Mitochondrial Calcium Waves by Electrical Stimulation in Cultured Hippocampal Neurons

Abstract

Mitochondria are critical to cellular Ca2+ homeostasis via the sequestering of cytosolic Ca2+ in the mitochondrial matrix. Mitochondrial Ca2+ buffering regulates neuronal activity and neuronal death by shaping cytosolic and presynaptic Ca2+ or controlling energy metabolism. Dysfunction in mitochondrial Ca2+ buffering has been implicated in psychological and neurological disorders. Ca2+ wave propagation refers to the spreading of Ca2+ for buffering and maintaining the associated rise in Ca2+ concentration. We investigated mitochondrial Ca2+ waves in hippocampal neurons using genetically encoded Ca2+ indicators. Neurons transfected with mito-GCaMP5G, mito-RCaMP1h, and CEPIA3mt exhibited evidence of mitochondrial Ca2+ waves with electrical stimulation. These waves were observed with 200 action potentials at 40 Hz or 20 Hz but not with lower frequencies or fewer action potentials. The application of inhibitors of mitochondrial calcium uniporter and oxidative phosphorylation suppressed mitochondrial Ca2+ waves. However, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors and N-methyl-d-aspartate receptor blockade had no effect on mitochondrial Ca2+ wave were propagation. The Ca2+ waves were not observed in endoplasmic reticula, presynaptic terminals, or cytosol in association with electrical stimulation of 200 action potentials at 40 Hz. These results offer novel insights into the mechanisms underlying mitochondrial Ca2+ buffering and the molecular basis of mitochondrial Ca2+ waves in neurons in response to electrical stimulation.