Probing metabolism in mouse embryos using Raman spectroscopy and deuterium tags

Abstract

The use of deuterated compounds is an interesting opportunity to expand the capabilities of Raman spectroscopy to study metabolism in living cells. Different biological objects have different tolerances to different deuterated compounds, and their metabolic chains may differ. Here, we explore the potential of this approach to probe metabolism in early mouse embryos. We investigated the Raman spectra of mouse embryos at different developmental stages cultured with deuterated amino acids, phenylalanine-d8 and leucine-d10, glucose-d7, and D₂O. Embryos after in vitro culture with 20 % v/v D₂O demonstrate Raman peak at 2186 cm^-1 corresponding to newly synthesized proteins. Deuterated amino acids can slow down the development rate in 4-8 cell stage embryos, and deuterated glucose can be used at 2 mM concentration. For blastocyst, it was possible to achieve 75 % fraction of deuterated phenylalanine, when cultured with glucose, the maximal intensity ratio between CD and CH bands was 13.7 %. To demonstrate the capabilities of Raman spectroscopy reinforced by deuterium labeling, we investigated the short-term effect of cryopreservation and revealed that cryopreservation decreases the amount of saccharides in embryos and does not affect the activity of protein de novo synthesis.

Keywords: Amino acids; Cryopreservation; Deuterium labeling; Deuterium oxide; Diapause; Glucose; Mouse embryos; Raman spectroscopy.