科研之友
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摘要
Mitochondria extrude protons across their inner membrane to generate the mitochondrial membrane potential (Delta Psi(m)) and pH gradient (Delta pH(m)) that both power ATP synthesis. Mitochondrial uptake and efflux of many ions and metabolites are driven exclusively by Delta pH(m), whose in situ regulation is poorly characterized. Here, we report the first dynamic measurements of Delta pH(m) in living cells, using a mitochondrially targeted, pH-sensitive YFP (SypHer) combined with a cytosolic pH indicator (5-(and 6)-carboxy-SNARF-1). The resting matrix pH (similar to 7.6) and Delta pH(m) (similar to 0.45) of HeLa cells at 37 degrees C were lower than previously reported. Unexpectedly, mitochondrial pH and Delta pH(m) decreased during cytosolic Ca2+ elevations. The drop in matrix pH was due to cytosolic acid generated by plasma membrane Ca2+-ATPases and transmitted to mitochondria by P-i/H+ symport and K+/H+ exchange, whereas the decrease in Delta pH(m) reflected the low H+-buffering power of mitochondria (similar to 5mM, pH 7.8) compared with the cytosol (similar to 20 mM, pH 7.4). Upon agonist washout and restoration of cytosolic Ca2+ and pH, mitochondria alkalinized and Delta pH(m) increased. In permeabilized cells, a decrease in bath pH from 7.4 to 7.2 rapidly decreased mitochondrial pH, whereas the addition of 10 mu M Ca2+ caused a delayed and smaller alkalinization. These findings indicate that the mitochondrial matrix pH and Delta pH(m) are regulated by opposing Ca2+-dependent processes of stimulated mitochondrial respiration and cytosolic acidification.
- 出版日期2011-4-1
