Fig. 6: Functions and properties of the neuromuscular junction (NMJ) derived from the coculture of SOD1G85G and SOD1G85R motor neurons (MNs) with SOD1G85G skeletal muscles (SKM).
A 3D confocal images of immunocytochemistry (ICC) staining for the SKM marker TTN; neuronal marker NF; and NMJ markers SV2 and AChR. B–J Calculations of the volume of SKM (B), MN (C), and MN/SKM ratio (D). Analysis of AChR properties, including number (E), volume (F), intensity (G), and AChR/SKM ratio (H) in coculture NMJ models. Distribution analysis of AChRs depicting the ratio of AChR on the SKM surface (I) and distances from the SKM surface (J). K Bright-field intensity changes in different ROIs after CaCl2 induction in coculture NMJs. L Spontaneous and CaCl2 induced SKM contraction ratios in NMJs. B–E, H, I For each group, 6 images were analyzed from 3 independent experimental replicates, with each data point representing the mean value of a single replicate. F, G, J A total of 333 (SOD1G85G MN + SOD1G85G SKM) and 74 (SOD1G85R MN + SOD1G85G SKM) AChRs from 6 images were analyzed, with each dot representing one AChR. L In total, 20 (SOD1G85G MN + SOD1G85G SKM), 29 (SOD1G85R MN + SOD1G85G SKM) and 34 (SOD1G85G SKM) ROIs were analyzed, with each dot representing one ROI. All data were collected from 3 independent experimental replicates. The unpaired 2-tailed Student’s t-test was used in (B–J) and one-way analysis of variance with Tukey’s post hoc test was used in (L). “−” indicating inside, “0” indicating directly on the surface, and “+” indicating outside the SKM surface in (J). Scale bar: 20 μm in confocal 3D images; and 5 μm in 2D image slices.
