Cerebral Cortex Advance Access published online on January 27, 2007
Cerebral Cortex, doi:10.1093/cercor/bhl164
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Nestin-CreER Mice Reveal DNA Synthesis by Nonapoptotic Neurons following Cerebral Ischemia–Hypoxia
1 Divisions of Developmental Biology and Pediatric Neurology, Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, 3333 Burnet Avenue, Cincinnati, OH 45229, USA, 2 Department of Neurobiology, Yale University School of Medicine, Kavli Institute for Neuroscience at Yale, New Haven, CT 06510, USA, 3 Physician Scientist Training Program, 4 Division of Molecular Cardiovascular Biology, Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, 3333 Burnet Avenue, Cincinnati, OH 45229, USA
Address correspondence to Dr Chia-Yi Kuan, MD, PhD, Division of Developmental Biology, Room 3464, Cincinnati Children's Hospital Research Foundation, 3333 Burnet Avenue, Cincinnati, OH 45229, USA. Email: alex.kuan{at}cchmc.org.
The standard method of detecting neurogenesis uses bromodeoxyuridine (BrdU) to label DNA synthesis followed by double labeling with neuronal markers. However, DNA synthesis may occur in events unrelated to neurogenesis including aneuploidy and abortive cell cycle reentry. Hence, it is important to confirm neurogenesis with methods other than BrdU incorporation. To this end, we have generated transgenic nestin-CreER mice that express tamoxifen-inducible Cre recombinase under the control of a nestin enhancer. When crossed with a ubiquitous Enhanced Green Fluorescent Protein (EGFP)-Cre-reporter line, the bitransgenic animals can reveal the nestin-positive progenitors and their progeny with EGFP after tamoxifen induction. This system has many applications including visualization of embryonic neural progenitors, detection of postnatally transformed radial glial cells, and labeling adult neural progenitors in the subventricular zone (SVZ). To examine the contribution of SVZ progenitors to cell replacement after stroke, tamoxifen-induced mice were challenged with focal ischemia or combined ischemia–hypoxia followed by BrdU injection. This analysis revealed only very few EGFP-positive cells outside the SVZ after focal ischemia but robust DNA synthesis by hippocampal neurons without immediate cell death following ischemia–hypoxia. These results suggest that the nestin-CreER system is a useful tool for detecting embryonic and adult neurogensis. They also confirm the existence of nonproliferative DNA synthesis by old neurons after experimental brain injury.
Key Words: BrdU • ischemia–hypoxia • lineage tracing • MCAO • neural stem cells • tamoxifen
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