Jesus Avila is a Professor and former Director of the Centre of Molecular Biology in Madrid. Over the last thirty years, his work has focused on the cytoskeleton of the neuron. He is now investigating on the role of tau proteins in neurodegenerative disorders (tauopathies) such as Alzheimer’s disease. Professor Avila is a member of several professional organizations including EMBO, the European Academy, and the Spanish Royal Academy of Sciences. He is also on the Editorial Board of a number of scholarly journals and has more than 300 publications to his name. Jose J. Lucas is Research Professor of the Spanish National Research Council (CSIC) at the Centre for Molecular Biology (CBMSO) in Madrid. He obtained his PhD from the Cajal Institute in 1993 and then moved to Columbia University in New York for his postdoctoral training. He subsequently returned to Spain to join CBMSO and, in recent years, his research has focused on the generation of mouse models to study neurodegeneration and other CNS diseases. Professor Lucas is a member of several professional organizations including the European Huntington’s Disease Network (EHDN) and the Spanish Royal Academy of Pharmacy. He has authored more than 70 papers including contributions on the potential for reversibility of neurodegenerative disorders. FÚlix Hernßndez lectures on Biochemistry and Molecular Biology at Autonoma University in Madrid. His main research focus is on neurodegenerative diseases, such as Alzheimer’s and related tauopathies, using genetically modified mouse models – especially those over-expressing mutated tau protein and its main kinase, GSK-3?.

Animal Models for Neurodegenerative Disease

By Jesús Avila, Jose J. Lucas, Félix Hernández

The Royal Society of Chemistry

Copyright © 2011 Royal Society of Chemistry
All rights reserved.
ISBN: 978-1-84973-184-3

Contents

Chapter 1 The Contribution of Transgenic Models to the Understanding of Alzheimer’s Disease Progression and Therapeutic Development Meredith A. Chabrier, Kara M. Neely, Nicholas A. Castello and Frank M. LaFerla, 1,
Chapter 2 Animal Models of Amyloid/PS-1 Pathology Akihiko Takashima, 15,
Chapter 3 The Ying and Yang of the Reelin Signalling Pathway in Alzheimer’s Disease Pathology Eduardo Soriano, Daniela Rossi and Lluís Pujadas, 39,
Chapter 4 Transgenic Mice Overexpressing GSK-3β as Animal Models for Alzheimer’s Disease Félix Hernández, 52,
Chapter 5 Invertebrate and Vertebrate Models of Tauopathies Jürgen Götz, Lars M. Ittner, Naeman N. Götz, Hong Lam and Hannah R. Nicholas, 69,
Chapter 6 Animal Models of Parkinson’s Disease Hardy J. Rideout and Leonidas Stefanis, 86,
Chapter 7 Animal Models and the Pathogenesis of Parkinson’s Disease José G. Castaño, Teresa Iglesias and Justo G. de Yébenes, 113,
Chapter 8 Neuroprotection in Parkinson’s Disease Alberto Pascual, Javier Villadiego, María Hidalgo-Figueroa, Simón Méndez-Ferrer, Raquel Gómez-Díaz, Juan José Toledo-Aral and José Lopez-Barneo, 162,
Chapter 9 Animal Models for ALS Ritsuko Fujii and Toru Takumi, 177,
Chapter 10 Animal Models for Huntington’s Disease Zaira Ortega and José J. Lucas, 214,
Chapter 11 Mouse Models of Prion Protein Related Diseases María Gasset and Adriano Aguzzi, 230,
Chapter 12 Mouse Models of Ischemia David C. Henshall and Roger P. Simon, 251,
Chapter 13 A Non-transgenic Rat Model of Sporadic Alzheimer’s Disease Khalid Iqbal, Xiaochuan Wang, Julie Blanchard and Inge Grundke-Iqbal, 274,
Subject Index, 284,

CHAPTER 1

The Contribution of Transgenic Models to the Understanding of Alzheimer’s Disease Progression and Therapeutic Development

MEREDITH A. CHABRIER, KARA M. NEELY, NICHOLAS A. CASTELLO AND FRANK M. LAFERLA

Department of Neurobiology and Behavior, Institute of Memory Impairments and Neurological Disorders, University of California Irvine, Irvine, CA, USA

1.1 Introduction

Alzheimer disease (AD), the most prevalent neurodegenerative disorder, is characterized by progressive memory loss and cognitive decline. Currently, there are over 35 million people throughout the world who suffer from AD and the prevalence is expected to increase by more than 50% by the year 2030.

