Covert strokes prior to Alzheimer's disease onset accelerate peri-lesional pathology but not cognitive deficits in an inducible APP mouse model.

It is estimated that up to 1 in 3 healthy middle-aged adults will have had a covert stroke during their lifetime. Furthermore, post-stroke, survivors are more than twice as likely to develop dementia. In the present study, we aimed to model the impact of focal subclinical ischemia prior to the onset of AD pathogenesis in a preclinical model. We utilized endothelin-1 to induce ischemia in an iducible transgenic mouse model of Alzheimer's disease, APPsi:tTA, allowing for temporal control of APP gene expression. We induced the focal subclinical ischemic events in the absence of APP expression, thus prior to AD onset. T2 structural magnetic resonance imaging confirmed the volume and location of focal subclinical ischemic lesions to the medial prefrontal cortex. Following recovery from surgery and 7 weeks of APP expression, we found that two subclinical ischemic lesions resulted in a significant localized increase in amyloid load and in microglial activation proximal to the lesion. However, no differences were found in astrogliosis. A battery of behaviour tests was conducted, in which no significant differences were detected in activities of daily living and cognitive function between stroked and sham cohorts. Overall, our results demonstrated that APP expression was the sole driving force behind behavioural deficits. In conclusion, our results suggest that a history of two subclinical strokes prior to AD onset does not worsen early disease trajectory in a mouse model.

The effects of voluntary running on cerebrovascular morphology and spatial short-term memory in a mouse model of amyloidosis.

Physical activity has been correlated with a reduced risk of cognitive decline, including that associated with vascular dementia, mild cognitive impairment (MCI) and Alzheimer's disease (AD); recent literature suggests this may in part result from benefits to the cerebrovascular network. Using a transgenic (Tg) mouse model of AD, we evaluated the effect of running on cortical and hippocampal vascular morphology, cerebral amyloid angiopathy, amyloid plaque load, and spatial memory. TgCRND8 mice present with progressive amyloid pathology, advancing from the cortex to the hippocampus in a time-dependent manner. We postulated that the characteristic progression of pathology could lead to differential, time-dependent effects of physical activity on vascular morphology in these brain regions at 6 months of age. We used two-photon fluorescent microscopy and 3D vessel tracking to characterize vascular and amyloid pathology in sedentary TgCRND8 mice compared those who have a history of physical activity (unlimited access to a running wheel, from 3 to 6 months of age). In sedentary TgCRND8 mice, capillary density was found to be lower in the cortex and higher in the hippocampus compared to non-transgenic (nonTg) littermates. Capillary length, vessel branching, and non-capillary vessel tortuosity were also higher in the hippocampus of sedentary TgCRND8 compared to nonTg mice. Three months of voluntary running resulted in normalizing cortical and hippocampal microvascular morphology, with no significant difference between TgCRND8 and nonTg mice. The benefits of physical activity on cortical and hippocampal vasculature in 6-month old TgCRND8 mice were not paralleled by significant changes on parenchymal and cerebral amyloid pathology. Short-term spatial memory- as evaluated by performance in the Y-maze- was significantly improved in running compared to sedentary TgCRND8 mice. These results suggest that long-term voluntary running contributes to the maintenance of vascular morphology and spatial memory in TgCRND8 mice, even in the absence of an effect on amyloid pathology.

Reduced Cerebrovascular Reactivity and Increased Resting Cerebral Perfusion in Rats Exposed to a Cafeteria Diet.

To better understand the effects of a diet high in fat, sugar, and sodium on cerebrovascular function, Sprague Dawley rats were chronically exposed to a Cafeteria diet. Resting cerebral perfusion and cerebrovascular reactivity was quantified using continuous arterial spin labeling (CASL) magnetic resonance imaging (MRI). In addition, structural changes to the cerebrovasculature and susceptibility to ischemic lesion were examined. Compared to control animals fed standard chow (SD), Cafeteria diet (CAF) rats exhibited increased resting brain perfusion in the hippocampus and reduced cerebrovascular reactivity in response to 10% inspired CO2 challenges in both the hippocampus and the neocortex. CAF rats switched to chow for one month (SWT) exhibited improved resting perfusion in the hippocampus as well as improved cerebrovascular reactivity in the neocortex. However, the diet switch did not correct cerebrovascular reactivity in the hippocampus. These changes were not accompanied by alterations in the structural integrity of the cerebral microvasculature, examined using rat endothelial cell antigen-1 (RECA-1) and immunoglobulin G (IgG) immunostaining. Also, the extent of tissue damage induced by endothelin-1 injection into sensorimotor cortex was not affected by the Cafeteria diet. These results demonstrate that short-term consumption of an ultra-processed diet reduces cerebrovascular reactivity. This effect persists after dietary normalization despite recovery of peripheral symptomatology.

