Differential regulation of glycinergic and GABAergic nanocolumns at mixed inhibitory synapses

• Published in: EMBO reports Vol. 22; no. 7; pp. e52154 - n/a
• Main Authors: Yang, Xiaojuan, Le Corronc, Hervé, Legendre, Pascal, Triller, Antoine, Specht, Christian G
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 05.07.2021; Springer Nature B.V; EMBO Press; John Wiley and Sons Inc
• Subjects: Clustering; direct stochastic optical reconstruction microscopy; Domains; EMBO27; Functional plasticity; GABAAR; Gephyrin; GlyR; Life Sciences; Phosphorylation; Proteins; Receptors; Scaffolds; single‐molecule localization microscopy; Spinal cord; sub‐synaptic domains; Synapses; Synaptic clustering; γ-Aminobutyric acid; γ-Aminobutyric acid A receptors; GABA; R; sub-synaptic domains; single-molecule localization microscopy; direct stochastic optical reconstruction microscopy; GlyR; sub-synaptic domains Subject Category Neuroscience; GABAAR; GABA A R; direct stochastic optical reconstruction microscopy GABA A R GlyR single-molecule localization microscopy sub-synaptic domains Subject Category Neuroscience
• ISSN: 1469-221X; 1469-3178; 1469-3178
• DOI: 10.15252/embr.202052154

Super-resolution imaging has revealed that key synaptic proteins are dynamically organized within sub-synaptic domains (SSDs). To examine how different inhibitory receptors are regulated, we carried out dual-color direct stochastic optical reconstruction microscopy (dSTORM) of GlyRs and GABA A Rs at mixed inhibitory synapses in spinal cord neurons. We show that endogenous GlyRs and GABA A Rs as well as their common scaffold protein gephyrin form SSDs that align with pre-synaptic RIM1/2, thus creating trans-synaptic nanocolumns. Strikingly, GlyRs and GABA A Rs occupy different sub-synaptic spaces, exhibiting only a partial overlap at mixed inhibitory synapses. When network activity is increased by 4-aminopyridine treatment, the GABA A R copy numbers and the number of GABA A R SSDs are reduced, while GlyRs remain largely unchanged. This differential regulation is likely the result of changes in gephyrin phosphorylation that preferentially occurs outside of SSDs. The activity-dependent regulation of GABA A Rs versus GlyRs suggests that different signaling pathways control the receptors' sub-synaptic clustering. Taken together, our data reinforce the notion that the precise sub-synaptic organization of GlyRs, GABA A Rs, and gephyrin has functional consequences for the plasticity of mixed inhibitory synapses. SYNOPSIS This study analyzes the internal organization of mixed inhibitory spinal cord synapses using dSTORM and reveals the existence of trans-synaptic nanocolumns containing GlyRs and GABA A Rs that are differentially regulated in response to altered network activity. Sub-synaptic domains (SSDs) are aligned in trans-synaptic nanocolumns at inhibitory synapses. SSDs formed by GlyRs and GABA A Rs are differentially spatially organized at mixed inhibitory synapses. GABA A Rs but not GlyRs show activity-dependent regulation at mixed inhibitory synapses. Gephyrin phosphorylation is compartmentalized in SSDs within the synaptic scaffold. Graphical Abstract This study analyzes the internal organization of mixed inhibitory spinal cord synapses using dSTORM and reveals the existence of trans-synaptic nanocolumns containing GlyRs and GABA A Rs that are differentially regulated in response to altered network activity.