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. 2015 Dec 2;35(48):15996–16011. doi: 10.1523/JNEUROSCI.2068-15.2015

Figure 2.

Figure 2.

Enhancing neuronal activity alters mitochondrial trafficking dynamics and morphology in astrocytic processes in situ. A, Representative confocal image of an organotypic slice hippocampal astrocyte AV-transduced with mtdsRed2 and EGFP. Scale bar, 5 μm. Ai, Imaging protocol where drug treatments (unless otherwise stated) were perfused while imaging for 5 min after a 5 min period of imaging basal trafficking and imaged 5 min after treatment. Example kymograph shows mitochondrial trafficking before (pre), during and after (post) glutamate treatment (100 μm glutamate + 1 μm glycine). Scale bar, 5 μm. B, Mitochondrial mobility in situ, defined as percentage moving, in the 5 min period post-treatment [excluding glutamate + 15 min and 4-AP + 30 min; control (combined pre): n = 39 cells, 36 slices; glutamate: n = 8 cells, 8 slices; glutamate + 15 min: n = 3 cells, 3 slices; NBQX (50 μm) + glutamate: n = 11 cells, 11 slices; MCPG (1 mm) + glutamate: n = 12 cells, 7 slices; D-APV (50 μm) + glutamate: n = 5 cells, 3 slices; 0 [Ca2+]e + glutamate: n = 4 cells, 3 slices; 4AP (100 μm): n = 7 cells, 7 slices; 4-AP + 30 min: n = 3 cells, 3 slices; 0 [Ca2+]e + 4-AP: n = 4 cells, 4 slices; TTX (1 μm): n = 3 cells, 3 slices). C, Velocity of moving mitochondria 5 min post-treatment (excluding glutamate + 15 min and 4-AP + 30 min; control (combined pre): n = 38 mitochondria, 36 slices; glutamate n = 13 mitochondria, 4 slices; glutamate + 15 min n = 23 mitochondria, 3 slices; 0 [Ca2+]e + glutamate: n = 11 mitochondria, 4 slices; 4AP: n = 11 mitochondria, 4 slices; 4-AP + 30 min: n = 18 mitochondria, 4 slices; 0 [Ca2+]e + 4-AP: n = 12 mitochondria, 4 slices; TTX: n = 16 mitochondria, 3 slices). D, Quantification of mitochondrial length post-treatment in situ (control: n = 53 cells, 53 slices; glutamate: n = 58 cells, 58 slices; glutamate + 24 h: n = 10 cells, 10 slices; 0 [Ca2+]e + glutamate: n = 24 cells, 24 slices; MK-801 (100 μm) + glutamate: n = 24 cells, 24 slices; NBQX + glutamate: n = 25 cells, 25 slices; 4AP: n = 11 cells, 11 slices). E, Normalized number of mitochondria before, during, and after live glutamate treatment in situ (pre: n = 3 cells, 3 slices; glutamate: n = 3 cells, 3 slices; post: n = 3 cells, 3 slices). F, Mitochondrial length post-treatment in pure cortical astrocyte cultures with glutamate treatment and glutamate + MK-801 (100 μm; control and glutamate treatment: n = 7 cells; glutamate + MK-801: n = 10 cells). *p < 0.05, **p < 0.01, ***p < 0.001.