Fig. 6
From: Astrocytes rescue neuronal health after cisplatin treatment through mitochondrial transfer
Mitochondrial transfer underlies the beneficial effects of astrocytes on calcium dynamics in neurons damaged by cisplatin. a. Neurons were treated with and without cisplatin for 24 h and co-cultured with astrocytes transfected with mito-mCherry (labeled mCh) for 17 h. The calcium response to 20 mM KCl was compared in cisplatin-treated neurons cultured without astrocytes (black bar), or with astrocytes separated into mCherry negative neurons (orange bar; do not contain astrocytic mitochondria) and mCherry positive neurons (blue bar; contain astrocyte-derived mitochondria). The mean increase in the F340:F380 ratio of Fura-2 fluorescence in response to 20 mM KCl was calculated as in Fig. 5. Data represents the mean ± SEM of 3 independent experiments with n = 24 neurons in total. One-way ANOVA, followed by Tukey’s post-hoc test: (**p < 0.01, ***p < 0.0001). b. Resting F340:F380 ratio of Fura-2 of neurons treated with cisplatin followed by co-culture with astrocytes transfected with mito-GFP and scr siRNA or Miro-1 siRNA. c. Mean F340:F380 ratio was calculated for each neuron for first 50 s and data represents the mean ± SEM of 3 independent experiments with n > 80 neurons for each group. Two-way ANOVA: cisplatin x transfected astrocyte interaction: ***p < 0.001 followed by Tukey’s post-hoc test: (***p < 0.001, ****p < 0.0001). d. Mean traces and SEM of the F340:F380 ratio in response to 20 mM KCl stimulus in neurons treated with and without cisplatin and co-cultured with astrocytes transfected with scrambled (Scr) or Miro-1 siRNA. The mean F340:F380 ratio KCl responses were normalized to baseline. e. The change in F340:F380 ratio between baseline and maximum peak value from the 20 mM KCl stimulus was calculated which correlated with [Ca2+]i increase in neurons. Mean increase in F340:F380 ratio ratio was calculated for each neuron for 3 s before 20 mM KCl and subtracted from mean F340:F380 ratio at the maximum response to 20 mM KCl. Data represents the mean ± SEM of 3 independent experiments with n > 80 neurons for each group. Two-way ANOVA: cisplatin x transfected astrocyte interaction: (****p < 0.0001), followed by Tukey’s post-hoc test (****p < 0.0001)