Thy1-mitoDsRed2 tg mice in studies of motor neurons. A. E13 tg Hb9-eGFP mouse embryo section showing motor neurons below the sulcus limitans (SL). Scale bar = 40 μm. B. Spinal cords of E13 Hb9-eGFP tg mouse embryos were subjected to flow cytometry and sorted by side scatter (SSC) and forward scatter (FSC) into small cells/debris and large cells (P1, blue and red dots). C. P1 (large) cells were subjected to FACS and sorted by viability based on phycoerythrin annexin (PE-A) negativity and eGFP (FITC)-positivity resulting in P2 cells. D. Cultured P2 cells were eGFP-positive motor neurons. Scale bar = 15 μm. E, F. Mitochondria (F, arrows) in living Hb9-eGFP motor neurons visualized by transfection with Thy1-mitoDsRed2 construct. Scale bar = 7 μm. G, H. Double tg mice expressing Hb9-eGFP and Thy1-mitoDsRed2 were created to image mitochondria (arrows) directly in living motor neurons. Scale bar = 7 μm. I, J. Mitochondrial visualization (hatched arrows) in situ in spinal cord motor neurons of Thy1-mitoDsRed tg mice (I) and tg ALS mice expressing human mutant SOD1 and Thy1-mitoDsRed (J). Asterisks mark nucleus. MitoDsRed reveals directly the mitochondrial swelling (J, hatched arrows) and fragmentation (J, open arrow) in ALS mouse motor neurons, and magnitude of mitochondrial swelling (K). Scale bars = 2 μm (I) and 2.5 μm (J). K. Mitochondrial diameters in motor neurons of Thy1-mitoDsRed2 tg mice (control) and SOD1-G93A:Thy1-mitoDsRed2 tg mice. Values are mean ± SD. Mitochondrial diameters in motor neurons were increased 5-fold (*, p < 0.01, n = 6 mice/group).