**6. Animal models for AD study**

AD investigations have been conducted traditionally by studying human brains (autopsy) or by producing specific brain lesions in mice. The generation of animal models is particularly relevant, because they have been designed to test neurodegeneration with characteristics similar to those in the human brain, allowing us to design new therapeutic approaches. These models are key tools for in‐depth studies of neurodegenerative diseases like AD.

Many studies of AD are based on experimental models in mice since their genome is nearly 99% homologous with human [161]. Transgenic mouse models recapitulate the major hall‐ marks of AD and have been utilized since the early 1990s to explore in detail mechanisms underlying the disease pathology; they have provided excellent opportunities to analyze the bases for the temporal evolution of AD brains and to delineate the basic mechanisms that cause cellular dysfunction.

At present, there are many transgenic mouse and knockout models to analyze certain aspects of AD pathology, allowing the exploration of uncharted territories; they have revealed new pathogenic possibilities, many of which have not yet been demonstrated in humans. On the other hand, some discrepancies between the data obtained in the mice and in man remain unexplained [162]. Mice lack certain important aspects of AD; for example, age is an important factor in AD, but these animals have a short life, between 2 and 4 years. Also, the amyloid protein in mice, derived from proteolysis of the APP precursor, is different from that in human [163]. In spite of that, diverse studies in this mouse model showed the presence of soluble Aβ oligomers at prefibrillar stages that can act as toxic ligands at postsynaptic compartments, driving the synaptic in neuronal populations localized in similar areas to those affected in the human pathology with memory alterations. They have also been instrumental in validating dug targets in special cerebral areas to control memory.

The triple transgenic (3xTg‐AD) mouse, which develops pathologies associated with AD, was created in 2003 (**Figure 1**). To produce this model, Oddo's team simultaneously microinjected two genes (APP and tau) into single‐cell PS1M146V mouse embryos (transgenic mice that overexpress human or wild‐type APP, and are hybrids from the 129/C57BL6 strain). These mice develop both amyloid plaques and NFT‐like pathology in a progressive and age‐ dependent manner associated with anatomical and temporal analogously to that observed in the human AD brain [16]. In this 3xTg‐AD, Aβ deposits initiate in the cortex and progress to the hippocampus with aging (**Figure 2**). Amyloid accumulation is localized in the basal neocortex as well as in entorhinal areas, but this accumulation can also expand into the hippocampus. The conformational or hyperphosphorylation changes characteristic of tau pathology occur particularly in pyramidal neurons of the hippocampal CA1 subfield and in cortical structures (**Figure 3**) and evolve in the AD brain [164].

**Figure 1.** Triple transgenic mouse (3xTg‐AD).

The quantitative aspects of the hypothesis imply that reducing the number of Aβ‐plaques or the concentration of Aβ‐oligomers should be sufficient to halt progression of AD. Thus, a minor increase in the Aβ42:Aβ<sup>40</sup> ratio stabilizes toxic oligomeric species with intermediate conformations. The toxic impact of these Abfix species on the synapse but can spread into cells, producing neuronal death; Kuperstein et al. [156], suggest that there is a dynamic

In addition, it is well known that diffusible Aβ oligomers are the major toxic agents in AD, and both monomers and oligomers are important for the early diagnosis of dementia because they are potential predictors for the progression of AD and are useful to evaluate new drugs against

A quarter to a third of older people has amyloid burdens without symptoms of dementia [159]. Various APP transgenic mice do not have all the characteristics of AD: they exhibit little or no neuron loss and not all of them develop cognitive impairments, even if for three‐quarters of their lives they have deposits of amyloid, suggesting that Aβ alone is not sufficient. Thus, they

AD investigations have been conducted traditionally by studying human brains (autopsy) or by producing specific brain lesions in mice. The generation of animal models is particularly relevant, because they have been designed to test neurodegeneration with characteristics similar to those in the human brain, allowing us to design new therapeutic approaches. These

Many studies of AD are based on experimental models in mice since their genome is nearly 99% homologous with human [161]. Transgenic mouse models recapitulate the major hall‐ marks of AD and have been utilized since the early 1990s to explore in detail mechanisms underlying the disease pathology; they have provided excellent opportunities to analyze the bases for the temporal evolution of AD brains and to delineate the basic mechanisms that cause

At present, there are many transgenic mouse and knockout models to analyze certain aspects of AD pathology, allowing the exploration of uncharted territories; they have revealed new pathogenic possibilities, many of which have not yet been demonstrated in humans. On the other hand, some discrepancies between the data obtained in the mice and in man remain unexplained [162]. Mice lack certain important aspects of AD; for example, age is an important factor in AD, but these animals have a short life, between 2 and 4 years. Also, the amyloid protein in mice, derived from proteolysis of the APP precursor, is different from that in human [163]. In spite of that, diverse studies in this mouse model showed the presence of soluble Aβ oligomers at prefibrillar stages that can act as toxic ligands at postsynaptic compartments, driving the synaptic in neuronal populations localized in similar areas to those affected in the human pathology with memory alterations. They have also been instrumental in validating

models are key tools for in‐depth studies of neurodegenerative diseases like AD.

equilibrium between toxic and non‐toxic intermediates.

are a model of asymptomatic AD [159, 160].

dug targets in special cerebral areas to control memory.

