Although mitochondria have been the object of intensive study over many decades, some aspects of their metabolism remain to be fully elucidated, including the L L-lactate metabolism. We review here the novel insights arisen from investigations on [7]. Our present position, based on our own work and that of others reviewed here is that the evidence for mitochondrial metabolism of L L-LAC, due to the existence of both L L-LAC carrier-mediated transport processes and of the mitochondrial L L-lactate dehydrogenase (mL-LDH) is now compelling. SpermatozoaThe first evidence in favour of the presence of a putative mL-LDH was from Clausen who found that about 40% of total activity in sperm cells was present in a particulate fraction showing high succinate dehydrogenase activity [8].The existence of the mL-LDH which can reduce pyridine nucleotide in the mitochondrial matrix was demonstrated in hypotonically treated rabbit epididymal spermatozoa by means of oxygen uptake and fluorimetric studies [9]. It was also shown that the mL-LDH functions actively in these cells. In sperm cells the cytosolic L L-lactate dehydrogenase (cL-LDH) and mL-LDH appear to be the same isoenzyme unique to those cells: L-LDH-X. The dual localization of the enzyme enables mammalian spermatozoa to exchange cytosolic and mitochondrial reducing equivalents by means of a L L-LAC/ pyruvate (PYR) shuttle, as suggested by Blanco et al. [10] and reconstituted later [11]. LiverSince liver possesses the enzymatic machinery for gluconeogenesis (GNG) and since L L-LAC is a major substrate for GNG, liver play a major role in L L-LAC metabolism. Indeed, metabolism of L L-LAC has traditionally been considered solely as a function of the cL-LDH in spite of the fact that the L L-LAC oxidation by mitochondria and the existence of an mL-LDH in the inner compartments of isolated rat liver mitochondria (RLM) has been widely reported [12][13][14][15]. In a recent detailed investigation de Bari et al. [16] showed that externally added L L-LAC can be oxidized in the matrix by an mL-LDH, with reduction of intramitochondrial NAD(P) +

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As part of an ongoing study of l-lactate metabolism both in normal and in cancer cells, we investigated whether and how l-lactate metabolism occurs in mitochondria of human hepatocellular carcinoma (Hep G2) cells. We found that Hep G2 cell mitochondria (Hep G2-M) possess an l-lactate dehydrogenase (ml-LDH) restricted to the inner mitochondrial compartments as shown by immunological analysis, confocal microscopy and by assaying ml-LDH activity in solubilized mitochondria. Cytosolic and mitochondrial l-LDHs were found to differ from one another in their saturation kinetics. Having shown that l-lactate itself can enter Hep G2 cells, we found that Hep G2-M swell in ammonium l-lactate, but not in ammonium pyruvate solutions, in a manner inhibited by mersalyl, this showing the occurrence of a carrier-mediated l-lactate transport in these mitochondria. Occurrence of the l-lactate/pyruvate shuttle and the appearance outside mitochondria of oxaloacetate, malate and citrate arising from l-lactate uptake and metabolism together with the low oxygen consumption and membrane potential generation are in favor of an anaplerotic role for l-LAC in Hep G2-M.

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