Ependymal damage with rhodamine dyes. (A) Timecourse of ependymal death in the lateral ventricle after rhodamine dye injection demonstrated with digital subtraction. Damage to the ependyma was evident at 12 h and rapidly progressed by 24 h. (B) Histology at 24 h demonstrates swollen ependyma with numerous pyknotic profiles in injected, but not the contralateral, hemisphere. e, ependyma; lv, lateral ventricle; p, parenchyma. RHO fluorescence overlaid on brightfield hematoxylin images. (C) Low-power view of lateral ventricles 3 d after injection demonstrates halo of rhodamine-positive cells around injected ventricle (white arrow). The contralateral ventricle demonstrates labelled ependyma in the absence of damage. (D) By 3 d, near-complete loss of the ependyma was evident. This coincided with the appearance of dye-laden SVMs, black arrowheads. The ependyma remained intact in the contralateral hemisphere (right panels). e, ependyma; lv, lateral ventricle; p, parenchyma; RhoB, rhodamine beads. RHO fluorescence overlaid on brightfield hematoxylin image (RhoB) and photoconverted DiI counterstained with hematoxylin. (E) Periventricular reactive astrocytes (black arrows) visualized with nestin immunohistochemistry (IHC) at 3d post-injection at wall of injected ventricle (left), but not in the contralateral hemisphere (right). lv, lateral ventricle; sp, septum. (F) IHC for ciliated cell-specific foxj1 28d after dye injection demonstrates persistent loss of ependyma in injected hemispere (left). cc, corpus callosum, cp, caudate/putamen; sp, septum. (G) Equivalent volume control injection of GFP-reporter adenovirus demonstrates no ependymal damage 3 weeks after injection. e, ependyma; lv, lateral ventricle; p, parenchyma. GFP fluorescence overlaid on brightfield hematoxylin image.