Figure 5.

The loss of BDNF expression correlates with an increased apoptotic cell death of mdDA neurons in the E12.5 Pitx3^GFP/GFP rostral VM. a–d, Triple immunostaining for cCasp3, GFP, and TH on representative midbrain coronal sections of Pitx3^+/GFP (a, b) and Pitx3^GFP/GFP (c, d) embryos at E12.5 revealed an increase of apoptotic Pitx3⁺ mdDA neurons [cCasp3⁺ (red) and GFP⁺ (green) double-labeled cells, white arrowheads) in the mutant VM. White arrows point at erythrocytes that were stained unspecifically for cCasp3. e, Quantification of cCasp3⁺/GFP⁺ double-labeled cells relative to the total amount of GFP⁺ (Pitx3⁺) cells in these sections showed a significant increase of apoptotic mdDA neurons in the rostral (cCasp3⁺/GFP⁺ cells: E12.5 Pitx3^+/GFP, 0.506 ± 0.047%; E12.5 Pitx3^GFP/GFP, 1.676 ± 0.185%; n = 3, mean ± SEM; ***p < 0.001 in the independent samples t test), but not caudal (cCasp3⁺/GFP⁺ cells: E12.5 Pitx3^+/GFP, 0.902 ± 0.068%; Pitx3^GFP/GFP, 1.056 ± 0.042%, n = 3, mean ± SEM), VM of the Pitx3^GFP/GFP embryos. Scale bar: (in d), 100 μm.