Molecular targets and mechanisms for ethanol action in glycine receptors

Abstract

Glycine receptors (GlyRs) are recognized as the primary mediators of neuronal inhibition in the spinal cord, brain stem and higher brain regions known to be sensitive to ethanol. Building evidence supports the notion that ethanol acting on GlyRs causes at least a subset of its behavioral effects and may be involved in modulating ethanol intake. For over two decades, GlyRs have been studied at the molecular level as targets for ethanol action. Despite the advances in understanding the effects of ethanol in vivo and in vitro, the precise molecular sites and mechanisms of action for ethanol in ligand-gated ion channels in general, and in GlyRs specifically, are just now starting to become understood. The present review focuses on advances in our knowledge produced by using molecular biology, pressure antagonism, electrophysiology and molecular modeling strategies over the last two decades to probe, identify and model the initial molecular sites and mechanisms of ethanol action in GlyRs. The molecular targets on the GlyR are covered on a global perspective, which includes the intracellular, transmembrane and extracellular domains. The latter has received increasing attention in recent years. Recent molecular models of the sites of ethanol action in GlyRs and their implications to our understanding of possible mechanism of ethanol action and novel targets for drug development in GlyRs are discussed.

Fig. 1. PMTS binding to cysteines substituted at position 52 and/or 267 reveals position-specific negative and positive modulation by ethanol in α1GlyRs

Mean ± SEM percent control glycine response for (A.) WT and (B. – D.) mutant α1GlyRs (n = 4 – 9). Shaded boxes indicate GlyR responses in the presence of PMTS (grey) or 100 mM ethanol (yellow). (* p < 0.05 vs. control glycine EC₁₀ response, † p < 0.05 versus residual effect of PMTS). Fig. from Crawford et al. 2007 (Crawford et al., 2007).

Fig. 2. Zoom view of a molecular model of a α1GlyR subunit with the alcohol pocket highlighted

The backbone atoms of one α1GlyR subunit are shown as a yellow ribbon. The largest cavity found by the Binding Site Analysis module of Insight 2005L is shown in red. The Cα atoms of A52 and S267 are separated by approximately 28Å. The interconnected sections of the pocket are large enough to accommodate ethanol and allow its passage between these regions. The ability of the A52C-S267C double mutant to gate currents and bind PMTS is consistent with there being enough distance between positions 52 and 267 to prevent spontaneous disulfide linkage between the cysteine substitutions. Figure modified from Crawford et al. 2007 (Crawford et al., 2007).

Fig. 3

(A) Loop 2 in a single GlyR WT subunit. Residues interacting with Loop 2 are rendered as colored stick models: Lys104, blue; Leu136, yellow; Arg218, pink, Lys276, green; (B) Delta Loop 2 in a single GlyR subunit. Residues interacting with Loop 2 are rendered as colored stick models: Lys104, blue; Leu136, yellow; Arg218, pink, Lys276, green (Figure modified from Perkins et al. 2009) (Perkins et al., 2009).