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Proper transport of RNAs to synapses is essential for localized translation of proteins in response to synaptic signals and synaptic plasticity. Alzheimer's disease (AD) is a neurodegenerative disease characterized by accumulation of amyloid aggregates and hyperphosphorylated tau neurofibrillary tangles followed by widespread synapse loss. To understand whether RNA synaptic localization is impacted in AD, we performed RNA sequencing on synaptosomes and brain homogenates from AD patients and cognitively healthy controls. This resulted in the discovery of hundreds of mislocalized mRNAs in AD among frontal and temporal brain regions. Similar observations were found in an APPswe/PSEN1dE9 mouse model. Furthermore, major differences were observed among circular RNAs (circRNAs) localized to synapses in AD including two overlapping isoforms of circGSK3β, one upregulated, and one downregulated. Expression of these distinct isoforms affected tau phosphorylation in neuronal cells substantiating the importance of circRNAs in the brain and pointing to a new class of therapeutic targets. Author summary: Synaptic plasticity is the foundation for cognition and describes the process by which synaptic connections between neurons are formed, strengthened, or weakened based on signals transmitted by neighboring neurons. Essential to this process is the ability of synapses to respond rapidly to signals and restructure their morphology by translating proteins from a pool of localized mRNAs which is orchestrated by a complex cytoskeletal trafficking system. Given that synapse loss is a cardinal feature of Alzheimer's disease (AD), we RNA sequenced fractionated synaptic particles from brain samples from AD patients and healthy controls to determine whether RNA localization to synapses is impacted in AD. We discovered hundreds of mislocalized mRNAs as well as circular RNAs (circRNAs) which are non-coding isoforms shown to play critical regulatory roles in the brain. Among our data, we discovered two isoforms of circGSK3β, one upregulated, and one downregulated in AD. Expression of these isoforms in neuronal culture was shown to affect tau phosphorylation, dysregulation of which is a hallmark of AD. These findings point to RNA localization and circRNAs as key components to understanding AD pathology and implicate new targets for AD therapies. [ABSTRACT FROM AUTHOR]

Comparing brain structure across species and regions enables key functional insights. Leveraging publicly available data from a novel mass cytometry-based method, synaptometry by time of flight (SynTOF), we applied an unsupervised machine learning approach to conduct a comparative study of presynapse molecular abundance across three species and three brain regions. We used neural networks and their attractive properties to model complex relationships among high dimensional data to develop a unified, unsupervised framework for comparing the profile of more than 4.5 million single presynapses among normal human, macaque, and mouse samples. An extensive validation showed the feasibility of performing cross-species comparison using SynTOF profiling. Integrative analysis of the abundance of 20 presynaptic proteins revealed near-complete separation between primates and mice involving synaptic pruning, cellular energy, lipid metabolism, and neurotransmission. In addition, our analysis revealed a strong overlap between the presynaptic composition of human and macaque in the cerebral cortex and neostriatum. Our unique approach illuminates species- and region-specific variation in presynapse molecular composition. [ABSTRACT FROM AUTHOR]

Background: Diabetes is a risk factor for Alzheimer's disease and related dementias (ADRD). Epidemiologic evidence shows an association between diabetes medications and ADRD risk; cell and mouse models show diabetes medication association with AD-related neuropathologic change (ADNC).Objective: This hypothesis-generating analysis aimed to describe autopsy-measured ADNC for individuals who used diabetes medications.Methods: Descriptive analysis of ADNC for Adult Changes in Thought (ACT) Study autopsy cohort who used diabetes medications, including sulfonylureas, insulin, and biguanides; total N = 118. ADNC included amyloid plaque distribution (Thal phasing), neurofibrillary tangle (NFT) distribution (Braak stage), and cortical neuritic plaque density (CERAD score). We also examined quantitative measures of ADNC using the means of standardized Histelide measures of cortical PHF-tau and Aβ1-42. Adjusted analyses control for age at death, sex, education, APOE genotype, and diabetes complication severity index.Results: Adjusted analyses showed no significant association between any drug class and traditional neuropathologic measures compared to nonusers of that class. In adjusted Histelide analyses, any insulin use was associated with lower mean levels of Aβ1-42 (-0.57 (CI: -1.12, -0.02)) compared to nonusers. Five years of sulfonylureas and of biguanides use was associated with lower levels of Aβ1-42 compared to nonusers (-0.15 (CI: -0.28, -0.02), -0.31 (CI: -0.54, -0.07), respectively).Conclusion: Some evidence exists that diabetes medications are associated with lower levels of Aβ1-42, but not traditional measures of neuropathology. Future studies are needed in larger samples to build understanding of the mechanisms between diabetes, its medications, and ADRD, and to potentially repurpose existing medications for prevention or delay of ADRD. [ABSTRACT FROM AUTHOR]