Over 100 rare mutations have been described in three AD-related genes that cause an autosomal dominant form of the disease, familial AD (fAD), which comprises less than 5% of all cases. The vast majority of AD cases are sporadic (sAD) and the causes underlying these cases remain unknown. Neuropathologically, AD is characterized by the accumulation of amyloid-beta (Aβ) plaques and neurofibrillary tangles, in addition to widespread synaptic loss, inflammation and oxidative damage, and neuronal death. Curiously, the neuropathological and clinical phenotype is indistinguishable in both types of the disease, with the major difference being the age of onset, which occurs at less than 65 in fAD.

As the cause of sAD is unknown, transgenic models have been based on the fAD component, utilizing fAD-associated genetic mutations to mimic specific elements of AD pathology, with the rationale that the events downstream of the initial trigger are quite similar. These genetic models have been invaluable in determining the molecular mechanisms of disease progression and for testing potential therapeutics. Although no single mouse model recapitulates all of the aspects of the disease spectrum, each model allows for in-depth analyses of one or two components of the disease, a feat not readily possible or ethical with human patients or samples.

1.2 Aβ and Tau Biology

Aβ is a central feature of AD pathology and is formed by the sequential cleavage of the amyloid precursor protein (APP) by β-secretase (BACE1) and γ-secretase, of which the catalytic component is either presenilin 1 or 2 (PS1 or PS2) (Figure 1.1A). The mutations found in fAD patients occur in APP, PS1 or PS2, and alter Aβ by either increasing its production, altering the ratio of Aβ42/Aβ40 or increasing its propensity to aggregate. Aβ42 has a higher propensity to aggregate and is considered the more pathological form. Although insoluble Aβ plaques are the most prominent pathological feature of AD, human evidence shows that the plaque burden does not correlate with cognitive decline6–10 and soluble forms of Aβ oligomers are now regarded as the primary pathological culprit, raising the possibility that plaques might actually be protective by sequestering the toxic oligomeric Aβ species. Importantly, this finding is mirrored in studies with transgenic mice, which provides us with an important tool for investigating the pathological effects of different Aβ assemblies.

Aβ oligomers are classified as either prefibrillar, which range in size from a small dimer to 75 kDa, or fibrillar which can assemble to a size greater than 500 kDa. The leading hypothesis for AD pathogenesis is the Aβ cascade hypothesis, which states that Aβ is the primary trigger that initiates a cascade of disease effects such as the formation of neurofibrillary tangles and chronic inflammation, eventually culminating in neurodegeneration and dementia.

Neurofibrillary tangles (NFTs) represent the second hallmark feature of AD pathology and consist of the microtubule binding protein tau. Tau is normally responsible for maintaining the structural integrity of cells, but in tauopathy disorders like AD, tau becomes aberrantly phosphorylated. Tau hyperphosphorylation renders it unable to bind to microtubules, causing their destablization and aggregation, eventually forming NFTs (Figure 1.1B). Transgenic models overexpressing mutant tau exhibit tangle formation, cognitive decline and neuronal loss.

1.3 Modelling Plaques and Tangles

Many transgenic models have successfully recapitulated amyloid pathology, generally by expressing mutated forms of APP and/or PS1. But models that develop both amyloid and tau pathology are rare, yet truly required for evaluation of the therapeutic efficacy of an anti-AD intervention. In 2003, our lab generated a triple-transgenic model of AD (3xTg-AD) that carries APPswe, tauP301L and PS1M146V mutant transgenes, which together promote the development of both Aβ and tau pathology. This model has allowed investigation of these two major pathological hallmarks within the same mouse, and most importantly, provided insight into the interaction between Aβ and tau (discussed below). The 3xTg-AD model also recapitulates other important features of AD, including age-dependent cognitive decline, chronic inflammation and neuronal loss.

It is important to note that the 3xTg-AD model, like all transgenic models of AD, does not replicate the initial cause of sporadic AD. Mutant forms of APP and PS1 are only carried by fAD patients, and the mutated forms of tau protein carried by many transgenic models, including the 3xTg-AD, are not found in AD patients, but instead are characteristic of frontotemporal dementia (FTD) with Parkinsonism linked to chromosome 17. However, the tau pathology produced by these mutations is highly similar to that found in AD and has allowed investigation of the interactions between Aβ and tau.

