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COVID-19, caused by the severe acute respiratory coronavirus 2 (SARS-CoV-2), currently has no treatment for acute infection. The main protease (Mpro) of SARS-CoV-2 is an essential enzyme for viral replication and an attractive target for disease intervention. The phenothiazine moiety has demonstrated drug versatility for biological systems, including inhibition of butyrylcholinesterase, a property important in the cholinesterase anti-inflammatory cascade. Nineteen phenothiazine drugs were investigated using in silico modelling techniques to predict binding energies and inhibition constants (Ki values) with SARS-CoV-2 Mpro. Because most side-effects of phenothiazines are due to interactions with various neurotransmitter receptors and transporters, phenothiazines with few such interactions were also investigated. All compounds were found to bind to the active site of SARS-CoV-2 Mpro and showed Ki values ranging from 1.30 to 52.4 µM in a rigid active site. Nine phenothiazines showed inhibition constants i values. Docking results were compared with remdesivir and showed similar interactions with key residues Glu-166 and Gln-189 in the active site. This work has identified several phenothiazines with limited neurotransmitter receptor and transporter interactions and that may provide the dual action of inhibiting SARS-CoV-2 Mpro to prevent viral replication and promote the release of anti-inflammatory cytokines to curb viral-induced inflammation. These compounds are promising candidates for further investigation against SARS-CoV-2.

Background: Microglia contribute to Alzheimer’s disease (AD) pathogenesis by clearing amyloid-β (Aβ) and driving neuroinflammation. Domestic dogs with age-related dementia (canine cognitive dysfunction (CCD)) develop cerebral amyloidosis like humans developing AD, and studying such dogs can provide novel information about microglial response in prodromal AD. Objective: The aim was to investigate the microglial response in the cortical grey and the subcortical white matter in dogs with CCD versus age-matched cognitively normal dogs. Methods: Brains from aged dogs with CCD and age-matched controls without dementia were studied. Cases were defined by dementia rating score. Brain sections were stained for Aβ, thioflavin S, hyperphosphorylated tau, and the microglial-macrophage ionized calcium binding adaptor molecule 1 (Iba1). Results were correlated to dementia rating score and tissue levels of Aβ. Results: Microglial numbers were higher in the Aβ plaque-loaded deep cortical layers in CCD versus control dogs, while the coverage by microglial processes were comparable. Aβ plaques were of the diffuse type and without microglial aggregation. However, a correlation was found between the %Iba1 area and insoluble Aβ 42 and N-terminal pyroglutamate modified Aβ(N3pE)-42. The %Iba1 area was higher in white matter, showing phosphorylation of S396 tau, versus grey matter. Perivascular macrophage infiltrates were abundant in the white matter particularly in CDD dogs. Conclusion: The results from this study of the microglial-macrophage response in dogs with CCD are suggestive of relatively mild microglial responses in the Aβ plaque-loaded deep cortical layers and perivascular macrophage infiltrates in the subcortical white matter, in prodromal AD.

In sporadic Alzheimer's disease (SpAD), acetylcholinesterase and butyrylcholinesterase, co-regulators of acetylcholine, are associated with β-amyloid plaques and tau neurofibrillary tangles in patterns suggesting a contribution to neurotoxicity. This association has not been explored in early-onset familial Alzheimer's disease (FAD). We investigated whether cholinesterases are observed in the neuropathological hallmarks in FAD expressing the presenilin 1 Leu235Pro mutation. Brain tissues from three FAD cases and one early-onset SpAD case were stained and analyzed for β-amyloid, tau, α-synuclein, acetylcholinesterase and butyrylcholinesterase. AD pathology was prominent throughout the rostrocaudal extent of all 4 brains but α-synuclein-positive neurites were present in only one familial case. In FAD and SpAD cases, cholinergic activity was associated with plaques and tangles but not with α-synuclein pathology. Both cholinesterases showed similar or decreased plaque staining than detected with β-amyloid immunostaining but greater plaque deposition than observed with thioflavin-S histofluorescence. Acetylcholinesterase and butyrylcholinesterase are highly associated with AD pathology in inherited disease and both may represent specific diagnostic and therapeutic targets for all AD forms.

Background: In Alzheimer’s disease (AD), and amyloid models such as the 5XFAD mouse, butyrylcholinesterase (BChE) is associated with β-amyloid (Aβ) plaques and has unique biochemical features which distinguish it from that found in neurons. It has been suggested that BChE associated with Aβ plaques may be involved in the maturation of this structure and thus disease progression. Objective: Currently, it is unknown whether BChE bound to Aβ plaques has altered biochemical properties due to a different primary structure or because of the association of this enzyme with Aβ plaques. Also, the source and binding mechanism of this BChE remains unknown. Methods: Brain tissue sections from the 5XFAD/BChE-KO mouse were incubated with exogenous sources of BChE and stained for this enzyme’s activity. Efforts were made to determine what region of BChE or Aβ may be involved in this association. Results: We found that incubation of 5XFAD/BChE-KO brain tissues with exogenous BChE led to this enzyme becoming associated with Aβ plaques and neurons. In contrast to neuronal BChE, the BChE bound to Aβ plaques had similar biochemical properties to those seen in AD. Mutations to BChE and efforts to block Aβ epitomes failed to prevent this association. Conclusion: The association of BChE with Aβ plaques, and the resultant biochemical changes, suggests that BChE may undergo a conformational change when bound to Aβ plaques but not neurons. The 5XFAD/BChE-KO model is ideally suited to explore the binding mechanism of BChE to Aβ plaques as well as the involvement of BChE in AD pathogenesis.