Mass‐tag cell barcoding has increased the throughput, multiplexing, and robustness of multiple cytometry approaches. Previously, we adapted mass cytometry for cells to analyze synaptosome preparations (mass synaptometry or SynTOF), extending mass cytometry to these smaller, anuclear particles. To improve throughput and individual event resolution, we report here the application of palladium‐based barcoding in human synaptosomes. Up to 20 individual samples, each with a unique combinatorial barcode, were pooled for labeling with an antibody cocktail. Our synaptosome protocol used six palladium‐based barcoding reagents, and in combination with sequential gating increased the identification of presynaptic events approximately fourfold. These same parameters also efficiently resolved two other anuclear particles: human red blood cells and platelets. The addition of palladium‐based mass‐tag barcoding to our approach improves mass cytometry of synaptic particles. [ABSTRACT FROM AUTHOR]

The late neuropathological effects of traumatic brain injury have yet to be fully elucidated, particularly with respect to community-based cohorts. To contribute to this critical gap in knowledge, we designed a multimodal neuropathological study, integrating traditional and quantitative approaches to detect pathologic changes in 532 consecutive brain autopsies from participants in the Adult Changes in Thought (ACT) study. Diagnostic evaluation including assessment for chronic traumatic encephalopathy (CTE) and quantitative immunoassay-based methods were deployed to examine levels of pathological (hyperphosphorylated) tau (pTau) and amyloid (A) β in brains from ACT participants with (n = 107) and without (n = 425) history of remote TBI with loss of consciousness (w/LOC). Further neuropathological assessments included immunohistochemistry for α-synuclein and phospho-TDP-43 pathology and astro- (GFAP) and micro- (Iba1) gliosis, mass spectrometry analysis of free radical injury, and gene expression evaluation (RNA sequencing) in a smaller sub-cohort of matched samples (49 cases with TBI and 49 non-exposed matched controls). Out of 532 cases, only 3 (0.6%–none with TBI w/LOC history) showed evidence of the neuropathologic signature of chronic traumatic encephalopathy (CTE). Across the entire cohort, the levels of pTau and Aβ showed expected differences for brain region (higher levels in temporal cortex), neuropathological diagnosis (higher in participants with Alzheimer's disease), and APOE genotype (higher in participants with one or more APOE ε4 allele). However, no differences in PHF-tau or Aβ1−42 were identified by Histelide with respect to the history of TBI w/LOC. In a subset of TBI cases with more carefully matched control samples and more extensive analysis, those with TBI w/LOC history had higher levels of hippocampal pTau but no significant differences in Aβ, α-synuclein, pTDP-43, GFAP, Iba1, or free radical injury. RNA-sequencing also did not reveal significant gene expression associated with any measure of TBI exposure. Combined, these findings suggest long term neuropathological changes associated with TBI w/LOC may be subtle, involve non-traditional pathways of neurotoxicity and neurodegeneration, and/or differ from those in autopsy cohorts specifically selected for neurotrauma exposure. [ABSTRACT FROM AUTHOR]

Analgesics are commonly used by older adults, raising the question of whether their use might contribute to dementia risk and neuropathologic changes of Alzheimer's disease (AD). The Adult Changes in Thought (ACT) study is a population-based study of brain aging and incident dementia among people 65 years or older who are community dwelling and not demented at entry. Amyloid-β (Aβ)42 and phospho-tau were quantified using Histelide in regions of cerebral cortex from 420 brain autopsies. Total standard daily doses of prescription opioid and non-aspirin nonsteroidal anti-inflammatory drug (NSAID) exposure during a defined 10-year exposure window were identified using automated pharmacy dispensing data and used to classify people as having no/low, intermediate, or high exposure. People with high NSAID exposure had significantly greater Aβ42 concentration in middle frontal gyrus and superior and middle temporal gyri, but not inferior parietal lobule; no Aβ42 regional concentration was associated with prescription opioid usage. People with high opioid usage had significantly greater concentration of phospho-tau in middle frontal gyrus than people with little-to-no opioid usage. Consistent with our previous studies, findings suggest that high levels of NSAID use in older individuals may promote Aβ42 accumulation in cerebral cortex. [ABSTRACT FROM AUTHOR]

