Mitochondria respond to Ca 2+ already in the submicromolar range: correlation with redox state

Rapid formation of high-Ca 2+ perimitochondrial cytoplasmic microdomains has been shown to evoke mitochondrial Ca 2+ signal and activate mitochondrial dehydrogenases, however, the significance of submicromolar cytoplasmic Ca 2+ concentrations in the control of mitochondrial metabolism has not been sufficiently elucidated. Here we studied the mitochondrial response to application of Ca 2+ at buffered concentrations in permeabilized rat adrenal glomerulosa cells, in an insulin-producing cell line (INS-1/EK-3) and in an osteosarcoma cell line (143BmA-13). Mitochondrial Ca 2+ concentration was measured with the fluorescent dye rhod-2 and, using an in situ calibration method, with the mitochondrially targeted luminescent protein mt-aequorin. In both endocrine cell types, mitochondrial Ca 2+ concentration increased in response to elevated cytoplasmic Ca 2+ concentration (between 60 and 740 nM) and an increase in mitochondrial Ca 2+ concentration could be revealed already at a cytoplasmic Ca 2+ concentration step from 60–140 nM. Similar responses were observed in the osteosarcoma cell line, although a clearcut response was first observed at 280 nM extramitochondrial Ca 2+ only. As examined in glomerulosa cells, graded increases in cytoplasmic Ca 2+ concentration were associated with graded increases in the reduction of mitochondrial pyridine nucleotides, consistent with Ca 2+ -dependent activation of mitochondrial dehydrogenases. Our data indicate that in addition to therecognized role of high-Ca 2+ cytoplasmic microdomains, also small Ca 2+ signals may influence mitochondrialmetabolism.