Mitochondria recycle Ca2+ to the endoplasmic reticulum and prevent the depletion of neighboring endoplasmic reticulum regions

To study Ca2+ fluxes between mitochondria and the endoplasmic reticulum (ER ), we used "cameleon" indicators targeted to the cytosol, the ER lumen, and the mitochondrial matrix. High affinity mitochondrial probes saturated in similar to 20% of mitochondria during histamine stimulation of HeLa cells, whereas a low affinity probe reported averaged peak values of 106 +/- 5 muM , indicating that Ca2+ transients reach high levels in a fraction of mitoch ondria. In concurrent ER measurements, [Ca2+](ER) averaged 371 +/- 21 muM: at rest and decreased to 133 +/- 14 muM and 59 +/- 5 muM upon stimulation w ith histamine and thapsigargin, respectively, indicating that substantial E R refilling occur during agonist stimulation. A larger ER depletion was obs erved when mitochondrial Ca2+ uptake was prevented by oligomycin and roteno ne or when Ca2+ efflux from mitochondria was blocked by CGP 37157, indicati ng that some of the Ca2+ taken up by mitochondria is re-used for ER refilli ng. Accordingly, ER regions close to mitochondria released less Ca2+ than E R regions lacking mitochondria. The ER heterogeneity was abolished by thaps igargin, oligomycin/rotenone, or CGP 37157, indicating that mitochondrial C a2+ uptake locally modulate ER refilling. These observations indicate that some mitochondria are very close to the sites of Ca2+ release and recycle a substantial portion of the captured Ca2+ back to vicinal ER domains. The d istance between the two organelles thus determines both the amplitude of mi tochondrial Ca2+ signals and the filling state of neighboring ER regions.