We previously reported that small peptides interfered with the binding of Gβγ to the GlyR and consequently inhibited the ethanol-induced potentiation of the receptor. Now, using virtual screening, we identified a subset of small molecules capable of interacting with the binding site of Gβγ. One of these compounds, M554, inhibited the ethanol potentiation of the GlyR in both evoked currents and synaptic transmission in vitro When this compound was tested in vivo in mice treated with ethanol (1-3.5 g/kg), it was found to induce a faster recovery of motor incoordination in rotarod experiments and a shorter sedative effect in loss of righting reflex assays. This study describes a novel molecule that might be relevant for the design of useful therapeutic compounds in the treatment of acute alcohol intoxication.

Highlights

Studied as a molecular target for ethanol, and electrophysiological experiments have demonstrated that ethanol potentiates glycine receptor (GlyR) activity [3], as determined by increased glycine-evoked currents [4, 5], an increased decay time constant in spontaneous synaptic events [6], increased probability of channel opening in single-channel analysis [7], and increased agonist affinity [8]
Our virtual screening produced a subset of small molecule leads that were examined as inhibitors for the ethanol effects on the GlyR using electrophysiological studies as a first approach to identify active compounds
Co-administration of M119 with morphine resulted in a leftward shift in the morphine antinociceptive dose-response curve [13], similar to the shift observed with phospholipase C␤ (PLC␤)3 knockout mice [18]

Summary

Introduction

Studied as a molecular target for ethanol, and electrophysiological experiments have demonstrated that ethanol potentiates GlyR activity [3], as determined by increased glycine-evoked currents [4, 5], an increased decay time constant in spontaneous synaptic events [6], increased probability of channel opening in single-channel analysis [7], and increased agonist affinity [8]. A small peptide (RQHc7) that binds G␤␥ inhibited the potentiation effect of ethanol on both evoked and synaptic currents [6, 11]. In this study, we aimed to identify small molecules capable of binding G␤␥ and able to inhibit the potentiation of the glycine current induced by ethanol. One of these compounds was assayed in in vivo pharmacology in mice using increasing concentrations of ethanol, demonstrating that it was able to partly reverse some intoxicating effects. The rationale for designing pharmacological modulators that interfere with the G␤␥-GlyR proteinprotein interaction is amenable for blocking acute ethanol intoxication

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Conclusion