1.4 Mechanisms of Disease Progression

1.4.1 Aβ and Tau

Essential to our understanding of Aβ and tau interactions, studies with the 3xTg-AD model show that the accumulation of Aβ begins first and coincides with cognitive deficits, while tau pathology develops several months later. Furthermore, characterization of the parental 3xTg-AD showed a significant increase in hyperphosphorylated tau compared with a derivative line without the PS1 mutation, suggesting that higher Aβ42 levels in the 3xTg-AD directly facilitates tau pathology. Blocking Aβ accumulation by immunizing against Aβ also decreases tau pathology. The mechanism for Aβ dependent tau pathology involves alterations in CHIP, a tau ubiquitin ligase as increased Aβ causes decreased levels of CHIP concurrent with increased tau. Restoring CHIP levels ameliorated the Aβ-induced tau pathology. This supports the amyloid cascade hypothesis and demonstrates that reducing Aβ levels can reverse tau pathology; it also implicates the ubiquitin proteasome system in disease progression.

Proteasome impairment has been implicated as an important mechanism for protein aggregation in AD. There is in vitro and in vivo evidence that Aβ blocks proteasome activity. Given that tau is largely degraded by the proteasome, Aβ-mediated proteasome impairment leads to tau accumulation. Using the 3xTg-AD model, our lab has shown that tau clearance follows Aβ clearance after Aβ immunotherapy, and that tau clearance is dependent on proteasome activity. Tau clearance is inhibited when the proteasome activity is blocked. Furthermore we showed that Aβ oligomers, but not monomers, specifically target the proteasome, thereby facilitating tau accumulation. In additional support of this mechanism of disease progression, Uch-L1, an enzyme responsible for ubiquitin recycling, is downregulated in the AD brain. Increasing expression of Uch-L1 rescues synaptic and cognitive deficits in the APP/PS1 model. This provides significant evidence that drugs targeting the ubiquitin–proteasome system may be a successful approach to AD therapy.

1.4.2 Inflammation

The immune system is chronically active in AD, leading to glial activation and the induction of pro-inflammatory cytokines near Aβ deposits. Transgenic models have provided evidence that there are specific interactions between the inflammatory pathway, Aβ and tau that consequently lead to neuronal loss. Studies in the 3xTg-AD model have found that TNF-α and MCP-I expression are highly upregulated by three months of age, which corresponds with the time course of microglial activation. The same study showed that neurons are a significant source of TNF-α, and chronic TNF-α exposure leads to neuronal cell death in transgenic but not non-transgenic mice. Another study in APP transgenic models demonstrated that specific targeting of the CD40 ligand by genetic and immunological approaches lead to attenuation of Aβ pathology and a decrease in amyloidogenic processing. While CD40 may be a future target for AD therapy, the effects on tau pathology are unknown. Together, these studies suggest that neuronal loss is caused by the upregulation of inflammatory proteins due to AD related pathology and targeting the immune system can attenuate disease progression.

Another pathway by which Aβ facilitates the development of tau pathology is through the activation of the CNS inflammatory system. Induction of CNS inflammation in the 3xTg-AD model causes tau hyperphosphorylation, while not affecting APP levels. The augmented tau pathology is due to increased formation of p25, which heightens the activity of Cdk5, a known kinase for tau phosphorylation (Figure 1.1B). Additionally, in the P301S tauopathy mouse model which accumulates tau pathology alone, microglial activation precedes tangle formation and neuronal loss. Administration of an immunosuppressant, FK-506, attenuates tau pathology and increases the lifespan of this model. Further studies are needed to elucidate the precise mechanism of steps from microglia activation to tau pathology, as these could be possible therapeutic targets.

1.4.3 Risk Factors

As 95% of AD cases are of unknown origin, it is highly relevant to determine risk factors that may trigger or amplify disease pathology. There are many known risk factors of AD; here we discuss stress, diet, exercise, environmental factors and low cerebral blood oxygen levels.

Chronic stress exacerbates disease progression by increasing Aβ production and tau accumulation. Administration of dexamethasone, a potent glucocorticoid, to 3xTg-AD mice selectively increases APP and BACE (betasite APP cleaving enzyme), causing Aβ levels to increase, leading to an increase in tau. To further show that the tau increases were due to increased Aβ levels, a control mouse with only the tauP301L and PS1M146V transgenes, without APP, was created. Dexamethasone treatment in tauP301L/PS1M146V mice did not cause an increase in tau pathology, suggesting that the increased tau accumulation is due to downstream effect of Aβ. Further evidence from the 3xTg-AD model shows that corticosterone levels correlate with cognitive performance on stressful tasks such as Morris water maze and inhibitory avoidance. These studies indicate that both therapeutically targeting stress-related hormones and/or lifestyle changes to reduce stress may slow AD disease progression.

