CHANGES IN MITOCHONDRIAL-FUNCTION IN RESPONSE TO CHANGES IN CYTOSOLICCA-2+ CONCENTRATION

Microfluorimetric techniques have been used to follow changes in mitoc hondrial potential (DELTApsi(m)) and in redox state in a variety of ma mmalian cell types. The characteristic properties of the fluorescence signals used are described in relation to chemiosmotic theory. These t echniques have been employed to examine the consequences for mitochond rial function of changing cytosolic Ca2+ concentration ([Ca2+]i), eith er 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 mi tochondria, probably due directly to mitochondrial Ca2+ uptake. This i s then usually followed by a secondary increase in the NADH/NAD ratio. This can be attributed to the stimulation of the rate-limiting enzyme s of the tricarboxylic acid (TCA) cycle, which are all up-regulated by Ca2+. The changes in NADH autofluorescence are blocked by intracellul ar application of ruthenium red, which blocks mitochondrial Ca2+ uptak e. These data suggest that changes in [Ca2+]i are associated in severa l cell types and in response to various stimuli, with significant mito chondrial Ca2+ uptake, and that this uptake has substantial consequenc es 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 dem and.