Quantifying antigens in formalin-fixed tissue is challenging and limits investigation in population-based studies of brain aging. To address this major limitation, we have developed a new technique that we call 'Histelide': immunohistochemistry (HIST-) and enzyme-linked immunosorbent assay (ELISA) (-EL-) performed on a glass slide (-IDE). We validated Histelide in sections of prefrontal cortex from 20 selected cases: 12 subjects with clinically and neuropathologically diagnosed Alzheimer's disease (AD), either autosomal dominant or late-onset forms, and 8 clinical and neuropathologic controls. AD cases had significantly increased amyloid beta (Aβ) peptide and paired helical filament- (PHF-) tau per area of neocortex that was proteinase K-sensitive, and significantly decreased amount of synaptophysin. We next investigated prefrontal cortex from 81 consecutive cases of high-cognitive performers from the Adult Changes in Thought (ACT) study, a population-based study of brain aging and incident dementia. As expected, latent AD was common in this group; however, our results quantified widely individually varying levels of Aβ peptides and PHF-tau among these high-cognitive performers. This novel approach obtains quantitative data from population-based studies, and our initial studies with high-cognitive performers provide important quantitative insights into latent AD that should help guide expectations from neuroimaging and prevention studies. [ABSTRACT FROM AUTHOR]

We previously showed that amyloid-β 1-42 (Aβ42) levels in cerebrospinal fluid (CSF) were markedly altered in response to a 4-week dietary intervention in normal aging and mild cognitive impairment (MCI). Here, we re-examined the data to assess whether diet-induced effects on CSF Aβ42 were modulated by high intensity physical activity (hi-PA). Normal older adults (n = 18, mean age = 68.6 ± 7.4 y) and adults with amnestic MCI (n = 23, mean age = 68.0 ± 6.5 y) received a low saturated fat/low glycemic index (LOW) diet or a high saturated fat/high glycemic index (HIGH) diet, and CSF levels of Aβ42, tau, and IL-8 were measured at baseline and week 4. Pre-study activity levels were assessed using a 7-d questionnaire, and weekly duration of hi-PA was quantified. At baseline, increased hi-PA in normals predicted lower CSF levels of tau (r = -0.54, p = 0.020) and IL-8 (r = -0.70, p = 0.025). Diet-induced effects on CSF Aβ42 during the intervention study were modulated by hi-PA, and the nature of this effect differed for normals and MCI (ANOVA, p = 0.039). That is, for normal adults, increased hi-PA attenuated the effects of the HIGH diet on CSF Aβ42 whereas in MCI, increased hi-PA potentiated the effects of the LOW diet. Our results suggest that normal adults who engage in hi-PA are less vulnerable to the pathological effects of an unhealthy diet, while in MCI, the benefit of a healthy diet on Aβ modulation is greatest when paired with hi-PA. Exercise may thus interact with diet to alter pathological processes that ultimately modify risk of Alzheimer's disease. [ABSTRACT FROM AUTHOR]

The striatum is composed predominantly of medium spiny neurons (MSNs) that integrate excitatory, glutamatergic inputs from the cortex and thalamus, and modulatory dopaminergic inputs from the ventral midbrain to influence behavior. Glutamatergic activation of AMPA, NMDA, and metabotropic receptors on MSNs is important for striatal development and function, but the roles of each of these receptor classes remain incompletely understood. Signaling through NMDA-type glutamate receptors (NMDARs) in the striatum has been implicated in various motor and appetitive learning paradigms. In addition, signaling through NMDARs influences neuronal morphology, which could underlie their role in mediating learned behaviors. To study the role of NMDARs on MSNs in learning and in morphological development, we generated mice lacking the essential NR1 subunit, encoded by the Grin1 gene, selectively in MSNs. Although these knockout mice appear normal and display normal 24-hour locomotion, they have severe deficits in motor learning, operant conditioning and active avoidance. In addition, the MSNs from these knockout mice have smaller cell bodies and decreased dendritic length compared to littermate controls. We conclude that NMDAR signaling in MSNs is critical for normal MSN morphology and many forms of learning. [ABSTRACT FROM AUTHOR]