Early neurovascular dysfunction in a transgenic rat model of Alzheimer's disease.

Alzheimer's disease (AD), pathologically characterized by amyloid-ß peptide (Aß) accumulation, neurofibrillary tangle formation, and neurodegeneration, is thought to involve early-onset neurovascular abnormalities. Hitherto studies on AD-associated neurovascular injury have used animal models that exhibit only a subset of AD-like pathologies and demonstrated some Aß-dependent vascular dysfunction and destabilization of neuronal network. The present work focuses on the early stage of disease progression and uses TgF344-AD rats that recapitulate a broader repertoire of AD-like pathologies to investigate the cerebrovascular and neuronal network functioning using in situ two-photon fluorescence microscopy and laminar array recordings of local field potentials, followed by pathological analyses of vascular wall morphology, tau hyperphosphorylation, and amyloid plaques. Concomitant to widespread amyloid deposition and tau hyperphosphorylation, cerebrovascular reactivity was strongly attenuated in cortical penetrating arterioles and venules of TgF344-AD rats in comparison to those in non-transgenic littermates. Blood flow elevation to hypercapnia was abolished in TgF344-AD rats. Concomitantly, the phase-amplitude coupling of the neuronal network was impaired, evidenced by decreased modulation of theta band phase on gamma band amplitude. These results demonstrate significant neurovascular network dysfunction at an early stage of AD-like pathology. Our study identifies early markers of pathology progression and call for development of combinatorial treatment plans.

Neurovascular unit remodelling in the subacute stage of stroke recovery.

Brain plasticity following focal cerebral ischaemia has been observed in both stroke survivors and in preclinical models of stroke. Endogenous neurovascular adaptation is at present incompletely understood yet its potentiation may improve long-term functional outcome. We employed longitudinal MRI, intracranial array electrophysiology, Montoya Staircase testing, and immunofluorescence to examine function of brain vessels, neurons, and glia in addition to forelimb skilled reaching during the subacute stage of ischemic injury progression. Focal ischemic stroke (~100mm3 or ~20% of the total brain volume) was induced in adult Sprague-Dawley rats via direct injection of endothelin-1 (ET-1) into the right sensori-motor cortex, producing sustained impairment in left forelimb reaching ability. Resting perfusion and vascular reactivity to hypercapnia in the peri-lesional cortex were elevated by approximately 60% and 80% respectively seven days following stroke. At the same time, the normal topological pattern of local field potential (LFP) responses to peripheral somatosensory stimulation was abolished and the average power of spontaneous LFP activity attenuated by approximately 50% relative to the contra-lesional cortex, suggesting initial response attenuation within the peri-infarct zone. By 21 days after stroke, perilesional blood flow resolved, but peri-lesional vascular reactivity remained elevated. Concomitantly, the LFP response amplitudes increased with distance from the site of ET-1 injection, suggesting functional remodelling from the core of the lesion to its periphery. This notion was further buttressed by the lateralization of spontaneous neuronal activity: by day 21, the average ipsi-lesional power of spontaneous LFP activity was almost twice that of the contra-lesional cortex. Over the observation period, the peri-lesional cortex exhibited increased vascular density, along with neuronal loss, astrocytic activation, and recruitment and activation of microglia and macrophages, with neuronal loss and inflammation extending beyond the peri-lesional cortex. These findings highlight the complex relationship between neurophysiological state and behaviour and provide evidence of highly dynamic functional changes in the peri-infarct zone weeks following the ischemic insult, suggesting an extended temporal window for therapeutic interventions.

Effects of Neurotrophic Support and Amyloid-Targeted Combined Therapy on Adult Hippocampal Neurogenesis in a Transgenic Model of Alzheimer's Disease.

