Lactic acidosis in vivo: testing the link between lactate generation and H+ accumulation in ischemic mouse muscle

The link between lactate generation and cellular acidosis has been questioned based on the possibility of H+ generation, independent of lactate production during glycolysis under physiological conditions. Here we test whether glycolytic H+ generation matches lactate production over a physiological pH and lactate range using ischemia applied to the hindlimb of a mouse. We measured the H+ generation and ATP level in vivo using 31P-magnetic resonance spectroscopy and chemically determined intracellular lactate level in the hindlimb muscles. No significant change was found in ATP content by chemical analysis (P > 0.1), in agreement with the stoichiometric decline in phosphocreatine (20.2 ± 1.2 mM) vs. rise in Pi (18.7 ± 2.0 mM), as measured by 31P-magnetic resonance spectroscopy. A substantial drop in pH from 7.0 to 6.7 and lactate accumulation to 25 mM were found during 25 min of ischemia. The rise in H+ generation closely agreed with the accumulation of lactate, as shown by a close correlation with a slope near identity (0.98; r2 = 0.86). This agreement between glycolytic H+ production and elevation of lactate is confirmed by an analysis of the underlying reactions involved in glycolysis in vivo and supports the concept of lactic acidosis under conditions that substantially elevate lactate and drop pH. However, this link is expected to fail with conditions that deplete phosphocreatine, leading to net ATP hydrolysis and nonglycolytic H+ generation. Thus both direct measurements and an analysis of the stoichiometry of glycolysis in vivo support lactate acidosis as a robust concept for physiological conditions of the muscle cell.