CHANGES IN MITOCHONDRIAL-FUNCTION IN RESPONSE TO CHANGES IN CYTOSOLIC CA-2+ CONCENTRATION

Microfluorimetric techniques have been used to follow changes in mitochondrial potential (DELTApsi(m)) and in redox state in a variety of mammalian cell types. The characteristic properties of the fluorescence signals used are described in relation to chemiosmotic theory. These techniques have been employed to examine the consequences for mitochondrial function of changing cytosolic Ca2+ concentration ([Ca2+]i), either following Ca2+ influx through voltage-gated channels, or by InsP3- or caffeine-induced mobilisation of Ca2+ from internal stores. In each case, a rise in [Ca2+]i is followed by a transient depolarisation of DELTApsi(m), which is associated with a transient uncoupling of the mitochondria, probably due directly to mitochondrial Ca2+ uptake. This is then usually followed by a secondary increase in the NADH/NAD ratio. This can be attributed to the stimulation of the rate-limiting enzymes of the tricarboxylic acid (TCA) cycle, which are all up-regulated by Ca2+. The changes in NADH autofluorescence are blocked by intracellular application of ruthenium red, which blocks mitochondrial Ca2+ uptake. These data suggest that changes in [Ca2+]i are associated in several cell types and in response to various stimuli, with significant mitochondrial Ca2+ uptake, and that this uptake has substantial consequences for the regulation of mitochondrial oxidative phosphorylation. This provides a direct mechanism to couple ATP supply with changes in [Ca2+]i typically associated with cell activation and increased energy demand.