Intracellular Abeta and cognitive deficits precede beta-amyloid deposition in transgenic arcAbeta mice

The brain pathology of Alzheimer's disease is characterized by abnormally aggregated Abeta in extracellular beta-amyloid plaques and along blood vessel walls, but the relation to intracellular Abeta remains unclear. To address the role of intracellular Abeta deposition in vivo, we expressed human APP with the combined Swedish and Arctic mutations in mice (arcAbeta mice). Intracellular punctate deposits of Abeta occurred concomitantly with robust cognitive impairments at the age of 6 months before the onset of beta-amyloid plaque formation and cerebral beta-amyloid angiopathy. beta-Amyloid plaques from arcAbeta mice had distinct dense-core morphologies with blood vessels appearing as seeding origins, suggesting reduced clearance of Abeta across blood vessels in arcAbeta mice. The co-incidence of intracellular Abeta deposits with behavioral deficits support an early role of intracellular Abeta in the pathophysiological cascade leading to beta-amyloid formation and functional impairment.     

Genotype-related changes of ganglioside composition in brain regions of transgenic mouse models of Alzheimer's disease

In this study, brain gangliosides of different transgenic mouse models of Alzheimer's disease (AD) were analyzed and compared with age-matched wild-type mice. Gangliosides were analyzed in cerebral cortex, a region with extensive Aβ plaques, and cerebellum, a non-vulnerable region with no Aβ containing plaques. There was a marked increase in simple gangliosides GM2 and GM3 only within the cortex of all mice expressing APP^SL. Additionally, loss of complex “a” gangliosides (GT1a, GD1a and GM1) was recorded in APP/PS1Ki model, whereas in APP^SL and APP/PS1 mice, the complex “b” gangliosides (GQ1b, GT1b and GD1b) moderately decreased. Surprisingly, expression of either mutant PS1^M146L or PS1 mutant FAD (Ki model) alone tended to lower the levels of both GM2 and GM3 within the cortex. Conversely, only slight changes of the ganglioside pattern were found in the cerebellum. Because ganglioside alterations occurring in APP transgenic mice were similar to those observed in human AD brain, these transgenic models would represent valuable tools to further investigate the role of altered ganglioside metabolism in the pathogenesis of AD.     

Overexpression of monocyte chemotactic protein-1/CCL2 in beta-amyloid precursor protein transgenic mice show accelerated diffuse beta-amyloid deposition

Microglia accumulation at the site of amyloid plaques is a strong indication that microglia play a major role in Alzheimer's disease pathogenesis. However, how microglia affect amyloid-beta peptide (Abeta) deposition remains poorly understood. To address this question, we developed a novel bigenic mouse that overexpresses both amyloid precursor protein (APP) and monocyte chemotactic protein-1 (MCP-1; CCL2 in systematic nomenclature). CCL2 expression, driven by the glial fibrillary acidic protein promoter, induced mononuclear phagocyte (MP; monocyte-derived macrophage and microglial) accumulation in the brain. When APP/CCL2 transgenic mice were compared to APP mice, a fivefold increase in Abeta deposition was present despite increased MP accumulation around hippocampal and cortical amyloid plaques. Levels of full-length APP, its C-terminal fragment, and Abeta-degrading enzymes (insulin-degrading enzyme and neprilysin) in APP/CCL2 and APP mice were indistinguishable. Sodium dodecyl sulfate-insoluble Abeta (an indicator of fibrillar Abeta) was increased in APP/CCL2 mice at 5 months of age. Apolipoprotein E, which enhances Abeta deposition, was also increased (2.2-fold) in aged APP/CCL2 as compared to APP mice. We propose that although CCL2 stimulates MP accumulation, it increases Abeta deposition by reducing Abeta clearance through increased apolipoprotein E expression. Understanding the mechanisms underlying these events could be used to modulate microglial function in Alzheimer's disease and positively affect disease outcomes.     

