Cellular maps of gastrointestinal organs: getting the most from tissue clearing

The development of modern methods to induce optical transparency ("clearing") in biological tissues has enabled the three-dimensional (3D) reconstruction of intact organs at cellular resolution. New capabilities in visualization of rare cellular events, long-range interactions, and irregular structures will facilitate novel studies in the alimentary tract and gastrointestinal systems. The tubular geometry of the alimentary tract facilitates large-scale cellular reconstruction of cleared tissue without specialized microscopy setups. However, with the rapid pace of development of clearing agents and current relative paucity of research groups in the gastrointestinal field using these techniques, it can be daunting to incorporate tissue clearing into experimental workflows. Here, we give some advice and describe our own experience bringing tissue clearing and whole mount reconstruction into our laboratory's investigations. We present a brief overview of the chemical concepts that underpin tissue clearing, what sorts of questions whole mount imaging can answer, how to choose a clearing agent, an example of how to clear and image alimentary tissue, and what to do after obtaining the image. This short review will encourage other gastrointestinal researchers to consider how utilizing tissue clearing and creating 3D "maps" of tissue might deepen the impact of their studies.