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

Abstract

The glycine receptor (GlyR), a ligand-gated ion channel, is critical for inhibitory neurotransmission in brainstem, spinal cord, and in supraspinal regions. Recent data from several laboratories have shown that GlyRs are expressed in the brain reward circuitry and that α1 and α2 are the principal subunits expressed in the nucleus accumbens (nAc). In the present study, we studied the sensitivity to ethanol of homomeric and heteromeric α3 GlyR subunits in HEK293 cells and dissociated neurons from the nAc. Finally, we explored ethanol-related behaviors in a Glra3 knockout mouse (Glra3–/–). Studies in HEK293 cells showed that while homomeric α3 GlyR subunits were insensitive to ethanol, heteromeric α3β GlyR subunits showed higher sensitivity to ethanol. Additionally, using electrophysiological recordings in dissociated accumbal neurons, we found that the glycine current density increased in Glra3–/– mice and the GlyRs were less affected by ethanol and picrotoxin. We also examined the effect of ethanol on sedation and drinking behavior in Glra3–/– mice and found that the duration in the loss of righting reflex (LORR) was unchanged compared to wild-type (WT) mice. On the other hand, using the drinking in the dark (DID) paradigm, we found that Glra3–/– mice have a larger ethanol consumption compared to WT mice, and that this was already high during the first days of exposure to ethanol. Our results support the conclusion that heteromeric α3β, but not homomeric α3, GlyRs are potentiated by ethanol. Also, the increase in GlyR and GABAAR mediated current densities in accumbal neurons in the KO mice support the presence of compensatory changes to α3 knock out. The increase in ethanol drinking in the Glra3–/– mice might be associated to the reduction in β and compensatory changes in other subunits in the receptor arrangement.

The glycine receptor alpha 3 subunit mRNA expression shows sex-dependent differences in the adult mouse brain

“…Previous GLRA3 studies have mainly focused on what function the subunit has in the brain, i.e. in the cortex [ 19 ], striatum, nucleus accumbens [ 15 , 19 ], hippocampus [ 19 , 25 , 28 ] and the brainstem [ 17 , 29 ]. For instance, McCracken et al (2017) showed that GLRA3-containing GlyRs are found in various areas of the forebrain.…”

“…These findings indicate that Glra3 participates in tonic inhibition in the prefrontal cortex and in both the dorsal striatum and nucleus accumbens [ 19 ]. San Martin et al (2021) investigated the potential role of the GLRA3 subunit in ethanol sensitivity by focusing on the nucleus accumbens [ 15 ]. They concluded that GLRA3 is expressed in low levels in the mouse nucleus accumbens [ 15 ].…”

“…Although GLRA3 is mainly known for its role in spinal circuits, the subunit is also expressed in the developing and adult brain [ 14 ], and a connection between GLRA3, ethanol-mediated effects [ 15 , 16 ] as well as respiratory rhythmic activity [ 17 ] has been reported. Immunohistochemical and electrophysiological analyses in mice have localized GLRA3 to post-synaptic sites in the inner plexiform layer of the retina [ 18 ], the nucleus accumbens [ 15 , 19 ], dorsal striatum and medial prefrontal cortex pyramidal neurons of layer II/III [ 19 ], as well as to both presynaptic glycine transporter 2 (GLYT2)-expressing neurons and postsynaptic neurons in brainstem areas important for respiratory rhythms [ 17 ]. In situ hybridization also locates Glra3 to the cortex, hypothalamus and midbrain ( https://mouse.brain-map.org/experiment/show/70723453 ) [ 20 ].…”

G protein‐coupled receptor 158 modulates sensitivity to the sedative‐hypnotic effect of ethanol in male mice

“…EtOH can pass the blood–brain barrier (BBB) to interact with the CNS and influence the psychological state (Popoola & Cameron, 2018; Schuckit, 2018; Thiele et al, 2000). EtOH can disinhibit dopaminergic neurons by acting on the mesocorticolimbic system including the prefrontal cortex (PFC), ventral tegmental area (VTA), and nucleus accumbens (NAc), while EtOH at a high dose may non‐specifically suppress cortical excitation and cause sedation (Pal et al, 2018; San Martin et al, 2021; Vore et al, 2021). Thus, sensitivity to EtOH is determined by both EtOH metabolism and individual vulnerability of the CNS to EtOH (Aguayo et al, 2014; Chen et al, 2021; Schuckit, 1984).…”

Loss, Gain and Altered Function of GlyR α2 Subunit Mutations in Neurodevelopmental Disorders

“…For example, some missense mutations increase the sensitivity to glycine (α1 I43F ), prolong the decay rate of inhibitory postsynaptic currents (IPSCs; α1 I43F , α1 W170S , α1 Q266E , α1 V280M , α1 R414H ), or induce spontaneous GlyR channel opening (α1 I43F , α1 Y128C , α1 W170S , α1 Q226E , α1 V280M , α1 R414H , β L285R ). GlyRs containing the α3 and α4 subunits have not yet been linked to human disease, but studies using knockout and knockin mice have implicated GlyR α3 in central pain sensitization (Harvey et al, 2004;Werynska et al, 2021), rhythmic breathing (Manzke et al, 2010), and ethanolmediated behaviors (Blednov et al, 2015;San Martin et al, 2021). GlyR α4 is a pseudogene in humans but contributes to touch-evoked escape behaviors in zebrafish (Leacock et al, 2018) and impacts embryonic development and litter sizes in rodents (Nishizono et al, 2020).…”