Effect of low temperatures on cytochrome photoresponse in mouse embryos

Embryos cryopreservation is a widely used technology for genetic resources storage. Cryopreservation suppresses cell respiration, but very little is known about the changes that occur with mitochondria at low temperatures. We used Raman spectroscopy to investigate photoresponse and redox state of cytochromes in the respiratory electron transport chain (ETC) in early mouse embryos during cooling. Redox state of cytochromes was probed by the intensity of cytochrome resonance Raman lines. Photoinduced reactions of cytochromes were used to study the changes in the rates of redox reactions. It is found that the rate of cytochrome photoresponse detected by Raman spectra abruptly changes when embryos are cooled below -50 °C. Raman mapping revealed that the average intensity of cytochrome Raman peaks at -65 °C is higher than at -40 °C. Cytochrome b reduction was found in embryos frozen below -50 °C when irradiated with 532 nm laser radiation. These effects were observed for cells frozen in aqueous solutions of two different cryoprotectants: glycerol and propylene glycol. Raman spectroscopy of cytochromes reveals the abrupt changes in the ETC work of frozen mouse embryos at temperatures below -50 °C. We suggest that similar phenomena can be found in various cell types.