A major consequence of brain hypoxia and hypoglycemia, which induces the detrimental effects of stroke, is impaired ATP supply. However, it is not yet clear to which degree reduced cellular ATP production affects Ca(2+) homeostasis and Ca(2+) signaling of glia cells. Here we studied in cultured hippocampal astrocytes the influence of inhibition of cellular energy supply on Ca(2+) load of intracellular stores. Inhibition of glycolysis in the presence of substrates for mitochondrial respiration resulted in an average drop of intracellular ATP levels by 35%. Inhibition of oxidative phosphorylation reduced intracellular ATP on average by 16%. With inhibition of both glycolysis and mitochondrial ATP production, intracellular ATP level was drastically reduced (84%). In astrocytes in Ca(2+)-free buffer, cytosolic [Ca(2+)](i) was dramatically increased due to inhibition of glycolysis, even in the presence of mitochondrial substrates. However, only a minor increase of [Ca(2+)](i) was observed with inhibitors of mitochondrial ATP synthesis. Remarkably, the moderate reduction of ATP levels found with inhibitors of glycolysis caused a severe loss of Ca(2+) from cyclopiazonic acid (CPA)-sensitive Ca(2+) stores. Consequently, inhibition of glycolysis reduced P2Y receptor- or thrombin receptor-evoked Ca(2+) responses on average by 95%, whereas a reduction of only 26% was found with mitochondrial inhibitors. In conclusion, glycolysis is the most important source of ATP for the maintenance of Ca(2+) load in stores that are required for transmitter-induced signaling. These results are consistent with the concept that a local ATP source in the vicinity of endoplasmic reticulum Ca(2+) pumps is required.
Copyright 2000 Wiley-Liss, Inc.