Figure 7. Overexpression of hPLS3... | Rockefeller University Press

Figure 7.

Overexpression of hPLS3 restores Cav2.2 “cluster-like” formations and spontaneous Ca2+transients in Smn-deficient growth cones. (A) LV-mCh and LV-hPLS3 transduced control and Smn−/−;SMN2 growth cones stained against Cav2.2 (gray) and F-actin (Phalloidin, yellow). Normalized mean gray values of Cav2.2 (N = 5, n = 105; ANOVA Kruskal–Wallis, ****P ≤ 0.0001). (B) Maximum intensity projections of SIM images of LV-mCh and LV-hPLS3 transduced control and Smn−/−;SMN2 growth cones stained against Cav2.2 (cyan) and F-actin (Phalloidin, yellow). Arrows indicate “cluster-like” accumulations. (C) Representative plots of Oregon Green 488 BAPTA-1 fluorescent changes over 5 min indicating spontaneous Ca2+ spikes of control and Smn−/−;SMN2 motoneurons transduced with LV-mCh and LV-hPLS3. Quantification of spontaneous Ca2+ spikes per minute (N = 4; n = 60; control LV-mCh: 5.140 ± 0.6073, control LV-hPLS3: 5.910 ± 0.5955, Smn−/−;SMN2 LV-mCh: 2.293 ± 0.3466, Smn−/−;SMN2 LV-hPLS3: 5.163 ± 0.5820; ANOVA Kruskal–Wallis, ***P ≤ 0.001). Bar represents the mean ± SEM. Scale bars: 5 µm.

Overexpression of hPLS3 restores Ca _v 2.2 “cluster-like” formations and spontaneous Ca ²⁺ transients in Smn-deficient growth cones. (A) LV-mCh and LV-hPLS3 transduced control and Smn^−/−;SMN2 growth cones stained against Ca_v2.2 (gray) and F-actin (Phalloidin, yellow). Normalized mean gray values of Ca_v2.2 (N = 5, n = 105; ANOVA Kruskal–Wallis, ****P ≤ 0.0001). (B) Maximum intensity projections of SIM images of LV-mCh and LV-hPLS3 transduced control and Smn^−/−;SMN2 growth cones stained against Ca_v2.2 (cyan) and F-actin (Phalloidin, yellow). Arrows indicate “cluster-like” accumulations. (C) Representative plots of Oregon Green 488 BAPTA-1 fluorescent changes over 5 min indicating spontaneous Ca²⁺ spikes of control and Smn^−/−;SMN2 motoneurons transduced with LV-mCh and LV-hPLS3. Quantification of spontaneous Ca²⁺ spikes per minute (N = 4; n = 60; control LV-mCh: 5.140 ± 0.6073, control LV-hPLS3: 5.910 ± 0.5955, Smn^−/−;SMN2 LV-mCh: 2.293 ± 0.3466, Smn^−/−;SMN2 LV-hPLS3: 5.163 ± 0.5820; ANOVA Kruskal–Wallis, ***P ≤ 0.001). Bar represents the mean ± SEM. Scale bars: 5 µm.