LEE, W-K.; HARRISON, J.; GREENLEE, K.; BETZ, O.; WESTNEAT, M.; Argonne National Laboratory; Arizona State University; Arizona State University; University of Kiel, Germany; Field Museum of Natural History: Biomechanics of insect respiration and feeding using synchrotron x-ray imaging

The motions of internal structures of insects and other small animals during behaviors such as respiration and feeding have largely remained a mystery due to a lack of high-resolution imaging approaches at the micron scale. We addressed questions of respiratory function and feeding behavior using x-ray imaging. We used 25 keV x-rays from the Advanced Photon Source to obtain high-resolution phase-enhanced x-ray videos of beetles, grasshoppers and other insects while restrained but capable of motion of the head, abdomen and legs. We optimized beam energy, scintillator screen composition (for x-ray to visible light conversion), camera optics, and live animal preparation to enable long survivorship in the beam (hours) and natural behaviors including feeding. Imaging involved 2X and 5X microscope objectives on digital cameras with field of view ranging from 1×1 to 3×5 mm. Video was recorded at 60Hz (digital video) or with lesser image quality up to 1000Hz HSV. We observed natural active respiratory behaviors such as abdominal compression in grasshoppers and the recently discovered tracheal compression in the head of beetles, and simultaneously recorded CO2 emissions to assess correlation between air sac or tracheal compression and gas exchange. We also analyzed mouthpart kinematics during feeding in ground beetles to test hypotheses of mouthpart musculoskeletal mechanics and reveal coordination of multiple internal mouthparts. A wide range of biological applications of the APS x-ray source are now underway and additional users are invited.