ApoE epsilon3-haplotype modulates Alzheimer beta-amyloid deposition in the brain

ApoE epsilon4 allele increases the risk of late-onset Alzheimer disease (AD) as well as the amount of beta-amyloid deposition in the brain. Because half of AD patients do not have ApoE epsilon4, it is important to search for other determinants of ApoE that modify AD risk. We tested whether the haplotype background of the most common ApoE allele, epsilon3, influences brain amyloid deposition or the risk of neuropathologically verified AD in a population-based sample of elderly Finns. To exclude the effects of ApoE protein polymorphism we focused these analyses on subjects homozygous for epsilon3. Haplotypes were defined using polymorphisms at positions - 491 and -219 of the ApoE promoter and at position +113 of intron-1. We found that epsilon3-haplotypes containing the promoter allele -219T were associated with reduced amyloid deposition and reduced risk of neuropathologically verified AD as compared to epsilon3-haplotypes containing -219G. The functional polymorphism(s) responsible for the haplotypic difference remains to be identified. These results indicate that there is significant allelic variation in the ApoE gene region, which modulates brain amyloid deposition and AD risk, independent of the ApoE protein polymorphism.     

Alterations of brain circuits in Down syndrome murine models

Trisomy 21, also referred to Down syndrome (DS), is the most common genetic cause of mental retardation, affecting 1 each 800-1000 newborn children all over the world. DS is a complex disease, determined by an extra copy of human chromosome 21 that causes an imbalanced gene dose effect. The syntenies that exist between mouse chromosomes 10, 16, and 17 and human chromosome 21 offer the opportunity for a genotype-phenotype correlation and several mouse models of DS have been developed to improve our knowledge about cognitive disabilities and brain alterations. We present here the different murine models available up to now and we discuss the neural alterations that have been described in these strains. The largest amount of studies involved the so called Ts65Dn mouse showing early alterations of nitrergic, noradrenergic and cholinergic systems at the level of the basal forebrain. Neurogenesis and spine formations are decreased in the hippocampus, as well as the whole size of the cerebellum and the number of granule cells.     

Genetic background regulates beta-amyloid precursor protein processing and beta-amyloid deposition in the mouse

Alzheimer's disease (AD) is a multigenic neurodegenerative disorder characterized by distinct neuropathological hallmarks including deposits of the beta-amyloid (A beta) peptide. A beta is a 39- to 43-amino acid peptide derived from the proteolytic processing of the amyloid precursor protein (APP). While increasing evidence suggests that altered APP processing and A beta metabolism is a common feature of AD, the relationship between the levels of A beta and various APP products and the onset of AD remains unclear. We have undertaken a screen to characterize genetic factors that modify APP processing, A beta metabolism and A beta deposition in a genomic-based yeast artificial chromosome (YAC) transgenic mouse model of AD. A mutant human APP YAC transgene was transferred to three inbred mouse strains. Despite similar levels of holo-APP expression in the congenic strains, the levels of APP C-terminal fragments as well as brain and plasma A beta in young animals varied by genetic background. Furthermore, we demonstrate that age-dependent A beta deposition in the APP YAC transgenic model is dramatically altered depending on the congenic strain examined. These studies demonstrate that APP processing, A beta metabolism and A beta deposition are regulated by genetic background and that analysis of these phenotypes in mice should provide new insights into the factors that regulate AD pathogenesis.     

Phagocytosis and deposition of vascular beta-amyloid in rat brains injected with Alzheimer beta-amyloid.

The presence of extracellular deposits of beta-amyloid protein in the brain is a hallmark of Alzheimer's disease (AD). In an effort to determine the effect of amyloid in an animal model, the authors injected amyloid cores isolated from AD brains into the cortex and hippocampus of rats. Lipofuscin, a major contaminant of the plaque core preparation, was injected on the contralateral side and used as a control to induce an analogous phagocytic cell response. Rats were sacrificed 2 days, 7 days, and 1 month after injection and amyloid located by four histochemical techniques. Amyloid and lipofuscin move from the site of injection into otherwise undamaged neuropil, persist for at least 1 month and are both associated with increases in glial fibrillary acidic protein and microglia (OX-42) staining. By 1 week, many of the amyloid cores are ingested by phagocytes. Some of the beta-amyloid-containing phagocytes migrate to the vessels and to the ventricles, and by 1 month, a significant amount of the amyloid is directly associated with the vessels. This suggests that phagocytic cells can internalize exogenous amyloid and attempt to clear it from the central nervous system (CNS). Therefore, the observed distribution of amyloid is not necessarily the initial site of deposition.     

