LETTER
Reply to Quistorff and Grunnet: Dysfunctional mitochondria, brain lactate, and lactate dehydrogenase isoforms Quistorff and Grunnet (1), on theoretical grounds, question our suggested explanation of the pathologically increased lactate levels that we reported in prematurely aging mtDNA mutator and normally aging mice (2). A clarification of the chain of events based on our experimental data is, therefore, warranted. The primary cause of lactate pathology in mtDNA mutator mice is precisely known: a mutation drastically decreasing the proofreading function of the catalytic subunit of Polγ. This leads to progressive increase of mtDNA mutation frequency in mitochondria of all cells and premature-aging phenotypes (3). Accumulating mtDNA mutations causes respiratory chain dysfunction, shown by cytochrome c oxidase (COX)/succinate dehydrogenase (SDH) staining (2). This, in turn, causes impaired pyruvate oxidation (3), and increased pyruvate levels would be expected. We have preliminary data to show such increase (measured in blood). Pyruvate to lactate conversion regenerates β-nicotinamide adenine dinucleotide (NAD+), which is essential for glycolysis (2). Twenty-three-week-old mtDNA mutator mice have a strong pathological increase of brain lactate. This is not caused by increased total LDH activity as suggested (1), because there is no such change of total pyruvate–lactate interconversion (figure 5c in ref. 2). Instead, there is a shift in the lactate dehydrogenase-A/ B (LDH-A/B) ratio, a corresponding shift in tetrameric isoenzyme types to those containing more M and fewer H units, and a similar shift in LDH1-5 isoenzyme function. Thus, we find that LDH-A and -B transcriptional activities are regulated to formation of isoenzymes better suited to counteract increased pyruvate levels. Although the isoenzymes have similar Keq values, Km values differ, and because LDH1-5 is inhibited differently by product/substrate, with LDH-1 being more easily inhibited by pyruvate, it is highly unlikely that the same large increase of lactate would have occurred without a changed A/B ratio. In 45-wk-old mtDNA mutator mice, when mtDNA mutation numbers have further increased, total LDH activity is also
E22 | PNAS | February 15, 2011 | vol. 108 | no. 7
increased. However, pyruvate to lactate activity increases much more than lactate to pyruvate activity; thus, the A/B ratio is also further altered and is accompanied by a further shift in LDH1-5 activities to even more efficient pyruvate conversion. The brain allows influx/efflux of lactate and pyruvate; therefore, equilibrium cannot be attained. Similarly, LDH-catalyzed in vivo reactions in cells with progressively dysfunctional mitochondria cannot be regarded as a dead end (4). Quistorff and Grunnet (1) refer to a theoretical study (5) claimed to state “that total LDH activity rather than the isoenzyme pattern determines the steady state lactate concentration” (5). However, Downer et al. (5) do not exclude a role for isoenzyme composition; they only state that “[t]aking into account total [LDH] concentration changes, as well as changes in isoform ratios, is essential” (5). The situation in the brain is particularly complex, because neurons and astrocytes have different metabolic roles. LDH-1, important for rapid build-up of pyruvate, is localized to neurons, whereas LDH-5, capable of mitigating high amounts of pyruvate, is found with LDH-1 in astrocytes (2). Quistorff and Grunnet (1) suggest that the lactate increase could be a consequence of a “changed pyruvate steady state concentration” (1). We concur and suggest that this change is an increase (supported by data) and that the LDH-A/B shift helps the tissue counter the pyruvate increase, thus contributing to the increased lactate levels. Jaime M. Rossa,b,1, Giuseppe Coppotellic, and Lars Olsona,1 Department of Neuroscience, Karolinska Institutet, SE-171 77 Stockholm, Sweden; bNational Institute on Drug Abuse, National Institutes of Health, Baltimore, MD 21224; and cDepartment of Cell and Molecular Biology, Karolinska Institutet, SE-171 77 Stockholm, Sweden a
1. Quistorff B, Grunnet N (2011) High brain lactate is not caused by a shift in the lactate dehydrogenase A/B ratio. Proc Natl Acad Sci USA 108:E21. 2. Ross JM, et al. (2010) High brain lactate is a hallmark of aging and caused by a shift in the lactate dehydrogenase A/B ratio. Proc Natl Acad Sci USA 107:20087–20092. 3. Trifunovic A, et al. (2004) Premature ageing in mice expressing defective mitochondrial DNA polymerase. Nature 429:417–423. 4. Schurr A (2006) Lactate: The ultimate cerebral oxidative energy substrate? J Cereb Blood Flow Metab 26:142–152. 5. Downer JD, Sevinsky JR, Ahn NG, Resing KA, Betterton MD (2006) Incorporating expression data in metabolic modeling: A case study of lactate dehydrogenase. J Theor Biol 240:464–474.
Author contributions: J.M.R., G.C., and L.O. wrote the paper. The authors declare no conflict of interest. 1
To whom correspondence may be addressed. E-mail:
[email protected] or Lars.Olson@ ki.se.
www.pnas.org/cgi/doi/10.1073/pnas.1018791108
