Calcium signalling in sarcoplasmic reticulum, cytoplasm and mitochondria during activation of rabbit aorta myocytes

This study investigated the relationship between cytoplasmic, mitochondrial, and sarcoplasmic reticulum (SR) [Ca2+] in rabbit aorta smooth muscle cells, following cell activation. Smooth muscle cells were loaded with the Ca2+-sensitive fluorescent indicator Mag-Fura-2-AM, and then either permeabilized by exposure to saponin, or dialyzed with a patch pipette in the whole-cell configuration to remove cytoplasmic indicator. When the intracellular solution contained millimolar EGTA or BAPTA, activation of SR Ca2+ release through IP3 or ryanodine receptors induced a decrease in the [Ca2+] reported by Mag-Fura-2. However, when EGTA was present at less than or equal to 100 mu M, the same stimuli caused an increase in the [Ca2+] reported by Mag-Fura-2. The increase in [Ca2+] caused by phenylephrine or caffeine was delayed, and prolonged, with respect to the cytoplasmic Ca2+ transient. Evidence is presented that this Mag-Fura-2 signal reflected a rise in mitochondrial [Ca2+]. Agents that inhibit mitochondrial function, such as FCCP or cyanide in combination with oligomycin B, converted the increase in organelle Mag-Fura-e fluorescence to a decrease, while also prolonging the cytoplasmic Ca2+ transient. There was considerable similarity between the localization of Mag-Fura-2 fluorescence and the mitochondria-selective indicator tetramethylrhodamine ethyl ester. Thus, we propose that there is close functional integration between the SR and mitochondria in aorta smooth muscle cells, with mitochondria taking up Ca2+ from the cytoplasm following cell activation. (C) 2000 Harcourt Publishers Ltd.