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. 2019 Apr 24;13:152. doi: 10.3389/fncel.2019.00152

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Copyright © 2019 Zhang, Bonadiman, Ciorraga, Benitez and Garrido.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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No correlation between P2Y1 suppression-mediated decrease of ankyrinG density and AIS length. (A) Graph represents AIS length of each 7 DIV neuron nucleofected with scrambled sh (black dots) or shP2Y1 (red dots). Lines represent the mean ± SEM. (B) Graph represents the mean ± SEM of AIS start point, maximum intensity point and end point of AIS in 7 DIV scrambled shRNA or P2Y1 shRNA nucleofected neurons. (C) Normalized ankyrinG intensity within the respective AIS length of neurons represented in (A). (D) Normalized AnkyrinG/AIS length ratio for each 7 DIV scrambled shRNA or P2Y1 shRNA nucleofected neuron. (E) Graph represents AIS length of each 14 DIV neuron nucleofected with scrambled sh (black dots) or shP2Y1 (red dots). Lines represent the mean ± SEM. (F) Normalized ankyrinG intensity within the respective AIS length of neurons represented in (E). Adjacent graph represents the mean ± SEM. (G) Normalized AnkyrinG/AIS length ratio for each 14 DIV scrambled shRNA or P2Y1 shRNA nucleofected neuron. Adjacent graph represents the mean ± SEM. (H) Correlation graph of AIS length and ankyrinG intensity for each individual 14 DIV neuron nucleofected with scrambled (black dots) or P2Y1 (red dots) shRNAs. Values are the same represented in (E,F). Data were analyzed first with two normality tests (Shapiro-Wilk and Kolmogorov-Smirnov). All data sets did fit a normal distribution and a Pearson correlation test was used. Regression lines are shown for shscr (black) and shP2Y1 (red) neurons. The number of neurons, Pearson r coefficient and p values are indicated in (H). A positive correlation is observed in control neurons, while there is no significant correlation in neurons expressing shP2Y1 interference RNA. (I) Graph represents the mean ± SEM of AIS start point, maximum intensity point and end point of AIS in 14 DIV scrambled shRNA or P2Y1 shRNA nucleofected neurons. Note a significant proximal shift of the maximum intensity point. (J) AnkyrinG profile along AIS length of 14 DIV neurons, indicating the proximal shift of the maximum ankyrinG intensity. Data are represented as the mean ± SEM of ankyrinG intensity every 1 micron. *p < 0.05, **p < 0.01, ***p < 0.001, n.s., non-significant.