Caffeine protects Alzheimer's mice against cognitive impairment and reduces brain beta-amyloid production

A recent epidemiological study suggested that higher caffeine intake over decades reduces the risk of Alzheimer's disease (AD). The present study sought to determine any long-term protective effects of dietary caffeine intake in a controlled longitudinal study involving AD transgenic mice. Caffeine (an adenosine receptor antagonist) was added to the drinking water of amyloid precursor protein, Swedish mutation (APPsw) transgenic (Tg) mice between 4 and 9 months of age, with behavioral testing done during the final 6 weeks of treatment. The average daily intake of caffeine per mouse (1.5 mg) was the human equivalent of 500 mg caffeine, the amount typically found in five cups of coffee per day. Across multiple cognitive tasks of spatial learning/reference memory, working memory, and recognition/identification, Tg mice given caffeine performed significantly better than Tg control mice and similar to non-transgenic controls. In both behaviorally-tested and aged Tg mice, long-term caffeine administration resulted in lower hippocampal beta-amyloid (Abeta) levels. Expression of both Presenilin 1 (PS1) and beta-secretase (BACE) was reduced in caffeine-treated Tg mice, indicating decreased Abeta production as a likely mechanism of caffeine's cognitive protection. The ability of caffeine to reduce Abeta production was confirmed in SweAPP N2a neuronal cultures, wherein concentration-dependent decreases in both Abeta1-40 and Abeta1-42 were observed. Although adenosine A(1) or A(2A) receptor densities in cortex or hippocampus were not affected by caffeine treatment, brain adenosine levels in Tg mice were restored back to normal by dietary caffeine and could be involved in the cognitive protection provided by caffeine. Our data demonstrate that moderate daily intake of caffeine may delay or reduce the risk of AD.     

Regional brain cytochrome oxidase activity in beta-amyloid precursor protein transgenic mice with the Swedish mutation

Cytochrome oxidase activity was examined in a transgenic mouse model of Alzheimer's disease with overexpression of the 751 amino acid isoform of beta-amyloid precursor protein with the Swedish mutation under control of the murine thy-1 promoter. The neuritic plaques, abundantly localized in the hippocampus and anterior neocortical areas, showed a core devoid of enzymatic activity surrounded by higher cytochrome oxidase activity at the sites of the dystrophic neurites and activated glial cells. Quantitative measures, taken only in the healthy-appearing regional areas without neuritic plaques, were higher in numerous limbic and non-limbic regions of transgenic mice in comparison with controls. Enzymatic activity was higher in the dentate gyrus and CA2-CA3 region of the hippocampus, the anterior cingulate and primary visual cortex, two olfactory structures, the ventral part of the neostriatum, the parafascicularis nucleus of the thalamus, and the subthalamic nucleus. Brainstem regions anatomically related with altered forebrain regions were more heavily labeled as well, including the substantia nigra, the periaqueductal gray, the superior colliculus, the medial raphe, the locus coeruleus and the adjacent parabrachial nucleus, as well as the pontine nuclei, red nucleus, and trigeminal motor nucleus. Functional brain organization is discussed in the context of Alzheimer's disease. Although hypometabolism is generally observed in this pathology, the increased cytochrome oxidase activity obtained in these transgenic mice can be the result of a functional compensation on the surviving neurons, or of an early mitochondrial alteration related to increased oxidative damage.     

Quantitative histological analysis of amyloid deposition in Alzheimer's double transgenic mouse brain

The development of transgenic mice has created new opportunities for the generation of animal models of human neurodegenerative diseases where previously there was no animal counterpart. The first successful transgenic mouse model of Alzheimer's disease expressed increased levels of mutant human amyloid precursor protein, exhibiting neuritic-type amyloid deposits and behavioral deficits at six to nine months of age. More recently, it was shown that transgenic mice expressing both mutant human amyloid precursor protein and presenilin 1 exhibit neuritic-type amyloid deposits and behavioral deficits in as little as 12 weeks. This accelerated Alzheimer phenotype greatly reduces the time necessary to conduct preclinical drug trials, as well as animal housing costs. The purpose of this study was to quantify the deposition of amyloid in five regions of the cortex and two regions of the hippocampus of transgenic mice expressing amyloid precursor protein (K670N, M671L) and presenilin 1 (M146L) mutations at various ages, using quantitative methods of confocal laser scanning microscopy and image analysis. Amyloid burden, expressed as the percentage area occupied by thioflavin S-positive amyloid deposits, increased an average of 179-fold from 12 to 54 weeks of age (0.02+/-0.01% to 3.57+/-0.29%, mean+/-S.E.M., respectively) in five regions of the cortex and two of the hippocampus. This was a function of increases in both deposit number and size.This transgenic mouse provides an ideal animal model for evaluating the efficacy of potential therapeutic agents aimed at reducing amyloid deposition, such as inhibitors of amyloid fibril formation or secretase inhibitors.     

