A 3D printable device for non-invasive longitudinal neuroimaging in adult zebrafish

Zebrafish are a powerful model for imaging studies of development and neurobiology. However, most studies have focused on developing zebrafish because of the technical and biological challenges of imaging adult stages. These include increased tissue opacity and illumination depth limitations, and difficulty maintaining life support and anesthesia in a 2-4 cm-long fish. There are currently limited tools for intravital imaging of the adult zebrafish brain. The ability to image the brain in the same individual repeatedly without physical damage would facilitate the use of zebrafish to study aging and neurodegenerative disease. We designed and applied a 3D printable device for non-invasive, repeatable multiphoton neural imaging of genetically non-pigmented adult zebrafish across multiple ages and sizes. Animals successfully recovered after multi-hour imaging sessions and could be imaged repeatedly over periods of weeks to months. We show the utility of this approach through imaging the dynamic cells of the neuroimmune system, as well as radial glial cells and oligodendrocytes. This technique could be widely used and beneficial for other cell-scale neuroimaging studies in adult fish.