A subset of octogenarians and older maintain normal cognitive function (CNOO) despite high prevalence and incidence of cognitive decline attributed to neurodegeneration or aging in the population. The rostral prefrontal cortex (rPFC) and hippocampal formation are brain regions integral to cognition, namely attention and memory, facilitated in part by cholinergic innervation. We hypothesized that preserved cholinergic neurotransmission in these regions contributes to intact cognition in the CNOO. To test this, we evaluated the burden of neuropathological and cholinesterase-associated protein aggregates in the rPFC and hippocampal formation. Tissues from age- and sex-matched CNOO and Alzheimer's disease (AD) rPFC and hippocampal formation were stained for β-amyloid (Aβ), tau, α-synuclein, phosphorylated TAR DNA-binding protein 43 (pTDP-43), acetylcholinesterase (AChE), and butyrylcholinesterase (BChE). The relative abundance of neuropathological aggregates was semi-quantitatively scored. Deposition of Aβ plaques, tau neurofibrillary tangles (NFT) and pTDP-43 inclusions were comparable between CNOO and AD cases. Intraneuronal Aβ and tau-positive thorny astrocytes consistent with aging-related tau astrogliopathy, were also noted in the rPFC. Abundance of BChE-positive plaque pathology was significantly higher in AD than in CNOO cases in most regions of interest, followed closely by abundance of AChE-positive plaque pathology. BChE and AChE activities were also associated with varied NFT morphologies. CNOO cases maintained cognition despite a high neuropathological burden in the rPFC and hippocampal formation. BChE-positive and, to a lesser extent, AChE-positive pathologies were significantly lower in most regions in the CNOO compared to AD. This suggests a specificity of cholinesterase-associated neuropathology with AD. We conclude that while CNOO have cholinesterase-associated neuropathology in the rPFC and hippocampal formation, abundance in this population is significantly lower compared to AD which may contribute to their intact cognition.

Intravenous thrombolysis with alteplase bolus followed by infusion is a global standard of care for patients with acute ischaemic stroke. We aimed to determine whether tenecteplase given as a single bolus might increase reperfusion compared with this standard of care.In this multicentre, open-label, parallel-group, registry-linked, randomised, controlled trial (AcT), patients were enrolled from 22 primary and comprehensive stroke centres across Canada. Patients were eligible for inclusion if they were aged 18 years or older, with a diagnosis of ischaemic stroke causing disabling neurological deficit, presenting within 4·5 h of symptom onset, and eligible for thrombolysis per Canadian guidelines. Eligible patients were randomly assigned (1:1), using a previously validated minimal sufficient balance algorithm to balance allocation by site and a secure real-time web-based server, to either intravenous tenecteplase (0·25 mg/kg to a maximum of 25 mg) or alteplase (0·9 mg/kg to a maximum of 90mg; 0·09 mg/kg as a bolus and then a 60 min infusion of the remaining 0·81 mg/kg). The primary outcome was the proportion of patients who had a modified Rankin Scale (mRS) score of 0-1 at 90-120 days after treatment, assessed via blinded review in the intention-to-treat (ITT) population (ie, all patients randomly assigned to treatment who did not withdraw consent). Non-inferiority was met if the lower 95% CI of the difference in the proportion of patients who met the primary outcome between the tenecteplase and alteplase groups was more than -5%. Safety was assessed in all patients who received any of either thrombolytic agent and who were reported as treated. The trial is registered with ClinicalTrials.gov, NCT03889249, and is closed to accrual.Between Dec 10, 2019, and Jan 25, 2022, 1600 patients were enrolled and randomly assigned to tenecteplase (n=816) or alteplase (n=784), of whom 1577 were included in the ITT population (n=806 tenecteplase; n=771 alteplase). The median age was 74 years (IQR 63-83), 755 (47·9%) of 1577 patients were female and 822 (52·1%) were male. As of data cutoff (Jan 21, 2022), 296 (36·9%) of 802 patients in the tenecteplase group and 266 (34·8%) of 765 in the alteplase group had an mRS score of 0-1 at 90-120 days (unadjusted risk difference 2·1% [95% CI - 2·6 to 6·9], meeting the prespecified non-inferiority threshold). In safety analyses, 27 (3·4%) of 800 patients in the tenecteplase group and 24 (3·2%) of 763 in the alteplase group had 24 h symptomatic intracerebral haemorrhage and 122 (15·3%) of 796 and 117 (15·4%) of 763 died within 90 days of starting treatment INTERPRETATION: Intravenous tenecteplase (0·25 mg/kg) is a reasonable alternative to alteplase for all patients presenting with acute ischaemic stroke who meet standard criteria for thrombolysis.Canadian Institutes of Health Research, Alberta Strategy for Patient Oriented Research Support Unit.

