Mitochondrial Ca²⁺ uptake prevents desynchronization of quantal release and minimizes depletion during repetitive stimulation of mouse motor nerve terminals

We investigated how inhibition of mitochondrial Ca²⁺ uptake affects transmitter release from mouse motor terminals during brief trains of action potentials (500 at 50 Hz) in physiological bath [Ca²⁺]. When mitochondrial Ca²⁺ uptake was inhibited by depolarizing mitochondria with antimycin A1 or carbonyl cyanide m-chlorophenyl-hydrazone, the stimulation-induced increase in cytosolic [Ca²⁺] was greater (> 10 μM, compared to ≤ 1μM in control solution), the quantal content of the endplate potential (EPP) depressed more rapidly (∼84% depression compared to ∼8% in controls), and asynchronous release during the stimulus train reached higher frequencies (peak rates of ∼6000 s^-1 compared to ∼75 s^-1 in controls). These effects of mitochondrial depolarization were not accompanied by a significant change in EPP quantal content or the rate of asynchronous release during 1 Hz stimulation, and were not seen in oligomycin, which blocks mitochondrial ATP synthesis without depolarizing mitochondria. Inhibition of endoplasmic reticular Ca²⁺ uptake with cyclopiazonic acid also had little effect on stimulation-induced changes in cytosolic [Ca²⁺] or EPP amplitude. We hypothesize that the high rate of asynchronous release evoked by stimulation during mitochondrial depolarization was produced by the elevation of cytosolic [ Ca²⁺], and contributed to the accelerated depression of phasic release by reducing the availability of releasable vesicles. During mitochondrial depolarization, the post-tetanic potentiation of the EPP observed under control conditions was replaced by a post-tetanic depression with a slow time course of recovery. Thus, mitochondrial Ca²⁺ uptake is essential for sustaining phasic release, and thus neuromuscular transmission, during and following tetanic stimulation.