  Selective plane illumination microscopy (SPIM), or light sheet, is a powerful
three-dimensional imaging approach. However, access to and interfacing
microscopes with microfluidics have remained challenging. Complex interfacing
with microfluidics has limited the SPIM's utility in studying the hydrodynamics
of freely moving multicellular organisms. We developed SPIM-Flow, an
inexpensive light sheet platform that enables easy integration with
microfluidics. We used SPIM-Flow to study the hydrodynamics of freely moving
Hydra polyps in millimeter-sized chambers (4 mm wide, 1.5 mm height). Our
initial experiments across multiple animals, feeding on a chip (Artemia
franciscana nauplius used as food), and baseline behaviors (eg., tentacle
swaying, elongation, and bending) indicated animals' health inside the system.
SPIM enabled easy imaging of the freely moving animal and tracer beads (for
fluid visualizations) inside the larger chambers. Next, using the chambers, we
investigated Hydra's response to flow. Results suggest that animals responded
to established flow by bending and swaying their tentacles in the flow
direction. Finally, we used a previously described video analysis software
(FlowTrace) to visualize path lines generated by (e.g., vortex generated) and
around (e.g., due to flow) Hydra. These results demonstrated the SPIM-Flow's
utility to study the hydrodynamics of freely moving animals.
