Beyond early development: observing zebrafish over 6 weeks with hybrid optical and optoacoustic imaging

Zebrafish animal models have traditionally been used in developmental biology studies but have recently become promising models of cancer, tissue regeneration and metabolic disorders, as well as efficient platforms for functional genomics and phenotype-based drug discovery. Most studies of zebrafish have examined only the embryonic or larval stages of development, yet many questions in developmental biology and biomedicine require analysis of adults, when zebrafish are large and opaque. Conventional microscopy methods are highly sensitive to light scattering and therefore cannot be applied to zebrafish older than a few weeks. We describe a novel multi-modality system that can observe zebrafish from the larval stage to adulthood. Using a hybrid platform for concurrent selective plane illumination microscopy (SPIM) and optoacoustic mesoscopy we show continuous imaging of fish growth over 47 days of development at a similar object size-to-resolution ratio. Using multiple wavelength illumination over the visible and short-wavelength infrared regions, we reveal that the optoacoustic method can follow GFP-based contrast used in SPIM, enabling molecular imaging interrogation in adult fish. Moreover we optoacoustically reveal many other features of zebrafish based on optical contrast not present in SPIM, including contrast from endogenous blood, water and lipids. The hybrid method presented can extend optical imaging to adult zebrafish employed as model systems for studying long-term processes in development, cancer, diabetes and other disorders.