ObjectiveTo test the hypothesis that degree of frailty and neuropathologic burden independently contribute to global cognition and odds of dementia.MethodsThis was a secondary analysis of a prospective cohort study of older adults living in Illinois. Participants underwent an annual neuropsychological and clinical evaluation. We included 625 participants (mean age 89.7 ± 6.1 years; 67.5% female) who died and underwent autopsy. We quantified neuropathology using an index measure of 10 neuropathologic features: β-amyloid deposition, hippocampal sclerosis, Lewy bodies, tangle density, TDP-43, cerebral amyloid angiopathy, arteriolosclerosis, atherosclerosis, and gross and chronic cerebral infarcts. Clinical consensus determined dementia status, which we coded as no cognitive impairment, mild cognitive impairment, or dementia. A battery of 19 tests spanning multiple domains quantified global cognition. We operationalized frailty using a 41-item frailty index. We employed regression analyses to model relationships between neuropathology, frailty, and dementia.ResultsBoth frailty and a neuropathology index were independently associated with global cognition and dementia status. These results held after controlling for traditional pathologic measures in a sample of participants with Alzheimer clinical syndrome. Frailty improved the fit of the model for dementia status (χ2[2] 72.64; p < 0.0001) and explained an additional 11%–12% of the variance in the outcomes.ConclusionDementia is a multiply determined condition, to which both general health, as captured by frailty, and neuropathology significantly contribute. This integrative view of dementia and health has implications for prevention and therapy; specifically, future research should evaluate frailty as a means of dementia risk reduction.

Upon publication of this article [1], it was brought to our attention that one of the 303 participants in the normative study should have been deleted from the database.

The disease, COVID-19, is caused by the severe acute respiratory coronavirus 2 (SARS-CoV-2) for which there is currently no treatment. The SARS-CoV-2 main protease (Mpro) is an important enzyme for viral replication. Small molecules that inhibit this protease could lead to an effective COVID-19 treatment. The 2-pyridone scaffold was previously identified as a possible key pharmacophore to inhibit SARS-CoV-2 Mpro. A search for natural, antimicrobial products with the 2-pyridone moiety was undertaken herein, and their calculated potency as inhibitors of SARS-CoV-2 Mpro was investigated. Thirty-three natural products containing the 2-pyridone scaffold were identified from the literature. An in silico methodology using AutoDock was employed to predict the binding energies and inhibition constants (Ki values) for each 2-pyridone-containing compound with SARS-CoV-2 Mpro. This consisted of molecular optimization of the 2-pyridone compound, docking of the compound with a crystal structure of SARS-CoV-2 Mpro, and evaluation of the predicted interactions and ligand-enzyme conformations. All compounds investigated bound to the active site of SARS-CoV-2 Mpro, close to the catalytic dyad (His-41 and Cys-145). Thirteen molecules had predicted Ki values, Highlights • 2-pyridone-scaffold is an inhibitory pharmacophore for SARS-CoV-2 Mpro. • Thirty-three natural, antimicrobial products identified with 2-pyridone moiety. • All 2-pyridone natural products bind to active site of SARS-CoV-2 Mpro in silico. • Thirteen molecules found to have potent inhibitory activity against SARS-CoV-2 Mpro. • Inhibition of SARS-CoV-2 by natural 2-pyridones may lead to treatment of COVID-19.

Kinetic studies and molecular modeling of human acetylcholinesterase (AChE) inhibition by a fluorinated acetophenone derivative, 1-(3-tert-butylphenyl)-2,2,2-trifluoroethanone (TFK), were performed. Fast reversible inhibition of AChE by TFK is of competitive type with Ki = 5.15 nM. However, steady state of inhibition is reached slowly. Kinetic analysis showed that TFK is a slow-binding inhibitor (SBI) of type B with Ki* = 0.53 nM. Reversible binding of TFK provides a long residence time, &tau, = 20 min, on AChE. After binding, TFK acylates the active serine, forming an hemiketal. Then, disruption of hemiketal (deacylation) is slow. AChE recovers full activity in approximately 40 min. Molecular docking and MD simulations depicted the different steps. It was shown that TFK binds first to the peripheral anionic site. Then, subsequent slow induced-fit step enlarged the gorge, allowing tight adjustment into the catalytic active site. Modeling of interactions between TFK and AChE active site by QM/MM showed that the &ldquo, isomerization&rdquo, step of enzyme-inhibitor complex leads to a complex similar to substrate tetrahedral intermediate, a so-called &ldquo, transition state analog&rdquo, followed by a labile covalent intermediate. SBIs of AChE show prolonged pharmacological efficacy. Thus, this fluoroalkylketone intended for neuroimaging, could be of interest in palliative therapy of Alzheimer&rsquo, s disease and protection of central AChE against organophosphorus compounds.

