Muscular twitching during the latter half of embryogenesis introduces key imaging and analysis challenges. First, the embryo moves rapidly within the eggshell, causing motion blur unless imaging is rapid. DiSPIM largely addresses the imaging challenge due to its rapid acquisition. Second, the contorted nature of the animal makes image analysis difficult since the coordinate system shifts from frame to frame. This problem also confounds comparisons across embryos (and combination of data derived from many embryos) since each embryo moves differently within the eggshell.
We have recently addressed these latter challenges by developing software that untwists the worm (1). A set of markers is expressed in the animal that allows us tomodel the twisted volume in the raw diSPIM images. After the volume is modeled in the twisted state, it can be resampled and linearized – this process is shown in Fig. 1. We are currently using the untwisting procedure to annotate and combine data from many different embryos in the post-twitching regime, with the goal of building an atlas of all nuclear positions and neurite outgrowths in the late developing embryo.
Fig. 1, Worm untwisting. A. Twisted-up embryo. B. The same embryo after a lattice has been added, defining the
coarse 3D shape. C. The same embryo after the worm model has been refined. D. The embryo after untwisting.
1. Untwisting the Caenorhabditis elegans embryo. Christensen RP, Bokinsky A, Santella A, Wu Y, Marquina-Solis J, Guo M, Kovacevic I, Kumar A, Winter PW, Tashakkori N, McCreedy E, Liu H, McAuliffe M, Mohler W, Colón-Ramos DA, Bao Z, Shroff H. Elife. 2015 Dec 3;4. pii: e10070. doi: 10.7554/eLife.10070.