Although it is recognized that multi-drug therapies may be necessary to combat AD, there is a paucity of preclinical proof of concept studies. We present a combination treatment paradigm, which temporally affects different aspects of Alzheimer's disease (AD)-like pathology, specifically Aß-toxicity and neurogenesis. At early stages of AD-like pathology, in TgCRND8 mice, we found that combating Aß pathology with scyllo-inositol ameliorated deficits in neurogenesis. Older TgCRND8 mice with established amyloid load had decreased progenitor cell proliferation and survival compared to non-transgenic mice, regardless of scyllo-inositol treatment. The prolonged exposure to Aß-pathology leads to deficits in the neurogenic niche, thus targeting Aß alone is insufficient to rescue neurogenesis. To support the neurogenic niche, we combined scyllo-inositol treatment with leteprinim potassium (neotrofin), the latter of which stimulates neurotrophin expression. We show that the combination treatment of scyllo-inositol and neotrofin enhances neuronal survival and differentiation. We propose this proof of concept combination therapy of targeting Aß-pathology and neurotrophin deficits as a potential treatment for AD.

Magnetic resonance spectroscopy reveals oral Lactobacillus promotion of increases in brain GABA, N-acetyl aspartate and glutamate.

The gut microbiome has been shown to regulate the development and functions of the enteric and central nervous systems. Its involvement in the regulation of behavior has attracted particular attention because of its potential translational importance in clinical disorders, however little is known about the pathways involved. We previously have demonstrated that administration of Lactobacillus rhamnosus (JB-1) to healthy male BALB/c mice, promotes consistent changes in GABA-A and -B receptor sub-types in specific brain regions, accompanied by reductions in anxiety and depression-related behaviors. In the present study, using magnetic resonance spectroscopy (MRS), we quantitatively assessed two clinically validated biomarkers of brain activity and function, glutamate+glutamine (Glx) and total N-acetyl aspartate+N-acetyl aspartyl glutamic acid (tNAA), as well as GABA, the chief brain inhibitory neurotransmitter. Mice received 1×10(9) cfu of JB-1 per day for 4weeks and were subjected to MRS weekly and again 4weeks after cessation of treatment to ascertain temporal changes in these neurometabolites. Baseline concentrations for Glx, tNAA and GABA were equal to 10.4±0.3mM, 8.7±0.1mM, and 1.2±0.1mM, respectively. Delayed increases were first seen for Glx (~10%) and NAA (~37%) at 2weeks which persisted only to the end of treatment. However, Glx was still elevated 4weeks after treatment had ceased. Significantly elevated GABA (~25%) was only seen at 4weeks. These results suggest specific metabolic pathways in our pursuit of mechanisms of action of psychoactive bacteria. They also offer through application of standard clinical neurodiagnostic techniques, translational opportunities to assess biomarkers accompanying behavioral changes induced by alterations in the gut microbiome.

Attenuation of functional hyperemia to visual stimulation in mild Alzheimer's disease and its sensitivity to cholinesterase inhibition.

Despite the growing recognition of the significance of cerebrovascular impairment in the etiology and progression of Alzheimer's disease (AD), the early stage brain vascular dysfunction and its sensitivity to pharmacological interventions is still not fully characterized. Due to the early and aggressive treatment of probable AD with cholinesterase inhibitors (ChEI), which in and of themselves have direct effects on brain vasculature, the vast majority of hemodynamic measurements in early AD subjects reported hitherto have consequently been made only after the start of treatment, complicating the disentanglement of disease- vs. treatment-related effects on the cerebral vasculature. To address this gap, we used pseudo continuous arterial spin labeling MRI to measure resting perfusion and visual stimulation elicited changes in cerebral blood flow (CBF) and blood oxygenation dependent (BOLD) fMRI signal in a cohort of mild AD patients immediately prior to, 6months post, and 12months post commencement of open label cholinesterase inhibitor treatment. Although patients exhibited no gray matter atrophy prior to treatment and their resting perfusion was not distinguishable from that in age, education and gender-matched controls, the patients' visual stimulation-elicited changes in BOLD fMRI and blood flow were decreased by 10±4% (BOLD) and 23±2% (CBF), relative to those in controls. Induction of cholinesterase inhibition treatment was associated with a further, 7±2% reduction in patients' CBF response to visual stimulation, but it stabilized, at this new lower level, over the follow-up period. Likewise, MMSE scores remained stable during the treatment; furthermore, higher MMSE scores were associated with higher perfusion responses to visual stimulation. This study represents the initial step in disentangling the effects of AD pathology from those of the first line treatment with cholinesterase inhibitors on cerebral hemodynamics and supports the use of arterial spin labeling MRI for quantitative evaluation of the brain vascular function in mild Alzheimer's disease. The findings provide evidence of a pronounced deficit in the visual cortex hyperemia despite the relative sparing of visual function in early stage AD, its reduction with ChEI treatment induction, and its stabilization in the first year of cholinesterase inhibition treatment. This article is part of a Special Issue entitled: Vascular Contributions to Cognitive Impairment and Dementia edited by M. Paul Murphy, Roderick A. Corriveau and Donna M. Wilcock.