Objective: To compare the effects of a 4-week high-saturated fat/high-glycemic index (HIGH) diet with a low-saturated fat/low-glycemic index (LOW) diet on insulin and lipid metabolism, cerebrospinal fluid (CSF) markers of Alzheimer disease, and cognition for healthy adults and adults with amnestic mild cognitive impairment (aMCI). Design: Randomized controlled trial. Setting: Veterans Affairs Medical Center clinical research unit. Participants: Forty-nine older adults (20 healthy adults with a mean [SD] age of 69.3 [7.4] years and 29 adults with aMCI with a mean [SD] age of 67.6 [6.8] years). Intervention: Participants received the HIGH diet (fat, 45% [saturated fat,>25%]; carbohydrates, 35%-40% [glycemic index,>70]; and protein, 15%-20%) or the LOW diet (fat, 25%; [saturated fat,<7%]; carbohydrates, 55%-60% [glycemic index,<55]; and protein, 15%-20%) for 4 weeks. Cognitive tests, an oral glucose tolerance test, and lumbar puncture were conducted at baseline and during the fourth week of the diet. Main Outcome Measures: The CSF concentrations of β-amyloid (Aβ42 and Aβ40), tau protein, insulin, F2-isoprostanes, and apolipoprotein E, plasma lipids and insulin, and measures of cognition. Results: For the aMCI group, the LOW diet increased CSF Aβ42 concentrations, contrary to the pathologic pattern of lowered CSF Aβ42 typically observed in Alzheimer disease. The LOW diet had the opposite effect for healthy adults, ie, decreasing CSF Aβ42, whereas the HIGH diet increased CSF Aβ42. The CSF apolipoprotein E concentration was increased by the LOW diet and decreased by the HIGH diet for both groups. For the aMCI group, the CSF insulin concentration increased with the LOW diet, but the HIGH diet lowered the CSF insulin concentration for healthy adults. The HIGH diet increased and the LOW diet decreased plasma lipids, insulin, and CSF F2-isoprostane concentrations. Delayed visual memory improved for both groups after completion of 4 weeks of the LOW diet. Conclusion: Our results suggest that diet may be a powerful environmental factor that modulates Alzheimer disease risk through its effects on central nervous system concentrations of Aβ42, lipoproteins, oxidative stress, and insulin. [ABSTRACT FROM AUTHOR]

Cognitive impairment and dementia are more common in patients with Parkinson disease (PD) than age-matched controls and appear to become more frequent as PD progresses. However, estimates of dementia in patients with PD have varied widely, likely due in part to differences in case definition, case ascertainment and methodology. First, we review investigations of usual pathologic correlates of dementia in patients with brainstem (b) Lewy Body Disease (LBD) and report our findings from the initial 266 brain autopsies from a population-based study of brain aging and incident dementia. Our results showed that 2.6% of subjects were diagnosed with PD during life but that 20% had bLBD at autopsy. Seventy percent of individuals with bLBD had high level of one or more cerebral pathologic changes significantly associated with dementia: Alzheimer's disease (AD), cerebral (c) LBD or microvascular brain injury (µVBI); these were commonly co-morbid. Next we consider proposed contributors to cognitive impairment and dementia in the approximately 30% of patients with only bLBD, including regionally selective dendritic degeneration of neostriatal medium spiny neurons. Diseases contributing to cognitive impairment and dementia in patients with bLBD are heterogeneous, providing diagnostic challenges as well as multiple opportunities for successful intervention in patients with PD. [ABSTRACT FROM AUTHOR]

In-cell immunoassays have become a valuable tool for protein expression analysis complementary to established assay formats. However, comprehensive molecular characterization of individual specimens has proven challenging and impractical due to, in part, a singleplex nature of reporter enzymes and technical complexity of alternative assay formats. Herein, we describe a simple and robust methodology for multiplexed protein expression profiling on the same intact specimen, employing a well-characterized enzyme alkaline phosphatase for accurate quantification of all targets of interest, while overcoming fundamental limitations of enzyme-based techniques by implementing the DNA-programmed release mechanism for segregation of sub-sets of target-bound reporters. In essence, this methodology converts same-sample multi-target labeling into a set of isolated singleplex measurements performed in a parallel self-consistent fashion. For a proof-of-principle, multiplexed detection of three model proteins was demonstrated on cultured HeLa cells, and two clinically-relevant markers of dementia, β-amyloid and PHF-tau, were profiled in formalin-fixed paraffin embedded brain tissue sections, uncovering correlated increase in abundance of both markers in the 'Alzheimer's disease' cohort. Featuring an analytically powerful yet technically simple and robust methodology, multiplexed in-cell immunoassay is expected to enable insightful same-sample protein profiling studies and become broadly adopted in biomedical research and clinical diagnostics. [ABSTRACT FROM AUTHOR]