Neuronal activity modulates the incorporation of newly translated PSD-95 into a robust structure as revealed by STED and MINFLUX

Abstract

The postsynaptic density component PSD-95 undergoes activity-dependent plasticity mechanisms that rely on protein synthesis and structural remodeling. How synaptic activity can influence these dynamics at the single synapse level remains unclear. Here we combine genome-editing, pulse-chase experiments, STED and 3D MINFLUX nanoscopy on hippocampal neuronal cultures to study the integration of newly translated PSD-95 molecules at postsynaptic sites and their rearrangement within individual clusters at near-molecular resolution. We show that the amount of newly translated PSD-95 recruited to individual synapses scales with synaptic size, and modulates in a bidirectional manner, resulting in less new protein following excitatory and more new protein following inhibitory stimulation. Furthermore, we show that within synaptic clusters PSD-95 has a dispersed organization that is largely robust to long-lasting changes in activity. Altogether, this work sheds new light on the mechanisms underlying plasticity at the single synapse level, adding previously inaccessible information.

Sequential replacement of PSD95 subunits in postsynaptic supercomplexes is slowest in the cortex

“…Two recent studies have provided insights into the spacing between PSD95 molecules. MINFLUX revealed a mean nearest neighbor distance of approximately 7 nm (Gürth et al, 2023), while one-step nanoscale expansion microscopy found a preferred spacing of 8-9 nm (Shaib et al, 2023). These measurements, taken within the PSD, may represent distances between neighboring supercomplexes rather than within a single supercomplex, as reported here.…”