Molecular imaging agents targeting butyrylcholinesterase (BChE) have shown promise in other neurodegenerative disorders and may have utility in detecting changes to normal appearing white matter in multiple sclerosis (MS). BChE activity is present in white matter and localizes to activated microglia associated with MS lesions. The purpose of this study was to further characterize changes in the cholinergic system in MS pathology, and to explore the utility of BChE radioligands as potential diagnostic and treatment monitoring agents in MS. Cortical and white matter lesions were identified using myelin staining, and lesions were classified based on microglial activation patterns. Adjacent brain sections were used for cholinesterase histochemistry and in vitro autoradiography using phenyl 4-[123I]-iodophenylcarbamate (123I-PIP), a previously described small-molecule cholinesterase-binding radioligand. BChE activity is positively correlated with microglial activation in white matter MS lesions. There is no alteration in cholinesterase activity in cortical MS lesions. 123I-PIP autoradiography revealed uptake of radioactivity in normal white matter, absence of radioactivity within demyelinated MS lesions, and variable uptake of radioactivity in adjacent normal-appearing white matter. BChE imaging agents have the potential to detect MS lesions and subtle pathology in normal-appearing white matter in postmortem MS brain tissue. The possibility of BChE imaging agents serving to supplement current diagnostic and treatment monitoring strategies should be evaluated.

The cetacean hippocampal formation has been noted to be one of the smallest relative to brain size of all mammals studied. This region, comprised of the dentate gyrus, hippocampus proper, subiculum, presubiculum, parasubiculum and the entorhinal cortex, is important in learning, memory, and navigation. There have been a number of studies detailing the distribution of acetylcholinesterase (AChE) in the hippocampal formation of terrestrial mammals with the goal of gaining a greater understanding of some aspects of the cholinergic innervation to this region, as well as its parcellation. The present study was undertaken to describe the organization, cytoarchitecture, and distribution of AChE in the hippocampal formation of the Atlantic white-sided dolphin (AWSD) with the view to understand similarities and differences between this aquatic mammal and terrestrial mammals. Nissl-staining demonstrated cytoarchitecture of the hippocampal formation in the AWSD comparable to that reported in other cetaceans. In addition, the AWSD had a rich pattern of AChE staining that distinctly varied between regions and laminae. A number of differences in the distribution of AChE staining in areas comparable to those of terrestrial species reported suggested possible alterations in connectivity of this region. Overall, however, AChE-staining suggested that cholinergic innervation, neural pathways and function of the hippocampal formation of the AWSD is conserved, similar to other mammals.

Alzheimer disease (AD) is the most common neurodegenerative disorder that causes dementia and is a significant cause of morbidity and mortality. The etiology of AD remains unknown and a definitive diagnosis is not yet achievable for the disease during life. As a consequence, intense focus has been placed on establishing reliable AD biomarkers that both improve the certainty of an AD diagnosis during life and improve our understanding of the pathogenesis of the disease. In this chapter we explore biomarkers currently being evaluated in AD. Components of the central cholinergic system will be emphasized, especially highlighting the enzyme butyrylcholinesterase as an important diagnostic biomarker in AD.

Objective: The Clock Drawing Test (CDT) is commonly used as a screening tool for the assessment of dementia. The association between the CDT in acute stroke and long-term functional and cognitive outcomes in this population is unknown. The present prospective study is the first to examine if CDT scores in the acute stage after stroke are related to long-term outcomes and to compare the predictive ability of two scoring systems in a large sample of stroke patients. Method: A total of 340 patients admitted to an acute stroke unit were included in the present study. Separate stepwise multiple linear regression analyses were performed with eight independent variables (demographic/pre-stroke variables - age, sex, premorbid functioning; stroke-related variables - stroke severity, localization; cognitive variables - Orientation Test, CDT [2 scoring systems]), and four dependent variables administered one year post-stroke (Barthel Index, modified Rankin Scale, Reintegration to Normal Living index, Global Deterioration Scale). Results: Although both CDT scoring methods were related to all long-term outcome measures, the more comprehensive scoring system was the only baseline variable that significantly explained the variance in outcome measures in all four multiple regression models. Conclusion: Performance on the CDT in acute stroke is related to long-term outcomes including patients' degree of independence in performing activities of daily living, the degree to which they achieved reintegration into daily occupations, and the degree of cognitive decline observed one-year post-stroke. Future studies are needed to clarify the nature of the relationship between different CDT scoring systems and post-stroke outcomes.

The basal forebrain (BFB) cholinergic neurotransmitter system is important in a number of brain functions including attention, memory, and the sleep-wake cycle. The size of this region has been linked to the increase in encephalization of the brain in a number of species. Cetaceans, particularly those belonging to the family Delphinidae, have a relatively large brain compared to its body size and it is expected that the cholinergic BFB in the dolphin would be a prominent feature. However, this has not yet been explored in detail. This study examines and maps the neuroanatomy and cholinergic chemoarchitecture of the BFB in the Atlantic white-sided dolphin (Lagenorhynchus acutus). As in some other mammals, the BFB in this species is a prominent structure along the medioventral surface of the brain. The parcellation and distribution of cholinergic neural elements of the dolphin BFB was comparable to that observed in other mammals in that it has a medial septal nucleus, a nucleus of the vertical limb of the diagonal band of Broca, a nucleus of the horizontal limb of the diagonal band of Broca, and a nucleus basalis of Meynert. The observed BFB cholinergic system of this dolphin is consistent with evolutionarily conserved and important functions for survival.