Sequence variants of IDE are associated with the extent of beta-amyloid deposition in the Alzheimer's disease brain

Insulin degrading enzyme, encoded by IDE, plays a primary role in the degradation of amyloid beta-protein (A beta), the deposition of which in senile plaques is one of the defining hallmarks of Alzheimer's disease (AD). We recently identified haplotypes in a broad linkage disequilibrium (LD) block encompassing IDE that associate with several AD-related quantitative traits. Here, by examining 32 polymorphic markers extending across IDE and testing quantitative measures of plaque density and cognitive function in three independent Swedish AD samples, we have refined the probable position of pathogenic sequences to a 3' region of IDE, with local maximum effects in the proximity of marker rs1887922. To replicate these findings, a subset of variants were examined against measures of brain A beta load in an independent English AD sample, whereby maximum effects were again observed for rs1887922. For both Swedish and English autopsy materials, variation at rs1887922 explained approximately 10% of the total variance in the respective histopathology traits. However, across all clinical materials studied to date, this variant site does not appear to associate directly with disease, suggesting that IDE may affect AD severity rather than risk. Results indicate that alleles of IDE contribute to variability in A beta deposition in the AD brain and suggest that this relationship may have relevance for the degree of cognitive dysfunction in AD patients.     

Enhanced nociceptive responding in two rat models of depression is associated with alterations in monoamine levels in discrete brain regions

Altered pain responding in depression is a widely recognized but poorly understood phenomenon. The present study investigated nociceptive responding to acute (thermal and mechanical) and persistent (inflammatory) noxious stimuli in two animal models of depression, the olfactory bulbectomized (OB) and the Wistar-Kyoto (WKY) rat. In addition, this study examined if altered nociceptive behaviour was associated with changes in monoamine levels in discrete brain regions. OB rats exhibited mechanical allodynia (von Frey test) but not thermal hyperalgesia (hot plate and tail-flick tests) when compared to sham-operated counterparts. Formalin-induced nociceptive behaviour was both heightened and prolonged in OB versus sham-operated controls. An inverse correlation was observed between 5-hydroxyindoleacetic acid (5-HIAA) concentration in the hippocampus and amygdaloid cortex and nociceptive behaviour in the formalin test. In comparison, WKY rats exhibited thermal hyperalgesia in the hot plate test, while behaviour in the tail-flick and von Frey tests did not differ between WKY and Sprague-Dawley rats. Furthermore, WKY rats exhibited enhanced formalin-evoked nociceptive responding up to 40 min post administration, an effect inversely correlated with serotonin and 5-HIAA levels in the hypothalamus. In conclusion, these findings demonstrate that altered pain responding observed in clinically depressed patients can be modelled pre-clinically, providing a means of investigating the neurochemical basis of, and possible treatments for, this phenomenon.     

Beta-amyloid deposition in brain is enhanced in mouse models of arterial hypertension

There are conflicting evidence regarding the association of hypertension with Alzheimer's disease (AD), and so far it is still unexplored whether increased blood pressure levels can be mechanistically related to the pathophysiology of AD. Since the deposition of beta-amyloid (A beta) in brain represents the first pathogenetic event in the onset of AD, in this study we investigated the role of hypertension in the brain deposition of A beta. We analyzed two independent mouse models of hypertension. In both models we observed an increased permeability of blood-brain barrier in cortex and hippocampus. More interestingly, in the same areas hypertensive mice showed a marked positivity to anti-A beta antibodies and the presence of A beta-like fragments. Finally, we analyzed mice after passive immunotherapy with anti-A beta IgG. We observed that this latter approach determined a markedly reduced A beta immunopositivity in both cortex and hippocampus. Our study demonstrates that chronic hypertension determines an impairment of the blood-brain barrier permeability with deposition of A beta in brain tissue and that passive immunotherapy prevents this latter phenomenon.     

