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. 2021 Mar 30;34(13):108923. doi: 10.1016/j.celrep.2021.108923

Figure 1.

Figure 1

Lesion-induced spine loss and subsequent recovery are accompanied by AMPAR redistribution

(A) OTCs from GFP-expressing mice (green) and stained with antibodies against the GluA2 subunit of AMPARs (magenta). Representative image of a dendritic stretch pre- (top; center: higher magnification) and post- (bottom) expansion.

(B) Representative example of a stained dendritic stretch with the original GluA2 staining (red, tGluA2), the GluA2 staining subdivided between surface (magenta, sGluA2) and internal (cyan, iGluA2) pools based on the GFP reconstruction (top), and their surface reconstruction using Imaris (below). The pseudocolor convention remains the same throughout the article.

(C) Representative surface reconstructions of stubby, thin, and mushroom spines and shafts (bottom row). GluA2 and GFP staining are shown in the top row; GluA2 staining is displayed in 2 pseudocolors representing the 2 different pools (surface GluA2 [sGluA2] in magenta and internal GluA2 [iGluA2] in cyan) based on the GFP reconstruction. The surface reconstruction of the GFP staining has been omitted in the center row to enable visualization of the complete GluA2 reconstruction.

(D) Quantification of GluA2 distribution normalized to GFP volume, shown as percentage of total GluA2/GFP corresponding to (C); n = 5 neurons, 2 experiments.

(E) Quantification of surface GluA2, shown as percentage of total GluA2 per compartment corresponding to (C); n = 5 neurons, 2 experiments.

(F) Anatomy of an entorhinal-hippocampal organotypic slice culture showing the position of the lesion and timeline of imaging experiments. CA, cornu ammonis; DG, dentate gyrus; DPL, day post-lesion; EC, entorhinal cortex; IML, inner molecular layer; OML, outer molecular layer.

(G) Representative pictures of dendritic stretches of dentate granule cells in non-denervated and denervated OTCs. D, denervated; ND, non-denervated.

(H) Quantification of total spine (upper graph) and mushroom spine (lower graph) density as relative ratios to baseline (DPL0) corresponding to (G); n = 5–7 neurons per condition, 4 experiments.

(I–K) Representative surface reconstructions of mushroom (I), thin (J), and stubby (K) spines from non-denervated and denervated OTCs at DPL2 and DPL21.

(L) Quantification of surface GluA2 in D and ND OTCs, represented as percentage of total GluA2 in each compartment corresponding to (I)–(K); n = 5 neurons per condition, 2 experiments.

Scale bars: 5 μm (A top and bottom), 2 μm (A center, B, and G), 1 μm (C and I–K). Graphs show means ± SEMs. ∗p < 0.05; ∗∗p < 0.01; ∗∗∗p < 0.001. Exact p values in Table S1.

See also Figures S1 and S2.