By using an endothelial cell line (ECV304), derived from human umbilical vein and transfected with recombinant aequorin targeted to the mitochondrial matrix, we find that stimulation with ATP evokes long lasting increases in mitochondrial Ca ([Ca]) that largely depend on Ca influx. In these cells, the release of stored Ca is inefficient at elevating [Ca]. Consequently it appears that in ECV304 cells, bulk cytosolic Ca ([Ca]) is the main determinant of [Ca] changes.

In ECV304 cells <4% of mitochondria are within 700 nm of the endoplasmic reticulum as opposed to 65% in HeLa cells, whereas 14% are within 700 nm of the inner surface of the plasma membrane, compared with <6% in HeLa cells. Following Ca depletion, readdition of extracellular Ca evokes an increase in [Ca] but not in [Ca]. Under these conditions, microdomains of high [Ca] may occur beneath the plasma membrane of ECV304 cells resulting in the preferential elevation of Ca in mitochondria located in this region.

A model is discussed in which the localization of mitochondria with respect to Ca sources is the main determinant of their in situ Ca uptake kinetics. Thus, in any given cell type mitochondria may be localized to suit the energy and metabolic demands of their physiological actions.

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