Actin mesh, rails, and corrals are present in various types of induced presynapses. (A) Three-color SMLM image of a bead-induced presynapse labeled for ß2-spectrin (DNA-PAINT image), synapsin (DNA-PAINT image), and actin (STORM image). The fourth column is an overlay of the three channels, while the fifth column shows the actin STORM image with highlighted presynaptic actin structures. (B) Actin nanostructures in A+ and A− induced presynapses along the same axon. The top-left panel shows the widefield image of ß2-spectrin (purple), synaptophysin (orange), and actin (gray). The top-center panel is a zoom of the highlighted area in the first image. The top-right panel shows an overlay of widefield images of ß2-spectrin (purple) and synaptophysin (orange) with the STORM image of actin (gray). The bottom-left panel is the isolated STORM image of actin (gray). The bottom-center panel is the isolated 3D-STORM image of actin, color-coded for depth (with Z scale in the top right corner). The bottom-right panel is the STORM image of actin with highlighted presynaptic actin structures in the A+ and A− induced presynapses: active zone mesh (red), actin rails (green), perisynaptic corral (blue). (C) Semi-quantitative assessment of the frequency of identification for presynaptic actin structures: actin mesh (red), rails (green), and corral (blue) from A+ and A− induced presynapses. (D) Quantification of the size of the mesh (red) and corral (blue) on 2D-projected STORM images of A+ and A− induced presynapses. Significance signs on the graph compare to the value of a given nanostructure size between A+ and A− synapses. See Data S1 file for detailed statistics. (E) Actin nanostructures in an identified bead-induced glutamatergic presynapse. First column shows the widefield image of ß2-spectrin (purple), VGLUT (orange), and actin (gray). The second column is a zoom of the highlighted area in the first image. The third column shows an overlay of widefield images of ß2-spectrin (purple) and VGLUT (orange) with the STORM image of actin (gray). The fourth column is 3D-STORM image of actin, color-coded for depth (with Z scale in the top right corner). The fifth column is the STORM image of actin with highlighted presynaptic actin structures: active zone mesh (red), actin rails (green), perisynaptic corral (blue). (F) Actin nanostructures in an identified bead-induced GABAergic presynapse. The first column shows the widefield image of ß2-spectrin (purple), VGAT (green), and actin (gray). The second column is a zoom of the highlighted area in the first image. The third column shows an overlay of widefield images of ß2-spectrin (purple) and VGAT (green) with the STORM image of actin (gray). The fourth column is 3D-STORM image of actin, color-coded for depth (with Z scale in the top right corner). The fifth column is the STORM image of actin with highlighted presynaptic actin structures: active zone mesh (red), actin rails (green), and perisynaptic corral (blue). Scale bars in A, 1 µm; in B, 5 µm (top left panel) and 1 µm (other panels); in E and F, 5 µm (left column panels) and 1 µm (other panels).
Figure 7.

Actin mesh, rails, and corrals are present in various types of induced presynapses. (A) Three-color SMLM image of a bead-induced presynapse labeled for ß2-spectrin (DNA-PAINT image), synapsin (DNA-PAINT image), and actin (STORM image). The fourth column is an overlay of the three channels, while the fifth column shows the actin STORM image with highlighted presynaptic actin structures. (B) Actin nanostructures in A+ and A− induced presynapses along the same axon. The top-left panel shows the widefield image of ß2-spectrin (purple), synaptophysin (orange), and actin (gray). The top-center panel is a zoom of the highlighted area in the first image. The top-right panel shows an overlay of widefield images of ß2-spectrin (purple) and synaptophysin (orange) with the STORM image of actin (gray). The bottom-left panel is the isolated STORM image of actin (gray). The bottom-center panel is the isolated 3D-STORM image of actin, color-coded for depth (with Z scale in the top right corner). The bottom-right panel is the STORM image of actin with highlighted presynaptic actin structures in the A+ and A− induced presynapses: active zone mesh (red), actin rails (green), perisynaptic corral (blue). (C) Semi-quantitative assessment of the frequency of identification for presynaptic actin structures: actin mesh (red), rails (green), and corral (blue) from A+ and A− induced presynapses. (D) Quantification of the size of the mesh (red) and corral (blue) on 2D-projected STORM images of A+ and A− induced presynapses. Significance signs on the graph compare to the value of a given nanostructure size between A+ and A− synapses. See Data S1 file for detailed statistics. (E) Actin nanostructures in an identified bead-induced glutamatergic presynapse. First column shows the widefield image of ß2-spectrin (purple), VGLUT (orange), and actin (gray). The second column is a zoom of the highlighted area in the first image. The third column shows an overlay of widefield images of ß2-spectrin (purple) and VGLUT (orange) with the STORM image of actin (gray). The fourth column is 3D-STORM image of actin, color-coded for depth (with Z scale in the top right corner). The fifth column is the STORM image of actin with highlighted presynaptic actin structures: active zone mesh (red), actin rails (green), perisynaptic corral (blue). (F) Actin nanostructures in an identified bead-induced GABAergic presynapse. The first column shows the widefield image of ß2-spectrin (purple), VGAT (green), and actin (gray). The second column is a zoom of the highlighted area in the first image. The third column shows an overlay of widefield images of ß2-spectrin (purple) and VGAT (green) with the STORM image of actin (gray). The fourth column is 3D-STORM image of actin, color-coded for depth (with Z scale in the top right corner). The fifth column is the STORM image of actin with highlighted presynaptic actin structures: active zone mesh (red), actin rails (green), and perisynaptic corral (blue). Scale bars in A, 1 µm; in B, 5 µm (top left panel) and 1 µm (other panels); in E and F, 5 µm (left column panels) and 1 µm (other panels).

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