Assessment of changes in zebrafish embryo biomechanical properties due to cold treatment using high-resolution Brillouin microscopy

The assessment of microscale biomechanical characteristics provides crucial information about the interplay between forces and tissue stiffness during development. These forces can regulate tissue dynamics and organogenesis. Many methods such as atomic force microscopy, optical tweezers, and optical coherence elastography have been utilized to extract mechanical parameters. However, these techniques often lack sufficient resolution or require external forces that involve contract, which can disrupt mechanosensitive cells and tissues. In this study, we have used a high-resolution Brillouin microscopy system enabling mapping of the stiffness of zebrafish embryos at cleavage periods 1 hour post fertilization (hpf), 1.75 hpf, and 2.25 hpf before and after a 1-hour cold treatment at 5°C. The axial and transverse resolution of Brillouin system was 2.3 and 1.8 μm, respectively, enabling cellular-level imaging of the zebrafish embryo mechanical properties. The results show that zebrafish embryo anomalies caused by the cold treatment were correlated to changes in the Brillouin frequency. The results show that the longitudinal modulus across the yolk has increased, and two various layers with different Brillouin frequency shifts at the blastomere are clearly observable. Furthermore, the results show an increasing stiffness at the interface of the blastomere and yolk regions, where the yolk syncytial layer (YSL) forms during early cleavage stages between the ninth and tenth cleavages, which is when the embryo has 512-1024 cells.