Extended Data Fig. 10. Presynaptic Mitochondria Contribute to the Recovery of Synaptic Efficacy During Prolonged Synaptic Activity.

a, Normalized EPSC amplitudes showing mitochondrial capacity of calcium clearance contributes to synaptic recovery after high-frequency stimulation (HFS). Cortical neurons were used for dual whole-cell patch clamp recording at DIV14–18. The mitochondrial calcium uniporter inhibitor RU360 (10 μM) was added to the intracellular recording solution to block mitochondrial calcium buffering capacity. Recording configuration (total 132 sec duration) consisted of a 2-sec 100 Hz train, a 10-sec recovery phase of four stimuli, and a 120-sec rest phase. The normalized EPSC amplitude was plotted for the first of each HFS and for each of the recovery EPSCs under each condition. Note that the application of RU360 induced a slower synaptic recovery after HFS. Two-way ANOVA revealed a main effect of RU360 treatment (F1, 41 = 10.64, P = 0.0022) and a significant interaction between RU360 treatment and stimuli (F14, 574 = 9.457, P < 0.001). b, Normalized EPSC amplitudes showing that myo6-SNPH-captured presynaptic mitochondria significantly contribute to synaptic recovery after HFS even their calcium buffering capacity is blocked. Cortical neurons were infected with lentiviruses encoding GFP, GFP-tagged myo6 and myo6-T405E, followed by dual whole-cell patch clamp recording at DIV14–18. 10 μM RU360 was applied to intracellular recording solution. The same recording configuration was used as described above. Note that enhanced presynaptic mitochondria anchoring through activating myo6-PAK pathway accelerates synaptic recovery after HFS even when calcium buffering capacity is blocked. Two-way ANOVA revealed a main effect of gene manipulation (F2, 60 = 3.506, P = 0.0363) and a significant interaction between gene manipulation and stimuli (F28, 840 = 2.074, P = 0.001). Data were quantified from the total pairs of neurons indicated in parentheses, and presented as mean ± SEM.