Background A need exists for easily administered assessment tools to detect mild cognitive changes that are more comprehensive than screening tests but shorter than a neuropsychological battery and that can be administered by physicians, as well as any health care professional or trained assistant in any medical setting. The Toronto Cognitive Assessment (TorCA) was developed to achieve these goals. Methods We obtained normative data on the TorCA (n = 303), determined test reliability, developed an iPad version, and validated the TorCA against neuropsychological assessment for detecting amnestic mild cognitive impairment (aMCI) (n = 50/57, aMCI/normal cognition). For the normative study, healthy volunteers were recruited from the Rotman Research Institute registry. For the validation study, the sample was comprised of participants with aMCI or normal cognition based on neuropsychological assessment. Cognitively normal participants were recruited from both healthy volunteers in the normative study sample and the community. Results The TorCA provides a stable assessment of multiple cognitive domains. The total score correctly classified 79% of participants (sensitivity 80%; specificity 79%). In an exploratory logistic regression analysis, indices of Immediate Verbal Recall, Delayed Verbal and Visual Recall, Visuospatial Function, and Working Memory/Attention/Executive Control, a subset of the domains assessed by the TorCA, correctly classified 92% of participants (sensitivity 92%; specificity 91%). Paper and iPad version scores were equivalent. Conclusions The TorCA can improve resource utilization by identifying patients with aMCI who may not require more resource-intensive neuropsychological assessment. Future studies will focus on cross-validating the TorCA for aMCI, and validation for disorders other than aMCI.

Background: Although mood and sleep disturbances are nearly universal among patients with Alzheimer's disease (AD), brain structures involved in non-cognitive processing remain under characterized in terms of AD pathology. Objectives: This study was designed to evaluate hallmarks of AD pathology in the brainstem of the APPswe/PS1dE9 mouse model of familial AD. Methods: Fresh-frozen sections from female, 12 month old, transgenic and control B6C3 mice (n=6/genotype) were examined for amyloid burden and neurofibrillary alterations, by using 6E10 immunohistochemistry and the Gallyas silver stain, respectively. Serotonin transporter (SERT) densities in the dorsal and the median raphe were quantified by [3H]DASB autoradiography. SERT mRNA expression was measured by RT-PCR and visualized by in situ hybridization. Neuroinflammation was evaluated by immunohistochemical staining for microglia and astrocytes, and by measuring mRNA levels of the proinflammatory cytokines TNF-α, IL-1β and IL-6. Results: No amyloid- and tau-associated lesions were observed in the midbrain raphe of 12 month old APPswe/PS1dE9 mice. SERT binding levels were reduced in transgenic animals compared to age-matched controls, and SERT mRNA levels were decreased by at least 50% from control values. Intense microglial, but not astrocytic immunoreactivity was observed in APPswe/PS1dE9 vs. wild-type mice. Levels of TNF-α mRNA were two-fold higher than control and correlated positively with SERT mRNA expression levels in transgenic animals. Conclusions: There was no amyloid accumulation and tau-associated pathology in the midbrain raphe of 12 month old APPswe/PS1dE9 mice. However, there was a local neuroinflammatory response with loss of serotonergic markers, which may partially account for some of the behavioral symptoms of AD.

Mutations in the microtubule-associated protein tau gene are known to cause progressive neurodegenerative disorders with variable clinical and neuropathological phenotypes, including the intronic 10 + 14 (IVS10 + 14) splice site mutation. Three families have been reported with the IVS10 + 14 microtubule-associated protein tau mutation. Here, we describe the clinical and neuropathological data from an additional family. Neuropathological data were available for 2 of the 3 cases, III-4, and III-5. While III-5 had widespread tau deposition and atrophy, III-4 exhibited more mild neuropathological changes except for the substantia nigra. The previously reported families that express the IVS10 + 14 mutation exhibited significant interfamilial heterogeneity, with symptoms including amyotrophy, dementia, disinhibition, parkinsonism, and breathing problems. In addition to expressing many of these symptoms, members of this fourth family experienced profound sensory abnormalities and sleep disturbance. Although there were probable clinicopathological correlates for the symptoms expressed by the earlier families and III-5 from our cohort, pathology in III-4 did not appear sufficient to explain symptom severity. This indicates the need to explore alternate mechanisms of tau-induced brain dysfunction.

