Fig. 3. Conditional deletion of v integrin in neural cells leads to cerebral hemorrhage. (A-F) The hGFAP-Cre transgene is expressed in central nervous system radial glial cells and post-natal astrocytes. (A,B) Sagittal sections through E15 embryonic neocortex immunostained with anti-Cre (green) and anti-brain lipid-binding protein (BLBP) (red, A) or anti-glutamate transporter (GLAST) (red, B). Expression of Cre is seen in radial glial cells within the embryonic neocortex (arrows). Regions highlighted within the dashed boxes are shown as higher magnification insets. (C) Cre is not expressed by embryonic neurons. E15 sagittal brain sections were immunostained with anti-Cre (green) and anti-ß-tubulin III (red). Cre expression is absent in ß-tubulin III-positive neurons (arrowheads in C). v, ventricle. (D,E) Postnatal astrocytes in the cerebellum (D) and cerebral cortex (E) express Cre. Sagittal sections from postnatal (P7) brains immunostained with anti-Cre (green) and anti-GFAP (red). There is co-localization of GFAP and Cre in cortical astrocytes in the cerebral cortex (arrows in E) as well as in Bergmann glia of the cerebellum (arrows in D). Boxed areas are shown as higher magnification insets. (F) Cultured astrocytes from hGFAP-Cre transgenic mice show mosaic Cre expression. There is co-localization of Cre and GFAP in most cells (arrows); however, some GFAP-positive cells show no detectable Cre protein expression (arrowhead). (G) Strategy for generating the conditional v mutant allele in central nervous system neural cells, particularly glia. (H) hGFAP-Cre-mediated recombination was monitored by PCR using DNA isolated from tails (T) or from mutant cultured astrocytes (A). DNA isolated from astrocytes shows reduced intensity of the 350 bp band, owing to recombination of the v-flox allele. We confirmed deletion of exon four using a primer pair that detects the deleted v-flox cassette (data not shown). (I) Cortical astrocyte lysates were immunoblotted with anti-v antibody. There is a significant reduction in v protein expression in mutants. (J,K) Brains dissected from control (J) and mutant (K) P5 neonates. Arrows in K indicate microhemorrhage in the mutant cerebral cortex. (L,M) Coronal sections from control (L) and mutant (M) mice indicate focal regions of hemorrhage in mutant cerebral cortex (arrows in M). Boxed area shown at higher magnification (inset in M). (N-Q) Gross analysis of adult brains from control and mutant mice. Adult control (N) or mutant (O) brains do not display overt signs of cerebral microhemorrhage. Hematoxylin and Eosin stained coronal sections from adult control (P) and mutant (Q) cerebral cortex reveal normal cytoarchitecture and no microscopic hemorrhage.