Optical Imaging of Metabolic Dynamics in Animals

Abstract

Direct visualization of metabolic dynamics in living tissues with high spatial and temporal resolution is essential to understanding many biological processes. Here we introduce a platform that combines deuterium oxide (D2O) probing with stimulated Raman scattering microscopy (DO-SRS) to image in situ metabolic activities. Enzymatic incorporation of D2O-derived deuterium into macromolecules generates carbondeuterium (C-D) bonds, which track biosynthesis in tissues and can be imaged by SRS in situ. Within the broad vibrational spectra of C-D bonds, we discovered lipid-, protein-, and DNA-specific Raman shifts and developed spectral unmixing methods to obtain C-D signals with macromolecular selectivity. DO-SRS enabled us to probe de novo lipogenesis in animals, image protein biosynthesis without tissue bias, and simultaneously visualize lipid and protein metabolism and reveal their different dynamics. DO-SRS, being noninvasive, universally applicable, and cost-effective, can be adapted to a broad range of biological systems to study development, tissue homeostasis, aging, and tumor heterogeneity. Results SRS imaging enables D2O to be a contrast agent for metabolic activitiesWater (H2O), the ubiquitous solvent of life, diffuses freely across cell and organelle membranes and participates in the vast majority of biochemical reactions. As an isotopologue of water, heavy water (D2O) can rapidly equilibrate with total body water in all cells within an organism and label cellular biomolecules with deuterium (D) by forming a variety of X-D bonds through non-enzymatic H/D exchange and enzymatic incorporation ( Figure 1A). The former is spontaneous and reversibly forms oxygen-deuterium (O-D), nitrogen-deuterium (N-D), and sulfur-deuterium (S-D) bonds on existing molecules, whereas the latter depends on enzyme-catalyzed chemical transformation that irreversibly breaks the O-D bond and forms C-D bonds on newly synthesized molecules 5 . Through such transformation, deuterium quickly labels the metabolic precursors, such as non-essential amino acids (NEAAs), acetyl-CoA, and deoxyribose, which are then slowly incorporated into proteins, lipids, and DNA,