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A novel technique for quantitative measurement of free Ca 2+ concentration in rat heart mitochondria
Author(s) -
Sheu SheyShing,
Sharma Virendra K.
Publication year - 1999
Publication title -
the journal of physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.802
H-Index - 240
eISSN - 1469-7793
pISSN - 0022-3751
DOI - 10.1111/j.1469-7793.1999.0577p.x
Subject(s) - thapsigargin , mitochondrion , xenopus , cytosol , endoplasmic reticulum , biophysics , calcium , chemistry , endocrinology , biology , biochemistry , organic chemistry , gene , enzyme
1 The free mitochondrial Ca 2+ concentration ([Ca 2+ ] m ) in rat heart mitochondria was measured quantitatively by loading the mitochondria with fura‐2 and then injecting them into Xenopus laevis oocytes. 2 When oocytes were incubated with a physiological solution, the free cytosolic Ca 2+ concentration ([Ca 2+ ] c ) in the oocytes was 82 ± 11 nM ( n = 20 , mean ± s.e.m. ) and the [Ca 2+ ] m of the injected rat heart mitochondria was 116 ± 10 nM ( n = 18 , mean ± s.e.m. ). 3 Inhibition of the oocyte endoplasmic reticular Ca 2+ ‐ATPase with thapsigargin produced a transient increase in averaged [Ca 2+ ] c at sub‐micromolar concentrations. 4 Injection of cardiac mitochondria blunted the peak and prolonged the duration of thapsigargin‐induced [Ca 2+ ] c transients as a result of Ca 2+ sequestration by the cardiac mitochondria. 5 These results demonstrate that the present technique provides a new approach for studying [Ca 2+ ] m regulation quantitatively under physiological environments. Furthermore, it clearly shows that cardiac mitochondria can modify the shape of thapsigargin‐induced cytosolic Ca 2+ pulses in Xenopus oocytes.