Venular degeneration leads to vascular dysfunction in a transgenic model of Alzheimer's disease.

Most patients with Alzheimer's disease exhibit accumulation of amyloid-ß peptide on leptomeningeal and cortical arterioles, or cerebral amyloid angiopathy, which is associated with impaired vascular reactivity and accelerated cognitive decline. Despite widespread recognition of the significance of vascular dysfunction in Alzheimer's disease aetiology and progression, much uncertainty still surrounds the mechanism underlying Alzheimer's disease vascular injury. Studies to date have focused on amyloid-ß-induced damage to capillaries and plaque-associated arterioles, without examining effects across the entire vascular bed. In the present study, we investigated the structural and functional impairment of the feeding arteriolar versus draining venular vessels in a transgenic murine Alzheimer's disease model, with a particular focus on the mural cell populations that dictate these vessels' contractility. Although amyloid-ß deposition was restricted to arterioles, we found that vascular impairment extended to the venules, which showed significant depletion of their mural cell coverage by the mid-stage of Alzheimer's disease pathophysiology. These structural abnormalities were accompanied by an abolishment of the normal vascular network flow response to hypercapnia: this functional impairment was so severe as to result in hypercapnia-induced flow decreases in the arterioles. Further pharmacological depletion of mural cells using SU6668, a platelet-derived growth factor receptor-ß antagonist, resulted in profound structural abnormalities of the cortical microvasculature, including vessel coiling and short-range looping, increased tortuosity of the venules but not of the arterioles, increased amyloid-ß deposition on the arterioles, and further alterations of the microvascular network cerebral blood flow response to hypercapnia. Together, this work shows hitherto unrecognized structural alterations in penetrating venules, demonstrates their functional significance and sheds light on the complexity of the relationship between vascular network structure and function in Alzheimer's disease.

Cerebrovascular contributions to Alzheimer's disease pathophysiology and potential therapeutic interventions in mouse models.

The inter-relationship between vascular dysfunction and Alzheimer's disease pathology is not clearly understood; however, it is clear that the accumulation of amyloid-beta peptide and loss of vascular function contribute to the cognitive decline detected in patients. At present, imaging modalities can monitor the downstream effects of vascular dysfunction such as cerebral blood flow alterations, white and gray matter lacunes, and ischemic lesions; however, they cannot distinguish parenchymal plaques from cerebrovascular amyloid. Much of our understanding regarding the relationship between amyloid and vascular dysfunction has come from longitudinal population studies and mouse models. In this review, we will discuss the breadth of data generated on vascular function in mouse models of Alzheimer's disease and cerebrovascular amyloid angiopathy. We will also discuss therapeutic strategies targeting the reduction of cerebrovascular amyloid angiopathy and improvement of vascular function.

Reactive astrocytes associated with plaques in TgCRND8 mouse brain and in human Alzheimer brain express phosphoprotein enriched in astrocytes (PEA-15).

To identify potential biomarkers associated with Alzheimer's disease (AD)-like neuropathologies in the murine brain, we conducted proteomic analyses of neocortices from TgCRND8 mice. Here we found that phosphoprotein enriched in astrocytes 15 kDa (PEA-15) is expressed at higher levels in the neocortical proteomes from 6-month old TgCRND8 mice, as compared to non-transgenic mice. Immunostaining for PEA-15 revealed reactive astrocytes associated with the neocortical amyloid plaques in TgCRND8 mice and in post-mortem human AD brains. This is the first report of increased phosphoprotein enriched in astrocytes (PEA-15) expression in reactive astrocytes of an AD mouse model and human AD brains.

scyllo-Inositol, preclinical, and clinical data for Alzheimer's disease.

Preclinical development of scyllo-inositol for the treatment of Alzheimer's disease (AD) has been investigated in both in vitro and in vivo models with positive results. scyllo-Inositol stabilized a small conformer of Aß42 in vitro, neutralized cell derived Aß trimers and promoted low molecular weight Aß species in vivo. These interactions resulted in decreased neuronal toxicity, increased long-term potentiation (LTP) and ablation of cognitive deficits in multiple mouse models of AD. scyllo-Inositol bioavailability, pharmacokinetics, and small animal toxicology studies demonstrated the potential for translation to human patients. The results of Phase I and Phase II clinical trials for AD are presented. Furthermore, the use of this compound for imaging and other amyloid related disorders is discussed.