Monica Garcia-Alloza

Rapid appearance and local toxicity of amyloid-beta plaques in a mouse model of Alzheimer's disease.

Senile plaques accumulate over the course of decades in the brains of patients with Alzheimer’s disease. A fundamental tenet of the amyloid hypothesis of Alzheimer’s disease is that the deposition of amyloid-β precedes and induces the neuronal abnormalities that underlie dementia. This idea has been challenged, however, by the suggestion that alterations in axonal trafficking and morphological abnormalities precede and lead to senile plaques. The role of microglia in accelerating or retarding these processes has been uncertain. To investigate the temporal relation between plaque formation and the changes in local neuritic architecture, we used longitudinal in vivo multiphoton microscopy to sequentially image young APPswe/PS1d9xYFP (B6C3-YFP) transgenic mice. Here we show that plaques form extraordinarily quickly, over 24 h. Within 1–2 days of a new plaque’s appearance, microglia are activated and recruited to the site. Progressive neuritic changes ensue, leading to increasingly dysmorphic neurites over the next days to weeks. These data establish plaques as a critical mediator of neuritic pathology.

Oligomeric amyloid β associates with postsynaptic densities and correlates with excitatory synapse loss near senile plaques

Synapse loss correlates with a cognitive decline in Alzheimers disease (AD), but whether this is caused by fibrillar deposits known as senile plaques or soluble oligomeric forms of amyloid β (Aβ) is controversial. By using array tomography, a technique that combines ultrathin sectioning of tissue with immunofluorescence, allowing precise quantification of small structures, such as synapses, we have tested the hypothesis that oligomeric Aβ surrounding plaques contributes to synapse loss in a mouse model of AD. We find that senile plaques are surrounded by a halo of oligomeric Aβ. Analysis of >14,000 synapses (represented by PSD95-stained excitatory synapses) shows that there is a 60% loss of excitatory synapses in the halo of oligomeric Aβ surrounding plaques and that the density increases to reach almost control levels in volumes further than 50 μm from a plaque in an approximately linear fashion (linear regression, r2 = 0.9; P < 0.0001). Further, in transgenic cortex, microdeposits of oligomeric Aβ associate with a subset of excitatory synapses, which are significantly smaller than those not in contact with oligomeric Aβ. The proportion of excitatory synapses associated with Aβ correlates with decreasing density (correlation, −0.588; P < 0.0001). These data show that senile plaques are a potential reservoir of oligomeric Aβ, which colocalizes with the postsynaptic density and is associated with spine collapse, reconciling the apparently competing schools of thought of “plaque” vs. “oligomeric Aβ” as the synaptotoxic species in the brain of AD patients.

Curcumin labels amyloid pathology in vivo, disrupts existing plaques, and partially restores distorted neurites in an Alzheimer mouse model

Alzheimer’s disease (AD) is characterized by senile plaques and neurodegeneration although the neurotoxic mechanisms have not been completely elucidated. It is clear that both oxidative stress and inflammation play an important role in the illness. The compound curcumin, with a broad spectrum of anti‐oxidant, anti‐inflammatory, and anti‐fibrilogenic activities may represent a promising approach for preventing or treating AD. Curcumin is a small fluorescent compound that binds to amyloid deposits. In the present work we used in vivo multiphoton microscopy (MPM) to demonstrate that curcumin crosses the blood–brain barrier and labels senile plaques and cerebrovascular amyloid angiopathy (CAA) in APPswe/PS1dE9 mice. Moreover, systemic treatment of mice with curcumin for 7 days clears and reduces existing plaques, as monitored with longitudinal imaging, suggesting a potent disaggregation effect. Curcumin also led to a limited, but significant reversal of structural changes in dystrophic dendrites, including abnormal curvature and dystrophy size. Together, these data suggest that curcumin reverses existing amyloid pathology and associated neurotoxicity in a mouse model of AD. This approach could lead to more effective clinical therapies for the prevention of oxidative stress, inflammation and neurotoxicity associated with AD.