Amyloid neuropathology in the single Arctic APP transgenic model affects interconnected brain regions

The Arctic APP mutation (E693G) within the amyloid β (Aβ) domain of amyloid precursor protein (APP) leads to dementia with clinical features similar to Alzheimer's disease (AD), which is believed to be mediated via increased formation of protofibrils. We have generated a transgenic mouse model, TgAPParc, with neuron-specific expression of human amyloid precursor protein with the Arctic mutation (hAPParc), showing mild amyloid pathology with a relatively late onset. Here we performed a detailed analysis of the spatiotemporal progression of neuropathology in homozygous TgAPParc, focusing on intracellular Aβ and diffuse Aβ aggregates rather than amyloid plaques. We show that the neuropathology in homozygous TgAPParc mice starts with intracellular Aβ aggregates, which is followed by diffuse extracellular Aβ deposits in subiculum that later expands to brain regions receiving neuronal projections from regions already affected. Together this suggests that the pathology in TgAPParc mice affects interconnected brain regions and may represent a valuable tool to study the spread and progression of neuropathology in Alzheimer's disease.     

Alterations of pulmonary zinc homeostasis and cytokine production following traumatic brain injury in rats

Previous studies have shown that labile zinc and inflammatory mediators participate in many pathophysiological processes. The present study investigated the effects of traumatic brain injury (TBI) on the levels of labile zinc and certain proinflammatory factors in rat lung. Male Wistar rats were randomly assigned to 7 groups as follows: normal group, group with sham operation, and TBI groups that were sacrificed respectively at 1, 6, 24, and 72 hr, and on day 7 post-injury. Pulmonary labile zinc, tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-8, and wet/dry weight ratio were measured at the specified time intervals. TBI caused a gradual increase of pulmonary labile zinc as demonstrated by fluorescence staining with Zinpyr-4 (ZP4). The levels of TNF-alpha and IL-8 and the lung wet/dry weight ratios were higher in the TBI groups compared to the normal and sham-operated groups (p <0.05). There were highly positive correlations between the intensity of ZP4 fluorescence and the pulmonary levels of TNF-alpha and IL-8. The results suggest that TBI induces rapid increases of labile zinc and inflammatory mediators in lung, which may participate in the pathogenesis of acute lung injury.     

A synthetic peptide blocking the apolipoprotein E/beta-amyloid binding mitigates beta-amyloid toxicity and fibril formation in vitro and reduces beta-amyloid plaques in transgenic mice

Alzheimer's disease (AD) is associated with accumulation of beta-amyloid (Abeta). A major genetic risk factor for sporadic AD is inheritance of the apolipoprotein (apo) E4 allele. ApoE can act as a pathological chaperone of Abeta, promoting its conformational transformation from soluble Abeta into toxic aggregates. We determined if blocking the apoE/Abeta interaction reduces Abeta load in transgenic (Tg) AD mice. The binding site of apoE on Abeta corresponds to residues 12 to 28. To block binding, we synthesized a peptide containing these residues, but substituted valine at position 18 to proline (Abeta12-28P). This changed the peptide's properties, making it non-fibrillogenic and non-toxic. Abeta12-28P competitively blocks binding of full-length Abeta to apoE (IC50 = 36.7 nmol). Furthermore, Abeta12-28P reduces Abeta fibrillogenesis in the presence of apoE, and Abeta/apoE toxicity in cell culture. Abeta12-28P is blood-brain barrier-permeable and in AD Tg mice inhibits Abeta deposition. Tg mice treated with Abeta12-28P for 1 month had a 63.3% reduction in Abeta load in the cortex (P = 0.0043) and a 59.5% (P = 0.0087) reduction in the hippocampus comparing to age-matched control Tg mice. Antibodies against Abeta were not detected in sera of treated mice; therefore the observed therapeutic effect of Abeta12-28P cannot be attributed to an antibody clearance response. Our experiments demonstrate that compounds blocking the interaction between Abeta and its pathological chaperones may be beneficial for treatment of beta-amyloid deposition in AD.     

