Presynaptic actin nanostructures are distinctly dependent on Arp2/3 and formins. (A–C) Images of induced presynapses from cultured neurons 2 d after bead seeding at 8 div. The first column shows a widefield image of ß2-spectrin (purple), synapsin (orange), and actin (gray). The second column is a zoom of the highlighted area in the first column image. The third column shows an overlay of widefield images of ß2-spectrin (purple) and synapsin (orange) with the STORM image of actin (gray). The fourth column is the isolated 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). (A) Representative examples from neuronal cultures treated with DMSO for 1 h (control condition). (B) Two examples after treatment with 50 μM CK666 for 1 h. (C) Two examples after treatment with 30 µM SMIFH2 for 1 h. (D) Semi-quantitative assessment of the frequency of identification for presynaptic actin structures: actin mesh, rails, and corrals, from all the presynapses imaged in the control condition (ctrl) or after treatment with CK666 (CK) or SMIFH2 (SMIF). (E) Two-color STORM images of actin and Arp2/3 (anti-p34 antibody) at two representative bead-induced presynapses. The first column shows an overlay of the widefield image of synaptophysin (purple) with the STORM images of actin (gray) and Arp2/3 (yellow). The second column shows the overlayed STORM images of actin (gray) and Arp2/3 (yellow). The third column adds highlighted presynaptic actin structures: active zone mesh (red), actin rails (green), and perisynaptic corral (blue) to the image in the second column. Scale bars in A–C, 5 µm (first column) and 1 µm (other columns); in E, 1 µm.
Figure 8.

Presynaptic actin nanostructures are distinctly dependent on Arp2/3 and formins. (A–C) Images of induced presynapses from cultured neurons 2 d after bead seeding at 8 div. The first column shows a widefield image of ß2-spectrin (purple), synapsin (orange), and actin (gray). The second column is a zoom of the highlighted area in the first column image. The third column shows an overlay of widefield images of ß2-spectrin (purple) and synapsin (orange) with the STORM image of actin (gray). The fourth column is the isolated 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). (A) Representative examples from neuronal cultures treated with DMSO for 1 h (control condition). (B) Two examples after treatment with 50 μM CK666 for 1 h. (C) Two examples after treatment with 30 µM SMIFH2 for 1 h. (D) Semi-quantitative assessment of the frequency of identification for presynaptic actin structures: actin mesh, rails, and corrals, from all the presynapses imaged in the control condition (ctrl) or after treatment with CK666 (CK) or SMIFH2 (SMIF). (E) Two-color STORM images of actin and Arp2/3 (anti-p34 antibody) at two representative bead-induced presynapses. The first column shows an overlay of the widefield image of synaptophysin (purple) with the STORM images of actin (gray) and Arp2/3 (yellow). The second column shows the overlayed STORM images of actin (gray) and Arp2/3 (yellow). The third column adds highlighted presynaptic actin structures: active zone mesh (red), actin rails (green), and perisynaptic corral (blue) to the image in the second column. Scale bars in A–C, 5 µm (first column) and 1 µm (other columns); in E, 1 µm.

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