Ethanol consumption and sedation are altered in mice lacking the glycine receptor α2 subunit

Abstract

Background and Purpose The precise mechanism/s of action of ethanol, although studied for many years, are not well understood. Like other drugs of abuse, ethanol affects dopamine levels in the nucleus accumbens (nAc), an important region of the mesolimbic system, causing a reinforcing effect. It has been shown that glycine receptors (GlyRs) present in the nAc are potentiated by clinically relevant concentrations of ethanol, where α1 and α2 are the predominant subunits expressed. Experimental Approach Using a combination of electrophysiology and behavioural assays, we studied the involvement of GlyR α2 subunits on the effects of low and high doses of ethanol, as well as on consumption using mice lacking the GlyR α2 subunit (male Glra2−/Y and female Glra2−/−). Key Results GlyR α2 subunits exist in accumbal neurons, since the glycine‐evoked currents and glycinergic miniature inhibitory postsynaptic currents (mIPSCs) in Glra2−/Y mice were drastically decreased. In behavioural studies, differences in ethanol consumption and sedation were observed between wild‐type (WT) and Glra2 knockout (KO) mice. Using the drinking in the dark (DID) paradigm, we found that Glra2−/Y mice presented a binge‐like drinking behaviour immediately when exposed to ethanol rather than the gradual consumption seen in WT animals. Interestingly, the effect of knocking out Glra2 in female (Glra2−/−) mice was less evident, since WT female mice already showed higher DID. Conclusion and Implications The differences in ethanol consumption between WT and KO mice provide additional evidence supporting the conclusion that GlyRs are biologically relevant targets for the sedative and rewarding properties of ethanol.

Contribution of GlyR α3 Subunits to the Sensitivity and Effect of Ethanol in the Nucleus Accumbens

“…While Western blot analysis showed no significant changes in global α subunit levels in the nAc in Glra3 −/− mice, the results showed a significant decrease in GlyR β subunits in the knockout mice. These results indicate that deletion of Glra3 only slightly reduced the presence of the α subunit pool in the nAc, thus suggesting the presence of a lower level of α3 as compared to α1 and α2 (Forstera et al, 2017;San Martin et al, 2020;Gallegos et al, 2021). However, the data does not allow us to rule out the possibility that compensatory up-regulation of α subunits occurred in the Glra3 −/− mice.…”

“…This unchanged duration of LORR in Glra3 −/− mice is consistent with previous published findings (Blednov et al, 2015) and might support a more restricted localization of α3 subunits in the CNS unable to produce a sedative action. On the other hand, previous studies have demonstrated that α1 and α2 subunits are important for the sedative effects of ethanol (Aguayo et al, 2014;San Martin et al, 2020;Gallegos et al, 2021). Thus, these findings suggest that α1 and α2 subunits are more relevant than α3 subunits in terms of alcohol effects on GlyRs in the CNS.…”

“…However, functional data show that PTX inhibited the current to a smaller extent in Glra3 −/− mouse. When compared to the Glra2 −/− mice (San Martin et al, 2020), the Glra3 −/− mouse showed a much higher glycine current density in dissociated accumbal neurons. Thus, the α3 subunit appears to control brain development in a divergent manner to α2 (Avila et al, 2013).…”

“…However, previous studies using recombinant α3 subunits in HEK293 cells suggested that homomeric α3 GlyRs were insensitive to pharmacologically relevant concentrations of ethanol (Sanchez et al, 2015). On the other hand, the presence of GlyR β subunits was shown in the nAc using immunodetection experiments (Forstera et al, 2017;Munoz et al, 2018;San Martin et al, 2020), thus indicating the presence of heteromeric α3β GlyRs in this region. Therefore, it is possible that ethanol potentiates α3β, but not α3 homomeric complexes, a subunit arrangement that is likely present in the nAc, but this possibility has not yet been examined.…”

“…The smaller effect of PTX found in the Glra3 −/− mice suggests an increase in αβ complexes in the membrane supporting a compensatory effect by different GlyR subtypes. Ethanol-Induced Sedation Is Not Altered in Male Glra3 −/− Mice The genetically modified Glra3 −/− mice did not display visible abnormalities or foot clasping behavior when the mouse was lifted by the tail (Supplementary Figure 1A) indicating the absence of alterations in muscle tone and coordination as seen in Glra1 mutant mice (Schaefer et al, 2018) and Glra2 −/− mice (San Martin et al, 2020). Additionally, this knockout mouse did not exhibit gross alterations in muscle strength when assayed by a qualitative grip test (Supplementary Figure 1B).…”