Alzheimer's disease (AD) is the most common neurodegenerative disorder causing dementia. One hallmark of the AD brain is the deposition of β-amyloid (Aβ) plaques. AD is also a state of cholinergic dysfunction and butyrylcholinesterase (BChE) associates with Aβ pathology. A transgenic mouse (5XFAD) is an aggressive amyloidosis model, producing Aβ plaques with which BChE also associates. A derived strain (5XFAD/BChE-KO), with the BChE gene knocked out, has significantly lower fibrillar Aβ than 5XFAD mice at the same age. Therefore, BChE may have a role in Aβ pathogenesis. Furthermore, in AD, diminished glucose metabolism in the brain can be detected in vivo with positron emission tomography (PET) imaging following 2-deoxy-2-(18F)fluoro-D-glucose (18FDG) administration. To determine whether hypometabolism is related to BChE-induced changes in fibrillar Aβ burden, whole brain and regional uptake of 18FDG in 5XFAD and 5XFAD/BChE-KO mice was compared to corresponding wild-type (WT5XFAD and WTBChE-KO) strains at 5months. Diminished fibrillar Aβ burden was confirmed in 5XFAD/BChE-KO mice relative to 5XFAD. 5XFAD and 5XFAD/BChE-KO mice demonstrated reduction in whole brain 18FDG retention compared to respective wild-types. Regional analysis of relevant AD structures revealed reduction in 18FDG retention in 5XFAD mice in all brain regions analyzed (save cerebellum) compared to WT5XFAD. Alternatively, 5XFAD/BChE-KO mice demonstrated a more selective pattern of reduced retention in the cerebral cortex and thalamus compared to WTBChE-KO, while retention in hippocampal formation, amygdala and basal ganglia remained unchanged. This suggests that in knocking out BChE and reducing fibrillar Aβ, a possible protective effect on brain function may be conferred in a number of structures in 5XFAD/BChE-KO mice.

Amyloid-β (Aβ) plaques are a neuropathological hallmark of Alzheimer's disease (AD); however, a significant number of cognitively normal older adults can also have Aβ plaques. Thus, distinguishing AD from cognitively normal individuals with Aβ plaques (NwAβ) based on Aβ plaque detection is challenging. It has been observed that butyrylcholinesterase (BChE) accumulates in plaques preferentially in AD. Thus, detecting BChE-associated plaques has the potential as an improved AD biomarker. We present Aβ, thioflavin-S, and BChE quantification of 26 postmortem brain tissues; AD (n = 8), NwAβ (n = 6), cognitively normal without plaques (n = 8), and other common dementias including corticobasal degeneration, frontotemporal dementia with tau, dementia with Lewy bodies, and vascular dementia. Pathology burden in the orbitofrontal cortex, entorhinal cortex, amygdala, and hippocampal formation was determined and compared. The predictive value of Aβ and BChE quantification was determined, via receiver-operating characteristic plots, to evaluate their AD diagnostic performance using sensitivity, specificity, and area under curve (AUC) metrics. In general, Aβ and BChE-associated pathology were greater in AD, particularly in the orbitofrontal cortex. In this region, the largest increase (9.3-fold) was in BChE-associated pathology, observed between NwAβ and AD, due to the virtual absence of BChE-associated plaques in NwAβ brains. Furthermore, BChE did not associate with pathology of the other dementias. In this sample, BChE-associated pathology provided better diagnostic performance (AUC = 1.0, sensitivity/specificity = 100% /100%) when compared to Aβ (AUC = 0.98, 100% /85.7%). These findings highlight the predictive value of BChE as a biomarker for AD that could facilitate timely disease diagnosis and management.

Introduction Diagnosis of Alzheimer's disease (AD) in vivo, by molecular imaging of amyloid or tau, is constrained because similar changes can be found in brains of cognitively normal individuals. Butyrylcholinesterase (BChE), which becomes associated with these structures in AD, could elevate the accuracy of AD diagnosis by focusing on BChE pathology in the cerebral cortex, a region of scant BChE activity in healthy brain. Methods N-methylpiperidin-4-yl 4-[123I]iodobenzoate, a BChE radiotracer, was injected intravenously into B6SJL-Tg(APPSwFlLon, PSEN1∗M146 L∗L286 V) 6799Vas/Mmjax (5XFAD) mice and their wild-type (WT) counterparts for comparative single photon emission computed tomography (SPECT) studies. SPECT, computed tomography (CT), and magnetic resonance imaging (MRI) enabled comparison of whole brain and regional retention of the BChE radiotracer in both mouse strains. Results Retention of the BChE radiotracer was consistently higher in the 5XFAD mouse than in WT, and differences were particularly evident in the cerebral cortex. Discussion Cerebral cortical BChE imaging with SPECT can distinguish 5XFAD mouse model from the WT counterpart.

Despite compelling evidence that the accumulation of amyloid-beta (Aβ) promotes neocortical MAPT (tau) aggregation in familial and idiopathic Alzheimer’s disease (AD), murine models of cerebral amyloidosis are not considered to develop tau-associated pathology. In the present study, we show that tau can accumulate spontaneously in aged transgenic APPswe/PS1ΔE9 mice. Tau pathology is abundant around Aβ deposits, and further characterized by accumulation of Gallyas and thioflavin-S-positive inclusions, which were detected in the APPswe/PS1ΔE9 brain at 18 months of age. Age-dependent increases in argyrophilia correlated positively with binding levels of the paired helical filament (PHF) tracer [18F]Flortaucipir, in all brain areas examined. Sarkosyl-insoluble PHFs were visualized by electron microscopy. Quantitative proteomics identified sequences of hyperphosphorylated and three-repeat tau in transgenic mice, along with signs of RNA missplicing, ribosomal dysregulation and disturbed energy metabolism. Tissue from the frontal gyrus of human subjects was used to validate these findings, revealing primarily quantitative differences between the tau pathology observed in AD patient vs. transgenic mouse tissue. As physiological levels of endogenous, ‘wild-type’ tau aggregate secondarily to Aβ in APPswe/PS1ΔE9 mice, this study suggests that amyloidosis is both necessary and sufficient to drive tauopathy in experimental models of familial AD.

