Effect of vitrification on mitochondrial distribution and membrane potential in mouse two pronuclear (2‐PN) embryos

Abstract

The present study was designed to investigate the effect of vitrification on mitochondrial distribution, membrane potential (Δψ) and microtubule distribution in mouse 2-PN embryos, as well as to document the relationship between mitochondrial distribution and developmental ability of those embryos. Mitochondrial distribution was examined by fluorescence microscopy technology. Results indicated that: (1) The rate of mitochondrial ring formation around pronuclei in vitrified 2-PN embryos was significantly lower than in fresh ones (67.3 ± 3.0% vs. 84.9 ± 3.1%) (P < 0.05). (2) Blastocyst development rate of vitrified 2-PN embryos without mitochondrial rings (61.7 ± 4.5%) was significantly lower than that of vitrified embryos with mitochondrial rings (82.1 ± 2.8%). (3) Following staining by 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethyl-imidacarbo-cyanine iodide (JC-1), most red-colored mitochondria (high Δψ) were distributed peripherally around pronuclei and along cell membranes of fresh 2-PN embryos. Conversely, red-colored mitochondria were greatly diminished in vitrified embryos, with green mitochondria (low Δψ) evenly distributed throughout the cytoplasm. The proportion of fresh 2-PN embryos with obvious aggregation of high Δψ mitochondria (84.2 ± 2.2%) was significantly higher than that of vitrified embryos (26.7 ± 3.0%) (P < 0.05). (4) The proportion of fresh embryos with microtubules distributed around pronuclei (83.5 ± 3.4%) was similar to that of vitrified embryos (74.7 ± 2.5%). In conclusion, vitrification affected mitochondrial distribution and decreased the mitochondrial membrane potential in mouse 2-PN embryos, events which may affect subsequent developmental viability of such embryos. Mol. Reprod. Dev. 76: 1056–1063, 2009.