Presynaptic mitochondrial calcium sequestration influences synaptic transmission. A, The postsynaptic MNTB neuron and presynaptic calyx were simultaneously voltage clamped. Scale bar, 10 μm. B, Trains of stimuli (as in Fig. 3C) produced a markedly depressing EPSC train. At 500 msec after the end of the train, the EPSC had recovered to 81% of control amplitude. C, D,Presynaptic calcium currents recorded in response to the train were unaffected by mitochondrial depolarization. E,Postsynaptic currents normalized to the magnitude of the first EPSC. The absolute magnitude of the EPSC was −4.6 ± 0.4 nA before mitochondrial depolarization and −4.3 ± 0.4 nA after. This 6 ± 2% reduction was a time-dependent run-down and was not statistically significant (n = 3;p = 0.07). Mitochondrial depolarization induced by rotenone and FCCP in the presence of oligomycin had no effect on the initial train amplitude and time course, but it significantly reduced the relative magnitude of the test EPSC. F, Identical results were observed after mitochondrial depolarization or by blocking the calcium uniporter with intracellular Ru360 (1 μm).G, Inclusion of 1 mm EGTA in the presynaptic patch pipette accelerated the decay and reduced the effect of mitochondrial depolarization on the presynaptic calcium transient.H, Presynaptic EGTA at 1 mm abolished the effects of mitochondrial depolarization on the rate of recovery from synaptic depression. Asterisks indicate statistical significance (p < 0.05).