ALTERED MITOGEN-ACTIVATED PROTEIN KINASE SIGNALING, TAU HYPERPHOSPHORYLATION AND MILD SPATIAL LEARNING DYSFUNCTION IN TRANSGENIC RATS EXPRESSING THE -AMYLOID PEPTIDE INTRACELLULARLY IN HIPPOCAMPAL AND CORTICAL NEURONS V. ECHEVERRIA, a A. DUCATENZEILER, a E. DOWD, a,d J. JÄNNE, e S. M. GRANT, a M. SZYF, a F. WANDOSELL, f J. AVILA, f H. GRIMM, g S. B. DUNNETT, d T. HARTMANN, g L. ALHONEN e AND A. C. CUELLO a,b,c * a Departments of Pharmacology and Therapeutics, McGill University, 3655 Promenade Sir-William-Osler, Montreal, Quebec, Canada H3G 1Y6 b Anatomy and Cell Biology, McGill University, Montreal, Canada H3G 1Y6 c Neurology and Neurosurgery, McGill University, Montreal, Canada H3G 1Y6 d Brain Repair Group, School of Biosciences, Cardiff University, Cardiff, Wales, UK e A. I. Virtanen Institute, University of Kuopio, Kuopio, Finland f Centro de Biologı ´a Molecular Severo Ochoa, CSIC-Universidad Autónoma de Madrid, Madrid, Spain g Center for Molecular Biology, University of Heidelberg, Heidelberg, Germany Abstract—The pathological significance of intracellular A accumulation in vivo is not yet fully understood. To address this, we have studied transgenic rats expressing Alzheimer’s- related transgenes that accumulate A intraneuronally in the cerebral and hippocampal cortices but do not develop extra- cellular amyloid plaques. In these rats, the presence of intra- neuronal A is sufficient to provoke up-regulation of the phosphorylated form of extracellular-regulated kinase (ERK) 2 and its enzymatic activity in the hippocampus while no changes were observed in the activity or phosphorylation status of other putative tau kinases such as p38, glycogen synthase kinase 3, and cycline-dependent kinase 5. The in- crease in active phospho-ERK2 was accompanied by in- creased levels of tau phosphorylation at S396 and S404 ERK2 sites and a decrease in the phosphorylation of the CREB kinase p90RSK. In a water maze paradigm, male transgenic rats displayed a mild spatial learning deficit relative to control littermates. Our results suggest that in the absence of plaques, intraneuronal accumulation of A peptide correlates with the initial steps in the tau-phosphorylation cascade, alterations in ERK2 signaling and impairment of higher CNS functions in male rats. © 2004 IBRO. Published by Elsevier Ltd. All rights reserved. Key words: -amyloid, transgenic rat, ERK/MAPK, tau phos- phorylation, Morris water maze. In genetically inherited cases of Alzheimer’s disease (AD), there is increased production of the -amyloid fragment (A) of the amyloid precursor protein (APP) and aggre- gated fibrils of this peptide form the core of the neuritic plaques which are widespread across the neocortex and hippocampus of all AD patients (for reviews see Walsh et al., 2002). Thus, strong evidence favors a central role for A in the AD disease process. The most established hy- pothesis of the pathophysiology of AD, the so-called amy- loid hypothesis, states that A plaques or A oligopoly- mers are somehow toxic to neurons, causing neuronal dysfunction, neurodegeneration and dementia (for review see Tanzi and Bertram, 2001). For this reason, the majority of studies have focused on the neurotoxic consequences of the aggregation of extracellular A fragments in AD and in AD animal models. However, there is no unequivocal correlation between plaque load and the degree of demen- tia (Hardy and Higgins, 1992) suggesting that other extra- cellular oligomeric forms of A other than the extracellular aggregated peptide could also play a role in AD pathology (for review see Walsh et al., 2002). Furthermore, there is evidence that A is generated and sequestered intracellu- larly in the early AD pathology (for reviews see Wilson et al., 1999; Hartmann, 1999; Echeverria and Cuello, 2002). A is derived by the sequential cleavage of APP in a number of sub-cellular organelles including the endoplas- mic reticulum, Golgi and endosomal/lysosomal system (Dickson et al., 1995). It has been shown that A is present intracellularly in Down syndrome preceding the appear- ance of amyloid plaques (Mori et al., 2002). A number of studies have also suggested that intracellular A accumu- lation would precede plaque formation in AD patients (D’Andrea et al., 2001; Takahashi et al., 2002) and a similar sequence has also been reported in transgenic (Tg) mice overexpressing AD-related genes (Gouras et al., 2000). Despite the mounting evidence for intracellular amyloid-induced cell dysfunction in vitro (Grant et al., 1999b; Glabe, 2001; Zhang et al., 2002), the pathological *Correspondence to: A. C. Cuello, Department of Pharmacology and Therapeutics, McGill University, 3655 Promenade Sir-William-Osler, Montreal, Quebec, Canada H3G 1Y6. Tel: +1-514-398-3618; fax: +1-514-398-8317. E-mail address: claudio.cuello@mcgill.ca (A. C. Cuello). Abbreviations: A, -amyloid; AD, Alzheimer’s disease; APP, amyloid  precursor protein; CDK5, cycline-dependent kinase 5; ERK, extracellular- regulated kinase; GSK3, glycogen synthase kinase 3; IR, immunoreac- tive; mAb, monoclonal antibody; MAPK, mitogen-activated protein kinase; NFT, neurofibrillary tangles; PBS, phosphate-buffered saline; PBST, 5% non-fat milk in phosphate-buffered saline, 0.2% Tween-20; PHF, paired helical filaments; PS1, Presenlin-1; SDS, sodium dodecyl sulfate; Tg, transgenic. Neuroscience 129 (2004) 583–592 0306-4522/04$30.00+0.00 © 2004 IBRO. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.neuroscience.2004.07.036 583