Figure 5

Quantification of commissural axon outgrowth defects after perturbation of MDGA2 function by RNAi and antibody injection. (A) Schematic representation of the areas used to measure fluorescence intensities in control, RNAi and antibody (Ab)-treated embryos. A, ipsilateral; B, floor plate; C, floor-plate exit; D, rostral longitudinal axis; E, caudal turn; F, no turn; G, 45° turn; H, -45° turn. Background fluorescence (A', B', C'...) at corresponding locations were subtracted from the values obtained at the different measurement sites (A, B, C....). The table shows the normalized intensities, with A_tot (A - A') of the control situation set as the reference value (100; for details see Materials and methods). Note that the fluorescent intensities (at sites B, C, D...) for all experimental conditions are adjusted by the same factor as calculated by the normalization at the site A_tot. (B) Data obtained by measurements of fluorescence intensity are presented as histograms to indicate the percentages of fluorescence intensity at particular locations. Values for control embryos were set to 100% at each of the analyzed sites. While in all analyzed conditions commissural axons were able to enter the floor plate (B, left panel), in MDGA2 RNAi-treated, as well as in embryos injected with the MDGA2 antibody fewer axons exited at the contralateral site (B, middle panel). The strongest difference between control and experimental animals was seen in commissural axon growth along the longitudinal axis, with only 15 to 30% of the axons extending rostrally after perturbation of MDGA2 function (B, right panel). While the effect on commissural axon stalling in the floor plate was not statistically significant, the lack of commissural axon growth along the longitudinal axis was highly significant (***P < 0.005; standard t-test). Error bars are given as standard errors (SEM).