Fast volume-scanning light sheet microscopy reveals transient neuronal events

Abstract

Light sheet fluorescence microscopy offers considerable potential to the cellular neuroscience community as it makes it possible to image extensive areas of neuronal structures, such as axons or dendrites, with a low light budget, thereby minimizing phototoxicity. However, the shallow depth of a light sheet, which is critical for achieving high contrast, well resolved images, adds a significant challenge if fast functional imaging is also required, as multiple images need to be collected across several image planes. Consequently, fast functional imaging of neurons is typically restricted to a small tissue volume where part of the neuronal structure lies within the plane of a single image. Here we describe a method by which fast functional imaging can be achieved across a much larger tissue volume; a custom-built light sheet microscope is presented that includes a synchronized galvo mirror and electrically tunable lens, enabling high speed acquisition of images across a configurable depth. We assess the utility of this technique by acquiring fast functional Ca2+ imaging data across a neuron’s dendritic arbour in mammalian brain tissue.