Background: Primary Progressive Aphasia (PPA) is a syndrome characterized by an isolated impairment of language function at disease onset. The cholinergic system is implicated in language function and cholinergic deficits are seen in the brains of individuals with PPA. One major source of cholinergic innervation of the cerebral cortex is the nucleus basalis of Meynert (NBM) within which lies the nucleus subputaminalis (NSP). This nucleus is postulated to be involved in language function. We compared the abundance of cholinergic neurons in the NBM and NSP of controls and individuals with PPA. Also explored was whether the individuals presenting with PPA, who subsequently developed different clinical and neuropathological profiles, showed similar cholinergic deficits in the NSP. Methods: Cytoarchitecture of the basal forebrain was studied using Nissl staining in control (n = 5) and PPA (n = 5) brains. Choline acetyltransferase (ChAT) immunohistochemical staining labeled cholinergic neurons were quantified using Neurolucida software. Results: In comparison to matched controls, PPA showed reduction of cholinergic neurons in the NBM (t(8) = 4.04, p = 0.0037; Cohen’s effect size value d = 2.62) and the NSP (t(6) = 4.62, p = 0.0042; Cohen’s d effect size d = 2.92). The average percent of cholinergic neuronal loss was relatively higher in the NSP (64.7%) compared to the NBM (47.7%). Conclusion: Regardless of underlying pathology, all cases presenting with PPA showed a marked loss of cholinergic neurons in the NSP, providing further evidence for the importance of this nucleus in language function.

Alzheimer's disease (AD) is an age-related neurodegenerative disorder that causes profound cognitive dysfunction. Deficits in olfactory memory occur in early stages of AD and may be useful in AD diagnosis. The 5xFAD mouse is a commonly used model of AD, as it develops neuropathology, cognitive and sensori-motor dysfunctions similar to those seen in AD. However, olfactory memory dysfunction has not been studied adequately or in detail in 5xFAD mice. Furthermore, despite sex differences in AD prevalence and symptom presentation, few studies using 5xFAD mice have examined sex differences in learning and memory. Therefore, we tested olfactory memory in male and female 5xFAD mice from 3 to 15 months of age using a conditioned odour preference task. Olfactory memory was not impaired in male or female 5xFAD mice at any age tested, nor were there any sex differences. Because early-onset impairments in very long-term (remote) memory have been reported in 5xFAD mice, we trained a group of mice at 3 months of age and tested olfactory memory 90 days later. Very long-term olfactory memory in 5xFAD mice was not impaired, nor was their ability to perform the discrimination task with new odourants. Examination of brains from 5xFAD mice confirmed extensive Aβ-plaque deposition spanning the olfactory memory system, including the olfactory bulb, hippocampus, amygdala and piriform cortex. Overall this study indicates that male and female 5xFAD mice do not develop olfactory memory deficits, despite extensive Aβ deposition within the olfactory-memory regions of the brain.

Despite compelling evidence that the accumulation of amyloid-beta (Aβ) promotes cortical MAPT (tau) aggregation in familial and idiopathic Alzheimer’s disease (AD), murine models of cerebral amyloidosis are not considered to develop tau-associated pathology. The absence of neurofibrillary lesions in amyloidosis mice remains a challenge for the amyloidocentric paradigm of AD pathogenesis. It has resulted in the generation of transgenic mice harboring mutations in theirtaugene, which may be inappropriate for studying a disease with no knownTAUmutations, such as AD. Here, we have usedAPPswe/PS1ΔE9mice to show that tau pathology can develop spontaneously in murine models of familial AD. Tauopathy was abundant around Aβ deposits, with Gallyas- and thioflavin-S-positive perinuclear inclusions accumulating in theAPPswe/PS1ΔE9cortex by 18 months of age. Age-dependent increases in Gallyas signal correlated positively with binding levels of the paired helical filament (PHF) ligand [18F]Flortaucipir, in all brain areas examined. Sarkosyl-insoluble PHFs were visualized by electron microscopy. Tandem mass tag proteomics identified sequences of hyperphosphorylated tau in transgenic mice, along with signs of RNA missplicing, ribosomal dysregulation and disturbed energy metabolism. Human frontal gyrus tissue was used to validate these findings, revealing primarily quantitative differences between the tauopathy observed in AD patient vs. transgenic mouse tissue. Levels oftaumRNA were not different betweenAPPswe/PS1ΔE9and littermate control animals. As physiological levels of endogenous, ‘wild-type’ tau aggregate secondarily to Aβ in transgenic mice, this study demonstrates that amyloidosis is both necessary and sufficient to drive tauopathy in experimental models of familial AD.

