Mitochondrial Ca2+ uptake depends on the spatial and temporal profile of cytosolic Ca2+ signals

Using confocal imaging of Rhod-2-loaded HeLa cells, we examined the ability of mitochondria to sequester Ca2+ signals arising from different sources. Mitochondrial Ca2+ (Ca-mit(2+)) uptake was stimulated by inositol 1,4,5-trisphosphate (InsP(3))-evoked Ca2+ release, capacitative Ca2+ entry, and Ca2+ leaking from the endoplasmic reticulum, For each Ca2+ source, the relationship between cytosolic Ca2+ (Ca-cyt(2+)) concentration and Ca-mit(2+) was complex. With Ca-cyt(2+) < 300 nM, a slow and persistent Ca-mit(2+) uptake was observed. If Ca-cyt(2+) increased above similar to 400 nM, Ca-mit(2+) uptake accelerated sharply. For equivalent Ca-mit(2+) increases, the rate of Ca-mit(2+) rise was greater with InsP(3)-evoked Ca2+ signals than any other source. Spatial variation of the Ca-mit(2+) response was observed within individual cells. Both the fraction of responsive mitochondria and the amplitude of the Ca-mit(2+) response were graded in direct proportion to stimulus concentration. Trains of repetitive Ca2+ oscillations did not maintain elevated Ca-mit(2+) levels. Only low frequency Ca2+ transients (<1/15 min) evoked repetitive Ca-mit(2+) signals. Our data indicate that there is a lag between Ca-cyt(2+) and Ca-mit(2+) increases but that mitochondria will accumulate calcium when it is elevated over basal levels regardless of its source. Furthermore, in addition to the characteristics of Ca2+ signals, Ca2+ uniporter desensitization and proximity of mitochondria to InsP(3) receptors modulate mitochondrial Ca2+ responses.