Activity-dependent inhibitory synapse remodeling through gephyrin phosphorylation

Abstract

Maintaining a proper balance between excitation and inhibition is essential for the functioning of neuronal networks. However, little is known about the mechanisms through which excitatory activity can affect inhibitory synapse plasticity. Here we used tagged gephyrin, one of the main scaffolding proteins of the postsynaptic density at GABAergic synapses, to monitor the activity-dependent adaptation of perisomatic inhibitory synapses over prolonged periods of time in hippocampal slice cultures. We find that learning-related activity patterns known to induce N-methyl-daspartate (NMDA) receptor-dependent long-term potentiation and transient optogenetic activation of single neurons induce within hours a robust increase in the formation and size of gephyrin-tagged clusters at inhibitory synapses identified by correlated confocal electron microscopy. This inhibitory morphological plasticity was associated with an increase in spontaneous inhibitory activity but did not require activation of GABA A receptors. Importantly, this activity-dependent inhibitory plasticity was prevented by pharmacological blockade of Ca 2+ /calmodulindependent protein kinase II (CaMKII), it was associated with an increased phosphorylation of gephyrin on a site targeted by CaMKII, and could be prevented or mimicked by gephyrin phosphomutants for this site. These results reveal a homeostatic mechanism through which activity regulates the dynamics and function of perisomatic inhibitory synapses, and they identify a CaMKIIdependent phosphorylation site on gephyrin as critically important for this process.inhibition | gabaergic synapse | plasticity | hippocampus | CaMKII S everal activity-dependent plasticity and homeostatic mechanisms (1, 2) contribute to regulate synaptic strength at excitatory synapses. Similar mechanisms are also expected to finely tune the level of inhibition in response to activity in individual neurons, but the mechanisms remain poorly understood. Different forms of plasticity at GABAergic synapses have been reported based on either presynaptic or postsynaptic mechanisms (3, 4). Similar to receptors at excitatory synapses, GABA A receptors (GABA A Rs), which mediate the fast component of inhibitory transmission, display complex trafficking mechanisms that affect the surface localization and diffusion of receptors (5). The distribution and clustering of GABA A Rs at synapses is tightly regulated through interactions with the scaffolding protein gephyrin, one of the main structural constituent of inhibitory postsynaptic densities. Gephyrin forms multimeric complexes that allow the anchoring of GABA A Rs (6) via molecular mechanisms that include phosphorylation and interactions with the guanine-nucleotide exchange factor collybistin (7-12). In addition to changes in inhibitory strength, more recent in vivo experiments revealed that inhibitory synapses are also dynamic structures that can be formed and eliminated in response to sensory experience (13-15). The mechanisms implicated in the coordinated regulation of excitatory an...

De novo synaptogenesis induced by GABA in the developing mouse cortex

“…Our findings suggest that GABA sets the balance between inhibitory and excitatory synapses in early postnatal stages, laying the foundation for later circuit development (6, 25–27). Because the developing dendrites in our study did not have predetermined spots for inhibitory or excitatory synapses, localization of axonal boutons may be an early step in the precise formation of inhibitory and excitatory circuits (9, 28, 29).…”

An NMDA Receptor-Dependent Mechanism Underlies Inhibitory Synapse Development

“…Finally, although synaptogenic adhesion molecules, transcription factors and signaling molecules have been identified that play important roles in inhibitory synapse development (Bloodgood et al, 2013; Fazzari et al, 2010; Gottmann et al, 2009; Huang and Scheiffele, 2008; Kuzirian et al, 2013; Lin et al, 2008; Siddiqui and Craig, 2011; Sudhof, 2008; Takahashi et al, 2012; Terauchi et al, 2010; Woo et al, 2013; Yim et al, 2013), possible functional interaction between NMDAR signaling and these molecules in the regulation of GABAergic synapse development remains unclear. Recent studies in mature neurons show that NMDARs can regulate GABAergic synapses through calcineurin- (Bannai et al, 2009; Muir et al, 2010), CaMKII- (Flores et al, 2015; Marsden et al, 2010; Petrini et al, 2014) or nitric oxide synthase-dependent mechanisms (Nugent et al, 2007). It would be interesting to examine the role of these signaling pathways in NMDAR-dependent GABAergic synapse development in early immature neurons in the future.…”

Several posttranslational modifications act in concert to regulate gephyrin scaffolding and GABAergic transmission

“…We have recently reported that NMDA ( N -methyl- D -aspartate) receptor-dependent neuronal depolarization regulates gephyrin phosphorylation in a CaMKII-dependent mechanism, to upregulate perisomatic inhibition and facilitate neuronal homeostasis34. Experimentally, mutations in GABA A R subunit genes that indirectly affect postsynaptic gephyrin clustering have been used to generate models for neurodevelopmental and neuropsychiatric disorders3536.…”