Mitochondrial Malate-Aspartate Shuttle Regulates Mouse Embryo Nutrient Consumption

Abstract

Pyruvate has been considered the sole substrate that can support development of the mouse zygote to the two-cell stage, with lactate able to support development from the two-cell stage. This study has determined for the first time that mitochondrial reducing equivalent shuttles regulate metabolism in the early embryo. Activity of the malate-aspartate shuttle was found to be essential for the metabolism of lactate in the two-cell embryo. Furthermore, the inability of the mouse zygote to use lactate as an energy source was a result of a lack of malate-aspartate shuttle activity. The mRNA for the four enzymes for shuttle activity were detected at all stages of development. It was determined that aspartate was a rate-limiting factor in the activity of the malate-aspartate shuttle in mouse zygotes probably due to the high K m of the cytoplasmic aspartate aminotransferase. Addition of high concentrations of exogenous aspartate to the culture medium enabled mouse zygotes to utilize lactate in the absence of pyruvate and develop normally to the blastocyst stage as well as produce normal viable offspring. This study determined that the malateaspartate shuttle is a key regulator of embryo metabolism and therefore viability and is the first report that mouse zygotes can develop normally to term in the absence of pyruvate.Studies on the preimplantation mouse embryo have indicated that the zygote has an absolute requirement for pyruvate to complete the first cleavage division (1-3). Significantly, lactate is only able to support development from the two-cell stage (4), and glucose could support development from the eight-cell stage (5). Interestingly, mouse zygotes can metabolize both pyruvate and lactate (6, 7) even though cleavage only occurs in the presence of pyruvate. While this has been documented for more than 30 years, there is no explanation for this unusual pattern of metabolism at the zygote nor is it known what changes occur in the embryo that enable the two-cell embryo to use lactate to support development.Energy metabolism in cells is compartmentalized to either the cytoplasm or the mitochondria. Pyruvate is transported into the mitochondria by specific transport proteins and is metabolized within the mitochondria. In contrast, lactate is converted to pyruvate within the cytosol with the concomitant conversion of NAD ϩ to NADH. Cells must maintain a balance of metabolic intermediates between the cytosol and the mitochondria to enable lactate metabolism to continue. An inability to maintain this balance results in impairment of either mitochondrial and/or cytoplasmic metabolism. In addition to many enzymes existing in either the cytoplasm or mitochondria, the pyridine nucleotides (NAD ϩ , NADH, NADP ϩ , and NADPH) are also compartmentalized between the cytoplasm and the mitochondria. Intact mitochondria are impermeable to NADH (8). Therefore to transfer NADH across the mitochondrial membrane for oxidation and ATP production and for the regeneration of cytoplasmic NAD ϩ , there is an indirect pathway involving th...

Regulation of redox metabolism in the mouse oocyte and embryo

“…5A). It also contradicts somehow the recent finding that cytosolic MDH is expressed in mouse oocytes (Lane and Gardner, 2005).…”

Malate dehydrogenase activity and isozyme content in uterus and ovarian tissues of cows during estral cycle

“…Встанов-лено, що активність ензиму забезпечує ріст і розвиток ооцитів, ембріонів та їх прижив-лення [7,9,15,6].…”

The Future of Human Embryo Culture Media – Or Have We Reached the Ceiling?

“…The beneficial effect of amino acids is believed to be due to their use as a metabolic substrate and also their ability to act as osmolyte and a regulator of intracellular pH (Edwards et al 1998;Van Winkle et al 1990). Amino acids also play an essential role as chelators and anti-oxidants and can regulate metabolism and cell differentiation (Gardner 1998;Lane and Gardner 1997a;Lane and Gardner 2005;Lindenbaum 1973;Liu and Foote 1995;Martin and Sutherland 2001).…”