**6. Animal models for AD study**

AD [157, 158].

206 Update on Dementia

cellular dysfunction.

**Figure 2.** Photomicrographs of the amyloid beta in triple transgenic mouse in the cerebral cortex of 11‐month‐old fe‐ male showing the staining for amyloid beta aggregates mice stained by immunohistochemistry using a BAM‐10 anti‐ body.

**Figure 3.** Photomicrographs of the cerebral cortex of an 11‐month‐old female mouse stained by immunohistochemistry using 499 tau antibody, showing the presence of human tau protein in two magnifications.

Anothercharacteristicofthe3xTg‐ADmouseisthatthebrainregionsseverelyaffected,including the hippocampus, entorhinal cortex, amygdala, neocortex, and some subcortical areas such as basal forebrain where the acetylcholine (Ach) neurotransmitter is altered in the brains of individuals with mild AD due to low choline acetyltransferase (ChAT) activity [165–167].

The 3xTg‐AD mouse has fewer ChAT‐immunopositive neurons in the Meynert nucleus (primary source of cholinergic neurons), as well as a reduced density of ChAT‐positive cholinergic fibers projecting to the primary motor cortex and the CA1 area of the hippocampus [168]. These cognitive dysfunctions are caused by massive loss of cholinergic neurons in the anterior basal brain, the area most vulnerable to the development of the pathological charac‐ teristics associated with AD. Alterations in cholinergic neurotransmission in the patients' neocortex and hippocampus are associated with the early stages of memory loss [168]. We also found a 50% reduction in nest‐building quality (a task controlled by the hippocampus), associated with a significant increase in damaged neurons in the CA1 hippocampal area (26%) compared to wild‐type mice [170]. The decreased ability to carry out activities of daily living (humans) or to perform nest building correctly (3xTg‐AD mice) are behavioral symptoms that can be studied and related to anatomical and morphological signs in the complex Alzheimer's disease syndrome.

#### **6.1. Sporadic models for Alzheimer's study**

A variety of animals can serve as experimental models of AD, which are valuable tools for the design of new therapeutic strategies and to explore some other aspects of the disease, as some specimens develop amyloid plaques in their brain and cognitive dysfunctions similar to those of AD. Like humans, dogs develop amyloid plaques in their brains with advancing age, and some specimens suffer sporadic cases of Alzheimer's disease, age‐related cognitive impair‐ ment with loss of short‐term memory or working memory, changes in behavior, irritability, incontinence, and orientation problems [171]. Sarasa cloned and sequenced the canine APP, finding it virtually identical to human APP, including the peptide sequence corresponding to β‐amyloid peptide. They analyzed the presence and distribution of amyloid plaques in the brains of healthy young and old dogs with severe cognitive dysfunction. With specific antibodies against AB40 and AB42, they found that the old demented animals had many amyloid and more mature plaques than older control dogs [163].

A nontransgenic rodent *Octodon degus*, which develops hallmarks of AD, could be a natural model to understand how sporadic AD, between 12 and 36 months of age, develop the accumulation of Aβ oligomers and phosphorylated tau proteins. Moreover, age‐ related changes in Aβ oligomers and tau phosphorylation levels are correlated with decreases in spatial and object recognition memory, postsynaptic function, and synaptic plasticity [172].

Sparks and Schreurs proposed studying AD in rabbits fed a diet rich in cholesterol and copper. These animals develop amyloid plaques in their brains and deficiencies in learning complex tasks. They exhibit increased immunoreactivity to amyloid β in neurons, the presence of extracellular plaques in the meninges, microgliosis, apoptosis, vascular activation of SOD, rupture of the blood‐brain barrier and elevated brain levels of cholesterol; these data provide strong support for the suggestion that copper is implicated in the accumulation of Abfix [173].

Alzheimer's disease is of special interest to neuroscientists, not only because it is the most common of the brain degenerations but also because it is a multifactorial disorder of unknown etiology. In addition, recent evidence supports the hypothesis that persistent chronic infections produce increased Aβ (amyloidosis) in brain, and may be mediated by a response of the innate immune system. This hypothesis may give an explanation of the common pathogenic mech‐ anisms and inflammatory gene polymorphisms involved in both AD and type 2 diabetes. In both diseases amyloidosis, that is, the accumulation of insoluble aggregates of fibrillar proteins, occurs in various organs and is often associated with bacterial infections [174]. Thus, the accumulation of intraneuronal amyloid‐β peptide (Aβ) appears to be an early event in AD, suggesting its important role in the neurodegenerative process of AD, because Aβ aggregates, particularly oligomers, may lead to synaptic dysfunction and neuronal loss, which are associated with memory and neural plasticity loss. Transgenic animal models are established to study the pathological role of intracellular Aβ and to screen for drugs against Aβ aggregation and associated toxicity, and they suggest that soluble, nonfibrillar Aβ oligomers may induce synaptic failure early in AD. Despite their undoubted value, the transgenic models rely on genetic manipulations that represent the inherited and familial but not the most abundant, sporadic form of AD [175].
