Diversity of glycine receptors in the mouse retina:: Localization of the α4 subunit

Glycine and gamma-aminobutyric acid (GABA) are the major inhibitory neurotransmitters in the retina. Approximately half of the amacrine cells release glycine at their synapses with bipolar, other amacrine, and ganglion cells. Whereas the retinal distributions of glycine receptor (GlyR) subunits alpha 1, alpha 2, and alpha 3 have been mapped, the role of the alpha 4 subunit in retinal circuitry remains unclear. A rabbit polyclonal antiserum was raised against a peptide that comprises the C-terminal 14 amino acids of the mouse GlyR alpha 4 subunit. Using immunocytochemistry, we localized the alpha 4 subunit in the inner plexiform layer IPL) in brightly fluorescent puncta, which represent postsynaptically clustered GlyRs. This was shown by double-labeling sections for GlyR alpha 4 and synaptic markers (bassoon, gephyrin). Double-labeling sections for GlyR alpha 4 and the other GlyR a subunits shows that they are mostly clustered at different synapses; however, similar to 30% of the alpha 4-containing synapses also express the alpha 2 subunit. We also studied the pre- and postsynaptic partners at GlyR alpha 4-containing synapses and found that displaced (ON-) cholinergic amacrine cells prominently expressed the alpha 4 subunit. The density of GlyR alpha 4-expressing synapses in wildtype, Glra1(ot/ot), and Glra3(-/-) mouse retinas did not differ significantly. Thus, there is no apparent compensation of the loss of alpha 1 or alpha 3 subunits by an upregulation of alpha 4 subunit gene expression; however, the alpha 2 subunit is moderately upregulated.