Benzodiazepine ligands rapidly influence GABAA receptor diffusion and clustering at hippocampal inhibitory synapses

Benzodiazepines (BZDs) are widely used in the treatment of a variety of neurological and psychiatric conditions including anxiety, insomnia and epilepsy. BZDs are thought to act predominantly by affecting the gating of GABAA receptor channels, resulting in enhanced GABA-mediated currents in neurons. However, mutations mimicking the effect of BZDs on GABAAR channel gating have been shown to also impact the membrane dynamics and synaptic anchoring of the receptors. Here, using single molecule tracking combined with electrophysiological recordings, we show that BZD ligands rapidly influence the dynamic behavior of GABAARs in hippocampal neurons. Application of the inverse BZD agonist DMCM rapidly increased the diffusion and reduced the clustering of GABAARs at synapses, resulting in reduced postsynaptic currents. Conversely, the BZD full agonist diazepam had little effect at rest but reduced lateral diffusion and increased synaptic stabilization and clustering of GABAARs upon sustained neuronal activity, resulting in enhanced potency of inhibitory synapses. These effects occurred in the absence of detectable changes in gephyrin clusters, suggesting they did not reflect a rapid dispersion of the synaptic scaffold. Thus, alterations of the diffusion and synaptic anchoring of GABAARs represent a novel, unsuspected mechanism through which BZDs rapidly modulate GABA signaling in central neurons.