Calpastatin levels affect calpain activation and calpain proteolytic activity in APP transgenic mouse model of Alzheimer’s disease Tali Vaisid a,1 , Nechama S. Kosower a, * , Aviva Katzav b , Joab Chapman b , Sivia Barnoy a,c a Department of Human Molecular Genetics and Biochemistry, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv 69978, Israel b Department of Physiology and Pharmacology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv 69978, Israel c Department of Nursing, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv 69978, Israel Received 9 January 2007; received in revised form 20 March 2007; accepted 10 April 2007 Available online 19 April 2007 Abstract The intracellular Ca 2+ -dependent protease calpain and the specific calpain endogenous inhibitor calpastatin are widely distributed, with the calpastatin/calpain ratio varying among tissues and species. Increased Ca 2+ and calpain activation have been implicated in Alzheimer’s disease (AD), with scant data available on calpastatin/calpain ratio in AD. Information is lacking on calpain activation and calpastatin levels in transgenic mice that exhibit AD-like pathology. We studied calpain and calpastatin in Tg2576 mice and in their wild type littermates (control mice). We found that in control mice calpastatin level varies among brain regions; it is significantly higher in the cerebellum than in the hippocampus, frontal and temporal cortex, whereas calpain levels are similar in all these regions. In the Tg2576 mice, calpain is activated, calpastatin is diminished, and calpain-dependent proteolysis is observed in brain regions affected in AD and in transgenic mice (especially hippocampus). In contrast, no differences are observed between the Tg2576 and the control mice in the cerebellum, which does not exhibit AD-like pathology. The results are consistent with the notion that a high level of calpastatin in the cerebellum renders the calpain in this brain region less liable to be activated; in the other brain parts, in which calpastatin is low, calpain is more easily activated in the presence of increased Ca 2+ , and in turn the activated calpain leads to further diminution in calpastatin (a known calpain substrate). The results indicate that calpastatin is an important factor in the regulation of calpain-induced protein degradation in the brains of the affected mice, and imply a role for calpastatin in attenuating AD pathology. Promoting calpastatin expression may be used to ameliorate some manifestations of AD. # 2007 Elsevier Ltd. All rights reserved. Keywords: Alzheimer’s disease; Mutant-APP transgenic mouse; Tg2576; Calpain; Calpastatin; Protein degradation 1. Introduction Alzheimer’s disease (AD) is a common neurodegenerative disorder, characterized by progressive mental deterioration. The neuropathologic alterations include the accumulation of amyloid-b-peptide (Ab), formed by abnormal proteolytic cleavage of the amyloid precursor protein (APP), extracellular neuritic plaques containing Ab, intracellular neurofibrillary tangles, composed mainly of the protein tau, and neuronal cell loss (Selkoe, 2001). Ab is considered to play a major role in the pathogenesis of AD, and is toxic to neurons (Glabe, 2005; Selkoe, 2001). Transgenic mice that exhibit accumulation of neuritic plaques and/or neurofibrillary tangles have been widely used as models for studying AD. The Tg2576 mouse model of AD (Hsiao et al., 1996) expresses the human APP double mutation found in a Swedish family with early onset AD (APP K670N, M671L). As these transgenic mice age, Ab is significantly increased, accompanied by the appearance of Ab-containing neuritic plaques in several brain regions. The plaques are similar to those found in AD. The Tg2576 mice also exhibit behavioral deficits (Hsiao et al., 1996; Lesne ´ et al., 2006). The appearance of age-dependent behavioral and pathological abnormalities in these mice have rendered them a suitable model for structural and biochemical alterations involved in the pathogenesis of AD, and for therapeutic trials (Gau et al., 2002; www.elsevier.com/locate/neuint Neurochemistry International 51 (2007) 391–397 * Corresponding author. Tel.: +972 3 640 9013; fax: +972 3 640 9900. E-mail address: nkosower@post.tau.ac.il (N.S. Kosower). 1 This work was performed in partial fulfillment of the requirements for a Ph.D. degree, Sackler Faculty of Medicine, Tel Aviv University. 0197-0186/$ – see front matter # 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.neuint.2007.04.004