Recovery and Brain Reorganization after Stroke in Adult and Aged Rats Tiffanie M. Markus, PhD, 1,2 Shih-Yen Tsai, MD, PhD, 2 Melanie R. Bollnow, BS, 2 Robert G. Farrer, PhD, 2 Timothy E. O’Brien, PhD, 3 Diana R. Kindler-Baumann, PhD, 4 Martin Rausch, PhD, 4 Markus Rudin, PhD, 4 Christoph Wiessner, PhD, 4 Anis K. Mir, PhD, 4 Martin E. Schwab, PhD, 5,6 and Gwendolyn L. Kartje, MD, PhD 1,2,7,8 Stroke is a prevalent and devastating disorder, and no treatment is currently available to restore lost neuronal function after stroke. One unique therapy that improves recovery after stroke is neutralization of the neurite in- hibitory protein Nogo-A. Here, we show, in a clinically relevant model, improved functional recovery and brain reorganization in the aged and adult rat when delayed anti–Nogo-A therapy is given after ischemic injury. These results support the efficacy of Nogo-A neutralization as treatment for ischemic stroke, even in the aged animal and after a 1-week delay, and implicate neuronal plastic- ity from unlesioned areas of the central nervous system as a mechanism for recovery. Ann Neurol 2005;58:950 –953 Cerebrovascular disease, or stroke, is one of the leading causes of death and is the most common cause of adult disability with increasing prevalence in the aged popu- lation. 1 Neuronal loss due to stroke may result in per- manent deficits in sensory, language, and motor capa- bilities, leading to profound economic and emotional costs. Despite tremendous research efforts and success with experimental animal models for stroke recovery, no specific treatment has emerged for improving func- tional recovery in patients beyond the acute stage of stroke. In many clinical settings, it often is not possible to treat stroke patients soon after the event due to de- lays in seeking medical care, delays in diagnosis, and relevant serious comorbidities that preclude treatment. Another problem that may affect the outcome of ani- mal studies and their application to human outcomes after stroke is the common use of young adult animals in experimental studies, rather than the more appropri- ate use of aged animals. Studies have shown that strategies to improve neu- ronal plasticity, that is, the reorganization of neuronal circuits from undamaged central nervous system (CNS) areas, could lead to enhanced rehabilitative potential. 2,3 We have shown that a unique way to improve func- tional recovery and neuronal plasticity after focal isch- emic stroke in the young adult rat is through neutral- ization of the neurite growth inhibitor Nogo-A 4 with specific anti–Nogo-A antibodies (Abs) administered ei- ther immediately, 5 24 hours after stroke, 6 or 1 week after stroke. 7 To study a more clinically relevant model for ischemic stroke, in this study, we tested the efficacy of anti–Nogo-A Abs delivered 1 week after ischemic damage in aged animals. Materials and Methods This study has been approved by the Joint Institutional An- imal Care and Use Committee of Loyola University and Hines Veterans Affairs Hospital and the veterinary authori- ties of Kanton Basel-Stadt, Switzerland. Aged rats (25 months of age at the time of stroke) were first tested on the skilled forelimb reaching test, a sensorimo- tor task requiring fine digital manipulation and the integrity of sensory and motor neuronal pathways 8 (Fig 1A). Animals then underwent permanent middle cerebral artery occlusion resulting in a focal ischemic stroke that impaired the trained limb (see Fig 1B). One week after stroke, animals received intracerebroventricular administration of purified anti– Nogo-A Ab 7B12 (monoclonal mouse IgG), a purified con- trol mouse IgG Ab, or no treatment. Using Alzet mini- osmotic pumps, (Durect Corporation, Cupertino, CA) as described previously 6 (model 2ML2; 5l/hour), we infused 5mg of the Ab (2.5mg/ml) at a rate of 15g/hour for 2 weeks, and then removed the pumps. Reaching performance was analyzed using a repeated-measures analysis of variance. To examine whether reorganization of neural circuits oc- curred as a result of anti–Nogo-A therapy, we evaluated adult rats (age, 3– 4 months) using functional magnetic res- onance imaging (fMRI) 8 weeks after stroke and delayed anti–Nogo-A therapy. Experimental groups included stroke/ anti–Nogo-A Ab (7B12), stroke/control Ab, and normal rats. Each animal was anesthetized and artificially ventilated while fMRI experiments were performed using a PharmaScan 70/16 system (PharmaScan, Bruker, Germany) operating at 7 Tesla. For functional imaging, a two-dimensional rapid ac- quisition with relaxation (RARE) sequence with the follow- ing parameters was used: matrix (MTX) = 128  128; 3 slices; field of view = 40  40mm; slice thickness = 1mm; slice spacing = 1mm; repetition time (TR) = 2,350 milli- seconds; Echo time (TE) = 76.8 milliseconds; RARE fac- From the 1 Neuroscience & Aging Institute, Loyola University Chi- cago, Maywood; 2 Neurology and Research Service, Hines Veteran Affairs Hospital, Hines; 3 Department of Mathematics and Statistics, Statistical Consulting Center, Loyola University Chicago, Chicago, IL; 4 Novartis Pharma AG, Nervous System Research, Neuroregen- eration Unit, Basel, Switzerland; 5 Brain Research Institute Univer- sity of Zurich; 6 Department of Biology, Swiss Federal Institute of Technology, Zurich, Switzerland; 7 Department of Cell Biology, Neurobiology & Anatomy, Loyola University Chicago; and 8 De- partment of Neurology, Loyola University Medical Center, May- wood, IL. Received Feb 23, 2005, and in revised form Jul 28. Accepted for publication Aug 7, 2005. Published online Nov 28, 2005 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/ana.20676 Address correspondence to Dr Kartje, Neurology Service #127, Build- ing 1, Room F201, Hines VA Hospital, Fifth Avenue and Roosevelt Road, Hines, IL 60141. E-mail: wendy.kartje@med.va.gov 950 © 2005 American Neurological Association Published by Wiley-Liss, Inc., through Wiley Subscription Services