Exercise and diet are known to improve cognitive function, possibly by increasing neurogenesis and synaptic plasticity. Epidemiological studies show that physical activity inversely correlates with risk for dementia, and specifically that AD patients were less active during their lifetimes than non-demented subjects. To investigate the specific mechanisms that link exercise and cognition in AD, evidence from several different mouse models correlate voluntary exercise with enhanced learning and a decrease in amyloid burden. Studies in the Tg2576 model have shown that these changes occur by altering APP processing in as little as one month of exercise. Dietary supplementation with a variety of antioxidants, fatty acids or teas has also been shown to modulate Aβ and improve learning and memory in transgenic models. For example, three-month-old 3xTg-AD mice treated with DHA, an omega-3 polyunsaturated fatty acid, over a period of 12 months had significantly reduced intraneuronal accumulation of Aβ and tau. However, it is important to note that this study shows that DHA is preventative for disease progression, not therapeutic. DHA given to mild to moderate AD patients in clinical trials did not significantly improve cognition. Changes in diet should therefore be considered as a preventative measure to decrease risk for AD and not as way to ameliorate pathology.

Interestingly, hypoperfusion events are also linked to the onset and progression of AD pathology. Epidemiological data suggest that patients are 2–5 times more likely to develop AD following a stroke or other ischemic episode. A number of studies examining hypoperfusion insults in AD models have shown robust increases in AD pathology, in part mediated via upregulation of BACE. Surprisingly, even non-ischemic mild hypoperfusion insults produce significant increases in insoluble Aβ42 up to three weeks after the insult, as well as significantly altering tau phosphorylation in the 3xTg-AD model. These data indicate a strong role for hypoperfusion on the development of AD pathology.

1.5 Therapeutic Development in Transgenic Models

Animal models have been extremely important for the discovery and development of novel AD therapeutics, particularly where studies in humans would be unethical, cost-prohibitive or too time-consuming. Here we highlight several promising treatment strategies that have been intensely investigated in transgenic mice. One such strategy is immunotherapy, which uses antibodies to target AD pathology and prevent or reverse disease progression. A second strategy uses histone deacetylase (HDAC) inhibitors to enhance synaptic plasticity in order to compensate for disease-associated cognitive deficits. A third strategy uses acetylcholinesterase inhibitors to compensate for aberrant acetylcholine signalling, which is an early and prominent feature of AD.

1.5.1 Aβ Immunotherapy

Immunotherapy against Aβ, either by active or passive immunization, has been well established as an effective method to prevent Aβ aggregation. Early studies actively immunized PD-APP mice against Aβ and found a significant reduction in Aβ aggregation in both young, pre-pathological mice, as well as aged mice with established pathology. Further studies in PD-APP mice found that passive delivery of Aβ antibodies to the periphery is also effective for reducing soluble and insoluble Aβ, and in Tg2576 mice led to a reversal in cognitive deficits.

Studies from our lab provided insight into the effect of immunotherapy on both Aβ and tau pathology in the 3xTg-AD model. Administration of Aβ antibodies into the hippocampus lead to reductions in soluble and insoluble Aβ, as well as phosphorylated tau. This finding supports the amyloid cascade hypothesis and suggests that targeting Aβ alone may be sufficient to affect multiple aspects of AD pathology. Interestingly, hyperphosphorylated tau was unaffected by immunotherapy, indicating that pathology may become more resistant to antibody-mediated clearance as the disease progresses. In a subsequent study, we demonstrated that clearance of soluble Aβ alone was insufficient to improve cognition in 3xTg-AD mice. Concomitant reduction in soluble Aβ and tau was necessary to rescue memory impairments, which highlights the utility of a mouse that models both Aβ and tau pathology.

(Continues…)Excerpted from Animal Models for Neurodegenerative Disease by Jesús Avila, Jose J. Lucas, Félix Hernández. Copyright © 2011 Royal Society of Chemistry. Excerpted by permission of The Royal Society of Chemistry.
All rights reserved. No part of this excerpt may be reproduced or reprinted without permission in writing from the publisher.
Excerpts are provided by Dial-A-Book Inc. solely for the personal use of visitors to this web site.