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. 2019 Jul 8;6(4):ENEURO.0014-19.2019. doi: 10.1523/ENEURO.0014-19.2019

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Copyright © 2019 Hess et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license, which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed.

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Figure 3. — Age-related morphologic abnormalities of PLM neurons are less severe in long-lived daf-2 mutants. A, Overview of morphologic features quantified in PLM neurons. Inset shows enlarged rectangular region with quantifications indicated (neurite-outgrowth count and length, red; sharp bends, blue; beads not shown). Yellow structure is the crossing PVM neuron. B, Neurite outgrowth counts at days 1 and 8 of adulthood for WT and daf-2(e1370) mutants. WT animals showed a significant increase in the number of neurite outgrowths between day 1 and day 8 (p = 3.15e-04), and higher neurite outgrowth counts than daf-2(e1370) mutants at day 8 (p = 1.12e-03). For direct comparison of this data with previous studies, we replotted this data in percentage of PLM neurons with neurite outgrowths (Extended Data Fig. 3-1). C, A significant decrease in bead density (bead count divided by the total length of the neurite) is found for WT animals (p = 8.25e-03), but not for daf-2(e1370) mutants comparing day 1 to day 8, respectively. However, no significant difference between genotypes at day 1 or at day 8 was found. Error bars are 95% bootstrapped confidence interval of the mean, n is the number of PLM neurons scored. D, Both WT and daf-2(e1370) mutants showed the same density of sharp bends (bend count divided by the total length of the neurite) at day 1 of adulthood (p = 0.903), whereas at day 8 of adulthood, there is a significant increase in bend density (day 1 vs 8: WT, p = 5.87e-18 vs daf-2, p = 8.34e-06). However, daf-2(e1370) mutants seems to protect against age-related wrinkly appearance of neuronal processes (daf-2 vs WT, p = 3.85e-07). This is in contrast to daf-2(RNAi) shown in Extended Data Figure 3-2. Error bars are 95% bootstrapped confidence interval of the mean, n is the number of PLM neurons scored. Data were collected in three independent trials and pooled for the analysis shown in this figure (B–D). See Extended Data Figure 3-3 for different angle threshold settings and Extended Data Figure 3-4 the histogram of angles and corresponding cumulative distributions. The waviness caused by osmotic shrinkage of C. elegans looks different to the age-related sharp bends (Extended Data Fig. 3-5). ns = p > 0.05, ** = p < 0.01, *** = p < 0.001, **** = p < 0.0001. Figure Contributions: Max Hess made all the figures.