Aberrant motor corticospinal axons in PLXA3 and PLXA4 double mutants are located in the ipsilateral, ventrolateral spinal cord. All panels represent cross sections of the pyramidal decussation and spinal cord. Unilateral BDA motor CST axon tracing was performed in (a-d"). (a, b) Crossing of motor CST axons at the pyramidal decussation (Pyr Dec; black arrow in (a)) and into the dorsal funiculus of the spinal cord (SpC; black arrow in (b)) of P45 WT mice. (c, d) Normal crossed (black arrows) and aberrant uncrossed (red arrows) motor CST axons at the pyramidal decussation and spinal cord of P45 PLXA3/PLXA4-/- mice. (a'-d") High power views of arrowed areas in (a-d). In the upper cervical spinal cord, many mutant axons branched out from the uncrossed CST (green arrowheads in (d")). Some of these axons crossed to the contralateral gray matter (green arrowheads in (d')). (e-h") CamKIIα immunohistochemistry of the pyramidal decussation and spinal cord of P25 WT and P25 PLXA3/PLXA4-/- mice. Crossed (black arrowheads) and uncrossed (red arrows) CamKIIα-immunolabeled CST axons are observed at the pyramidal decussation and spinal cord in PLXA3/PLXA4-/-'s. High power views of the spinal cord in (f, h) are shown in (f', h', h"). Black dashed lines in (f', h') outline positive CamKIIα immunostaining of the dorsal CST in the dorsal funiculus of the spinal cord. (i) Comparison of percentages of 4- to 6-week old WT, PLXA3-/-, PLXA4-/-, and PLXA3/PLXA4-/- mice with an abnormal ventral CST apparent with BDA tracing. Numbers in parentheses indicate the number of mice analyzed. (j) Average normalized areas (see Materials and methods) of CamKIIα-labeled dorsal CST axons in WT, PLXA3-/-, PLXA4-/-, and PLXA3/PLXA4-/- mice. The dorsal CST area in each animal (mean ± standard error of the mean) is indicated by a black circle. The overall average dorsal CST areas (black lines) are decreased in the cervical spinal cords of PLXA3-/- (n = 6 mice), PLXA4-/- (n = 5 mice), and PLXA3/PLXA4-/- (n = 8 mice) versus WT (n = 6 mice) mice. **p < 0.01, Student's t-test. cc, central canal; DF, dorsal funiculus. Scale bars: 500 μm (a-h); 100 μm (a'-d", f'-h").