Objective: Neuronal ceroid-lipofuscinoses are a heterogeneous group of inherited disorders in which abnormal lipopigments form lysosomal inclusion bodies in neurons. Kufs disease is rare, and clinical symptoms include seizures, progressive cognitive impairment, and myoclonus. Most cases of Kufs disease are autosomal recessive; however, there have been a few case reports of an autosomal dominant form linked to mutations within the DNAJC5 gene. Methods: We describe a family with Kufs disease in which the proband and three of her four children presented with cognitive impairment, seizures, and myoclonus. Results: Genetic testing of all four children was positive for a c.346_348delCTC(p.L116del) mutation in the DNAJC5 gene. The proband brain had an abundance of neuronal lipofuscin in the cerebral cortex, striatum, amygdala, hippocampus, substantia nigra, and cerebellum. There were no amyloid plaques or neurofibrillary tangles. Immunohistochemistry demonstrated that the cholinergic neurons and cholinergic projection fibers were spared, but there was a profound loss of choline acetyltransferase within the caudate, putamen, and basal forebrain. This suggests a loss of choline acetyltransferase as opposed to a loss of the neurons. Conclusions: This report describes the clinical history of autosomal dominant Kufs disease, the genetic mutation within the DNAJC5 gene, and the neuropathological findings demonstrating depletion of choline acetyltransferase in the brain.

In Alzheimer’s disease (AD), numerous β-amyloid (Aβ) plaques are associated with butyrylcholinesterase (BChE) activity, the significance of which is unclear. A mouse model, containing five human familial AD genes (5XFAD), also develops Aβ plaques with BChE activity. Knock-out of BChE in this model showed diminished fibrillar Aβ plaque deposition, more so in males than females. This suggests that lack of BChE reduces deposition of fibrillar Aβ in AD and this effect may be influenced by sex.

Aims Alzheimer's disease (AD) is characterized by cholinergic dysfunction and deposition of β-amyloid (Aβ) plaques and tau neurofibrillary tangles (NFTs) in the brain. Olfactory abnormalities often precede cognitive symptoms in AD, indicating early involvement of pathology in olfactory structures. The cholinergic system is important not only in cognition but also in modulation of the olfactory system. The primary olfactory gyrus (POG) is comprised of the olfactory tract, anterior olfactory nucleus (AON) and olfactory area (OA). Because of the importance of the olfactory and cholinergic systems, we examined the anatomical and cholinergic organization of the POG in normal human brain and neuropathology in AD. Methods Cytoarchitecture of the POG was studied using Nissl staining in normal (n=8) and AD (n=6) brains. Distributions of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) were determined using histochemical methods. Aβ plaques and tau NFTs were detected using immunohistochemistry. Abundance of AD pathology was assessed using a semi-quantitative approach. Results Nissl staining showed pyramidal cells in the AON and paleocortical organization of the OA. AChE stained neurons and neuropil in the AON and OA, while BChE activity was noted in the olfactory tract and in AON and OA neurons. Pathology was frequent in the AD POG and the abundance of BChE-associated AD pathology was greater than that associated with AChE. Conclusions AChE and BChE activities in normal POG recapitulated their distributions in other cortical regions. Greater abundance of BChE-associated, in comparison to AChE-associated, AD pathology in the POG suggests preferential involvement of BChE in olfactory dysfunction in AD. This article is protected by copyright. All rights reserved.

Cholinesterase inhibitors are the standard of care for Alzheimer's disease (AD). Acetylcholinesterase (AChE) catalyzes the hydrolysis of the cholinergic neurotransmitter acetylcholine. However, the related enzyme butyrylcholinesterase (BuChE) also breaks down acetylcholine and is likewise targeted by the same clinical cholinesterase inhibitors. The lack of clinical efficacy for the highly specific and potent AChE inhibitor, (-) huperzine A, is intriguing, given the known cholinergic deficit in AD. Based on the proven efficacy of inhibitors affecting both cholinesterases and the apparent failure of specific AChE inhibition, focused BuChE inhibition seems important for more effective treatment of AD. Therefore, BuChE-selective inhibitors provide promise for improved benefit.

Brain glucose hypometabolism has been observed in Alzheimer’s disease (AD) patients, and is detected with 18 F radiolabelled glucose, using positron emission tomography. A pathological hallmark of AD is deposition of brain β - amyloid plaques that may influence cerebral glucose metabolism. The five times familial AD (5XFAD) mouse is a model of brain amyloidosis exhibiting AD-like phenotypes. This study examines brain β -amyloid plaque deposition and 18 FDG uptake, to search for an early biomarker distinguishing 5XFAD from wild-type mice. Thus, brain 18 FDG uptake and plaque deposition was studied in these mice at age 2, 5 and 13 months. The 5XFAD mice demonstrated significantly reduced brain 18 FDG uptake at 13 months relative to wild-type controls but not in younger mice, despite substantial β - amyloid plaque deposition. However, by comparing the ratio of uptake values for glucose in different regions in the same brain, 5XFAD mice could be distinguished from controls at age 2 months. This method of measuring altered glucose metabolism may represent an early biomarker for the progression of amyloid deposition in the brain. We conclude that brain 18 FDG uptake can be a sensitive biomarker for early detection of abnormal metabolism in the 5XFAD mouse when alternative relative uptake values are utilized.