Rapid Microglial Response Around Amyloid Pathology after Systemic Anti-Aβ Antibody Administration in PDAPP Mice

Aggregation of amyloid-β (Aβ) peptide in the brain in the form of neuritic plaques and cerebral amyloid angiopathy (CAA) is a key feature of Alzheimers disease (AD). Microglial cells surround aggregated Aβ and are believed to play a role in AD pathogenesis. A therapy for AD that has entered clinical trials is the administration of anti-Aβ antibodies. One mechanism by which certain anti-Aβ antibodies have been proposed to exert their effects is via antibody-mediated microglial activation. Whether, when, or to what extent microglial activation occurs after systemic administration of anti-Aβ antibodies has not been fully assessed. We administered an anti-Aβ antibody (m3D6) that binds aggregated Aβ to PDAPP mice, an AD mouse model that was bred to contain fluorescent microglia. Three days after systemic administration of m3D6, there was a marked increase in both the number of microglial cells and processes per cell visualized in vivo by multiphoton microscopy. These changes required the Fc domain of m3D6 and were not observed with an antibody specific to soluble Aβ. These findings demonstrate that some effects of antibodies that recognize aggregated Aβ are rapid, involve microglia, and provide insight into the mechanism of action of a specific passive immunotherapy for AD.

Cholinergic-serotonergic imbalance contributes to cognitive and behavioral symptoms in Alzheimer's disease

Neuropsychiatric symptoms seen in Alzheimers disease (AD) are not simply a consequence of neurodegeneration, but probably result from differential neurotransmitter alterations, which some patients are more at risk of than others. Therefore, the hypothesis of this study is that an imbalance between the cholinergic and serotonergic systems is related to cognitive symptoms and psychological syndromes of dementia (BPSD) in patients with AD. Cholinergic and serotonergic functions were assessed in post-mortem frontal and temporal cortex from 22 AD patients who had been prospectively assessed with the Mini-Mental State examination (MMSE) for cognitive impairment and with the Present Behavioral Examination (PBE) for BPSD including aggressive behavior, overactivity, depression and psychosis. Not only cholinergic deficits, but also the cholinacetyltransferase/serotonin ratio significantly correlated with final MMSE score both in frontal and temporal cortex. In addition, decreases in cholinergic function correlated with the aggressive behavior factor, supporting a dual role for the cholinergic system in cognitive and non-cognitive disturbances associated to AD. The serotonergic system showed a significant correlation with overactivity and psychosis. The ratio of serotonin to acetylcholinesterase levels was also correlated with the psychotic factor at least in women. It is concluded that an imbalance between cholinergic-serotonergic systems may be responsible for the cognitive impairment associated to AD. Moreover, the major findings of this study are the relationships between neurochemical markers of both cholinergic and serotonergic systems and non-cognitive behavioral disturbances in patients with dementia.

Differential involvement of 5-HT1B/1D and 5-HT6 receptors in cognitive and non-cognitive symptoms in Alzheimer's disease

Growing evidence suggests that a compromised serotonergic system plays an important role in the pathophysiology of Alzheimers disease (AD). We assessed the expression of 5-HT1B/1D and 5-HT6 receptors and cholinacetyltransferase (ChAT) activity in post-mortem frontal and temporal cortex from AD patients who had been prospectively assessed for cognitive function using the Mini-Mental State Examination (MMSE) and behavioral changes using the Present Behavioral Examination (PBE). 5-HT1B/1D and 5-HT6 receptor densities were significantly reduced in both cortical areas. 5-HT1B/1D receptor density was correlated to MMSE decline in the frontal cortex, supporting its implication in memory impairment. The best predictor for lowered 5-HT6 receptor density in the temporal cortex was the PBE measure of overactivity. The 5-HT6/ChAT ratio was related to aggression both in the frontal and temporal cortex. Therefore, antagonists acting at 5-HT6 receptors could be useful in the treatment of non-cognitive symptoms associated to AD.

Lack of localization of 5‐HT6 receptors on cholinergic neurons: implication of multiple neurotransmitter systems in 5‐HT6 receptor‐mediated acetylcholine release

