Supplementary material for Lein et al. (1999) Proc. Natl. Acad. Sci. USA 96 (23), 13491-13495.
Fig. 4.
Specificity of subplate neuron ablation. (A) ISEL (in situ end labeling) of visual cortex at P3, 24 hr after kainic acid injection. Many cells in the subplate zone, both in the white matter (lower subplate) and at the base of the cortical plate (upper subplate) are ISEL labeled (red cells), indicating that they are dying. Note that at this early age, the bottom of the cortical plate is not layer 6, but the upper subplate, which will disappear in the ensuing weeks [Luskin, M. B. & Shatz, C. J. (1985) J. Neurosci. 5, 1062-1075; Chun, J. J. & Shatz, C. J. (1989) J. Comp. Neurol. 282, 555-569]. Sections are counterstained with TOPRO3 (blue; Molecular Probes). Circles of green fluorescent microspheres mark the two kainic acid injection sites. Posterior is to the right. (B) Magnification of the area boxed in A. Most ISEL labeling is confined to the upper subplate, although some labeled cells are present in deep layer 6. Only rarely are labeled cells seen in the upper cortical plate (layers 2-4). ISEL-treated control sections from neonatal mouse thymus (C), where many dying cells are expected, and liver (D), where few dying cells are expected. (E and F) Ablation of identified subplate neurons by kainic acid. Immunohistochemical visualization of subplate neurons labeled at their birthdate (E24) with BrdU in a normal P28 animal (E) and a P28 animal that received kainate injections into the subplate at P7 (F). Many BrdU-labeled cells are visible in the upper subplate in E, while virtually no labeled cells are present after kainate treatment in F. Numbers refer to cortical layers. SPU, upper subplate zone. [Bars = 1 mm (A) and 200 mm (B-F).]