Mitochondrial activation and the pyruvate paradox in a human cell line

Pyruvate promotes hyperpolarization of the inner mitochondrial membrane. However, in isolated mitochondria, pyruvate could participate in a futile cycle leading to mitochondrial depolarization. Here, we investigated this paradox in intact human cells by measuring parameters reflecting mitochondrial activation in response to 1 mM pyruvate and 5 mM glucose. NAD(P)H levels were elevated similarly by both substrates. Conversely, pyruvate induced a first transient phase of mitochondrial depolarization before the establishment of the expected sustained hyperpolarization. This correlated with kinetics of cytosolic ATP levels exhibiting a first phase decrease followed by an increase. Therefore, pyruvate transiently depolarizes mitochondria and reduces ATP in intact cells.