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Notizen: Abstract: The effect of [H+] on the rate of glycolysis was investigated in glioma C6 and fibroblast BHK-21 cells and in synaptosomes from rat brain. The rates of lactate production at an extracellular pH (pHe) of 6.2, 7.4, and 7.8 were correlated with intracellular [ATP], [ADP], and [Pi] ([ATP]i, [ADP]i, and [Pi]i, respectively) and, when relevant, creatine phosphate (PCr) as well as with the levels of several glycolytic intermediates. In C6 cells cytosolic [H+] was measured simultaneously together with [Ca2+], [K+], [Na+], and membrane potentials. In all three systems studied, an increase in [H+]e suppressed whereas a fall enhanced the rate of lactate generation. Changes in pHe produced no simple correlation between the amount of lactate formed and alterations either in the absolute [ATP], [ADP], [Pi], and [PCr] or their ratios but did correlate with the levels of glycolytic intermediates. Higher [fructose-1,6-bisphosphate] and [glyceraldehyde-3-phosphate] and lower [glucose-6-phosphate] and [fructose-6-phosphate] accompanied faster glycolytic activity. Addition of rotenone markedly enhanced glycolysis at all pHe values studied. The increases were larger at higher [H+] so that the rate of lactate generation was only slightly lower at pH 6.2 than at 7.4 or 7.8. With rotenone present, [ATP] (and where relevant [PCr]) fell and [ADP] and [Pi] rose under all pHe conditions. Simultaneously [glucose-6-phosphate] and [fructose-6-phosphate] decreased whereas [fructose-1,6-bisphosphate] and [glyceraldehyde-3-phosphate] increased; the levels of the last two were similar at pH 6.2 and 7.4. Alterations in concentrations of cations other than H+ were small and unlikely to contribute to the regulation of glycolysis. It is concluded that (a) under intracellular conditions, in the presence of a high [ATP] and a low [ADP] and [AMP], a fall in [H+] powerfully inhibits phosphofructokinase; lactate production correlates with the levels of glycolytic intermediates. (b) A small decline in cellular energy state is sufficient to release phosphofructokinase inhibition by protons. (c) Once activated by the decrease in energy level, the steady-state glycolytic rate is dependent on other factors, such as glycolytic intermediates and enzymes.