Advanced maternal age and in vitro embryo production (IVP) predispose to pregnancy loss in horses. We
investigated whether mare age and IVP were associated with alterations in mitochondrial (mt) DNA copy number or
function that could compromise oocyte and embryo development. Effects of mare age (,12 vs $12 years) on mtDNA
copy number, ATP content and expression of genes involved in mitochondrial replication (mitochondrial transcription
factor (TFAM), mtDNA polymerase g subunit B (mtPOLB) and mitochondrial single-stranded DNA-binding protein
(SSB)), energy production (ATP synthase-coupling factor 6, mitochondrial-like (ATP-synth_F6)) and oxygen free radical
scavenging (glutathione peroxidase 3 (GPX3)) were investigated in oocytes before and after in vitro maturation (IVM),
and in early embryos. Expression of TFAM, mtPOLB and ATP-synth-F6 declined after IVM (P,0.05). However,
maternal age did not affect oocyte ATP content or expression of genes involved in mitochondrial replication or function.
Day 7 embryos from mares $12 years had fewer mtDNA copies (P¼0.01) and lower mtDNA : total DNA ratios
(P,0.01) than embryos from younger mares, indicating an effect not simply due to lower cell number. Day 8 IVP
embryos had similar mtDNA copy numbers to Day 7 in vivo embryos, but higher mtPOLB (P¼0.013) and a tendency to
reduced GPX3 expression (P¼0.09). The lower mtDNA number in embryos from older mares may compromise
development, but could be an effect rather than cause of developmental retardation. The general down-regulation of genes
involved in mitochondrial replication and function after IVM may compromise resulting embryos.