The involvement of the cholinergic system in learning and memory together with the cognitive enhancing properties of 5‐HT6 receptor antagonists led us to study the relationship between 5‐HT6 receptors and cholinergic neurotransmission. A selective cholinergic lesion, induced by injection of the immunotoxin 192‐IgG‐Saporin into the nucleus basalis magnocellularis, failed to alter the density of 5‐HT6 receptor mRNA or protein expression in the deafferentated frontal cortex, suggesting that 5‐HT6 receptors are not located on cholinergic neurons. The 5‐HT6 receptor antagonist SB‐357134 (0.001–1 µm) induced a concentration‐dependant K+‐evoked [3H]acetylcholine (ACh) release in vitro in rat cortical and striatal slices, which was blocked by tetrodotoxin. SB‐357134, up to 1 µm, stimulated glutamate release in cortical and striatal slices. In the cortex, riluzole (1 µm) blocked the SB‐357134‐induced K+‐stimulated [3H]ACh release, and simultaneous administration of MK‐801 (1 µm) and SB‐357134 (0.05 µm) elicited an increase in K+‐evoked ACh release. In the striatum, SB‐357134, 1 µm, decreased dopamine release, and the increase in K+‐evoked [3H]ACh release induced by 5‐HT6 receptor blockade was reversed by the D1 receptor antagonist, SCH23390 (1 µm). In both the frontal cortex and striatum, bicuculline, 1 µm, showed no effect on SB‐357134‐evoked [3H]ACh. These results are discussed in terms of neurochemical mechanisms involved in 5‐HT6 receptor functions.

Cerebrovascular lesions induce transient β-amyloid deposition

Previous clinical studies have documented a close relationship between cerebrovascular disease and risk of Alzheimers disease. We examined possible mechanistic interactions through use of experimental stroke models in a transgenic mouse model of β-amyloid deposition (APPswe/PS1dE9). Following middle cerebral artery occlusion, we observed a rapid increase in amyloid plaque burden in the region surrounding the infarction. In human tissue samples, however, we were unable to detect a localized increase in amyloid burden adjacent to cerebral infarcts. To resolve this discrepancy, we generated cerebral microstrokes in amyloid precursor protein mouse models with the photosensitive dye Rose bengal, and monitored plaque formation in real time using multiphoton microscopy. We observed a striking increase in the number of new plaques and amyloid angiopathy in the area immediately surrounding the infarcted area; however, the effect was transient, potentially resolving the discord between mouse and human tissue. We did not detect changes in candidate proteins related to β-amyloid generation or degradation such as β-amyloid-converting enzyme, amyloid precursor protein, presenilin 1, neprylisin or insulin-degrading enzyme. Together, these results demonstrate that strokes can trigger accelerated amyloid deposition, most likely through interference with amyloid clearance pathways. Additionally, this study indicates that focal ischaemia provides an experimental paradigm in which to study the mechanisms of plaque seeding and growth.

Detection of isolated cerebrovascular β-amyloid with pittsburgh compound B†

Imaging of cerebrovascular β‐amyloid (cerebral amyloid angiopathy) is complicated by the nearly universal overlap of this pathology with Alzheimers pathology. We performed positron emission tomographic imaging with Pittsburgh Compound B on 42‐year‐old man with early manifestations of Iowa‐type hereditary cerebral amyloid angiopathy, a form of the disorder with little or no plaque deposits of fibrillar β‐amyloid. The results demonstrated increased Pittsburgh Compound B retention selectively in occipital cortex, sparing regions typically labeled in Alzheimers disease. These results offer compelling evidence that Pittsburgh Compound B positron emission tomography can noninvasively detect isolated cerebral amyloid angiopathy before overt signs of tissue damage such as hemorrhage or white matter lesions. Ann Neurol 2008;64:587–591

Kinetics of Cerebral Amyloid Angiopathy Progression in a Transgenic Mouse Model of Alzheimer Disease

Cerebral amyloid angiopathy (CAA), the deposition of cerebrovascular β-amyloid (Aβ) in the walls of arterial vessels, has been implicated in hemorrhagic stroke and is present in most cases of Alzheimer disease. Previous studies of the progression of CAA in humans and animal models have been limited to the comparison of pathological tissue from different brains at single time points. Our objective was to visualize in real time the initiation and progression of CAA in Tg2576 mice by multiphoton microscopy through cranial windows. Affected vessels were labeled by methoxy-X04, a fluorescent dye that selectively binds cerebrovascular β-amyloid and plaques. With serial imaging sessions spaced at weekly intervals, we were able to observe the earliest appearance of CAA in leptomeningeal arteries as multifocal deposits of band-like Aβ. Over subsequent imaging sessions, we were able to identify growth of these deposits (propagation), as well as appearance of new bands (additional initiation events). Statistical modeling of the data suggested that as the extent of CAA progressed in this vascular bed, there was increased prevalence of propagation over initiation. During the early phases of CAA development, the overall pathology burden progressed at a rate of 0.35% of total available vessel area per day (95% confidence interval, 0.3–0.4%). The consistent rate of disease progression implies that this model is amenable to investigations of therapeutic interventions.