Accumulation of amyloid beta-protein in the low-density membrane domain accurately reflects the extent of beta-amyloid deposition in the brain

To learn more about the process of amyloid beta-protein (Abeta) deposition in the brain, human prefrontal cortices were fractionated by sucrose density gradient centrifugation, and the Abeta content in each fraction was quantified by a two-site enzyme-linked immunosorbent assay. The fractionation protocol revealed two pools of insoluble Abeta. One corresponded to a low-density membrane domain; the other was primarily composed of extracellular Abeta deposits in those cases in which Abeta accumulated to significant levels. Abeta42 levels in the low-density membrane domain were proportional to the extent of total Abeta42 accumulation, which is known to correlate well with overall amyloid burden. In PDAPP mice that form senile plaques and accumulate Abeta in a similar manner to aging humans, Abeta42 accumulation in the low-density membrane domain also increased as Abeta deposition progressed with aging. These observations indicate that the Abeta42 associated with low-density membrane domains is tightly coupled with the process of extracellular Abeta deposition.     

Neuronal and microglial involvement in beta-amyloid protein deposition in Alzheimer's disease.

This study was undertaken to localize amyloid precursor protein (APP) and to determine how APP might be released and proteolyzed to yield the beta-amyloid protein deposits found in senile plaques in the brains of Alzheimer's disease patients. We found that antibodies to recombinantly expressed APP labeled many normal neurons and neurites. In addition, dystrophic neurites in different types of senile plaques and degenerating neurons in the temporal cortex and hippocampus of Alzheimer's disease patients were immunostained. We also detected small clusters of dystrophic APP immunoreactive neurites that were not associated with beta-amyloid protein deposits. Microglia was involved in different types of senile plaques and often were associated closely with APP immunoreactive neurites and neurons. The greatest concurrence of APP immunoreactivity and reactive microglia was seen in the subiculum and area CA1, regions with a high density of congophilic plaques and subject to intense Alzheimer's pathology. Our findings suggest that neuronally derived APP is the source for senile plaque beta-amyloid protein, while microglia may act as processing cells.     

Cerebrovascular changes in the rat brain in two models of ischemia

BackgroundVascular occlusion and cyanide neurotoxicity induces oxidative stress and degeneration in the brain. This oxidant induced stress changes the vascular dynamics of cerebral blood vessels, and participates in homeostatic response mechanisms which balance oxygen supply to hypoxic stress-sensitive neurons. The associated changes in vascular morphology include remodeling of the microvasculature and endothelial changes, alterations in regional circulation and variations in the blood brain barrier (BBB). This study compares alterations in physiology of the cerebral artery after a short-term oxidative stress induced by cyanide toxicity and vascular occlusion.MethodAdult Wistar rats (N = 30) were divided into three groups; vascular occlusion (VO) (n = 12), potassium cyanide administration (CN) (n = 12) and Control-CO (n = 6). The CN rates were treated with 30 mg/kg of orally administered KCN while the VO was subjected to global vascular occlusion, both for a duration of 10 days, described as the treatment phase. Control animals were fed on normal rat chow and water for 10 days. At the end of the treatment phase, n = 6 animals in each of the VO, CN and VO groups were anesthetized with sodium pentobarbital (50IP) and the CCA exposed, after which pin electrodes were implanted to record the spikes form the tunica media of the CCA. After day 10, treatment was discontinued for these animals, each remaining in the VO and CN groups (VO-I and CN-I) until day 20 (withdrawal phase) following which the spikes were recorded using the procedure described above.Results/DiscussionVascular occlusion and cyanide toxicity increased vascular resistance in the MCA (reduced lumen thickness ratio) and increased the diameter of the CCA after the treatment phase of 10 days. After 10 days of withdrawal, the VO group showed a reduction in resistance and an increase in the lumen width/wall thickness ratio (LWR) while the CN group showed increased resistance and a reduction in LWR.ConclusionCyanide toxicity increased vascular resistance by inducing degenerative changes in the wall of the artery while vascular occlusion increased resistance through mechanical stress and increased thickness of arterial wall. After the withdrawal phase, vascular resistance diminished in the VO to a significantly greater extent than the CN.