Figure 3.

Only a subpopulation of release sites is enriched with GluN2 subunits

a Example synapse of DNA-PAINT localizations of PSD-95, GluN2A, GluN2B, and Munc13–1 showing arrangement of receptor subunits relative to release sites. Markers as described in Fig. 2a. b Munc13–1 autocorrelation decayed rapidly and plateaued below one, consistent with its c numerous and small detected Munc13–1 NCs. d-e Munc13–1 NCs were, on average, de-enriched with GluN2A and GluN2B at distances <55 nm (shading). f Schematic indicates possible configurations of GluN2 and Munc13–1 NCs. g Schematic of NC pairing to reveal stereotyped distances of closely-associated NCs. h Paired Munc13–1 NCs were enriched with GluN2A as well as i GluN2B within 55 nm of their peak. j-k 20.5% and 24.1% of Munc13–1 NCs were statistically enriched with GluN2A or GluN2B. l 26.7% and 32.7% of Munc13–1 NCs had a GluN2A or GluN2B NC peak within 60 nm. m-n 10.9% and 10.6% of Munc13–1 NCs spatially overlapped with GluN2A or GluN2B NCs. Data in b, d-e, and h-i are means ± SEM shading. Points in c (left) are synapses and (right) NCs, with lines at mean ± SEM. Data in l are shown as frequency histograms (20 nm bins), with dashed line indicating the division summarized in inset pie charts. N = 173 GluN2A, 274 GluN2B, 478 Munc13–1 NCs from 74 synapses throughout, except h-i, where N = 113 Munc13–1 NCs when paired with GluN2A and 152 when paired with GluN2B. Specific n per data point in b varies from 74 due to data processing, see methods for details. *p<0.05, **p<0.01, ***p<0.001.