Reduction In Synaptophysin Research Articles

Cigarette Smoke Exposure Induces Neurocognitive Impairments and Neuropathological Changes in the Hippocampus

Background and ObjectiveNeurocognitive dysfunction is present in up to ∼61% of people with chronic obstructive pulmonary disease (COPD), with symptoms including learning and memory deficiencies, negatively impacting the quality of life of these individuals. As the mechanisms responsible for neurocognitive deficits in COPD remain unknown, we explored whether chronic cigarette smoke (CS) exposure causes neurocognitive dysfunction in mice and whether this is associated with neuroinflammation and an altered neuropathology.MethodsMale BALB/c mice were exposed to room air (sham) or CS (9 cigarettes/day, 5 days/week) for 24 weeks. After 23 weeks, mice underwent neurocognitive tests to assess working and spatial memory retention. At 24 weeks, mice were culled and lungs were collected and assessed for hallmark features of COPD. Serum was assessed for systemic inflammation and the hippocampus was collected for neuroinflammatory and structural analysis.ResultsChronic CS exposure impaired lung function as well as driving pulmonary inflammation, emphysema, and systemic inflammation. CS exposure impaired working memory retention, which was associated with a suppression in hippocampal microglial number, however, these microglia displayed a more activated morphology. CS-exposed mice showed changes in astrocyte density as well as a reduction in synaptophysin and dendritic spines in the hippocampus.ConclusionWe have developed an experimental model of COPD in mice that recapitulates the hallmark features of the human disease. The altered microglial/astrocytic profiles and alterations in the neuropathology within the hippocampus may explain the neurocognitive dysfunction observed during COPD.

Altered Brain Cholinergic and Synaptic Markers in Obese Zucker Rats.

The association between obesity and loss of cognitive performance has been recognized. Although there are data regarding the metabolic alterations in obese conditions and the development of neuroinflammation, no clear evidence concerning obesity-related cholinergic and synaptic impairments in the frontal cortex and hippocampus has been reported yet. Here, we investigate different cholinergic and synaptic markers in 12-, 16-, and 20-week-old obese Zucker rats (OZRs) compared with lean littermate rats (LZRs), using immunochemical and immunohistochemical analysis. Consequently, OZRs showed body weight gain, hypertension, and dysmetabolism. In 20-week-old OZRs, the reduction of vesicular acetylcholine transporter (VAChT) and alpha7 nicotinic acetylcholine receptors (α7nAChR) occurred both in the frontal cortex and in the hippocampus, suggesting a cognitive dysfunction due to obesity and aging. Among the muscarinic receptors analyzed, the level of expression of type 1 (mAChR1) was lower in the hippocampus of the older OZRs. Finally, we showed synaptic dysfunctions in OZRs, with a reduction of synaptophysin (SYP) and synaptic vesicle glycoprotein 2B (SV2B) in 20-week-old OZRs, both in the frontal cortex and in the hippocampus. Taken together, our data suggest specific alterations of cholinergic and synaptic markers that can be targeted to prevent cognitive deficits related to obesity and aging.

PD44-11 THERAPEUTIC POTENTIAL OF NITRIC OXIDE AGAINST NEUROENDOCRINE PROSTATE CANCER

You have accessJournal of UrologyProstate Cancer: Basic Research & Pathophysiology II (PD44)1 Sep 2021PD44-11 THERAPEUTIC POTENTIAL OF NITRIC OXIDE AGAINST NEUROENDOCRINE PROSTATE CANCER Fabio Frech, Omar Rosete, Khushi Shah, Fakiha Firdaus, Joshua Hare, Ranjith Ramasamy, and Himanshu Arora Fabio FrechFabio Frech More articles by this author , Omar RoseteOmar Rosete More articles by this author , Khushi ShahKhushi Shah More articles by this author , Fakiha FirdausFakiha Firdaus More articles by this author , Joshua HareJoshua Hare More articles by this author , Ranjith RamasamyRanjith Ramasamy More articles by this author , and Himanshu AroraHimanshu Arora More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002058.11AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Neuroendocrine prostate cancer (NEPC) is an aggressive variant of prostate cancer that may arise de novo or in patients treated with hormonal therapies for prostate adenocarcinoma as a mechanism of resistance. The underlying mechanisms and effective therapies against NEPC are largely underexplored. We showed that S-nitrosoglutathione (GSNO), an NO donor, reduced CRPC tumor burden by targeting the tumor microenvironment (TME). In the present study, we evaluated the impact of increased nitric oxide on NEPC reduction. We hypothesized that increased nitric oxide reduces NEPC tumor burden by suppressing NEPC TME. METHODS: Cell proliferation assay (MTT) was performed using H660 cells (NEPC cells) in the presence of increasing concentration of GSNO (from 0, 5, 10, 25, 50, 100, 200, and 500 uM respectively). Expression of neuroendocrine and proliferative markers (chromogranin A, synaptophysin, and pERK) was checked at RNA and protein levels after treating H660 cells with increasing concentrations of GSNO (from 0, 25, 50, 100, 200, and 500uM respectively). For in-vivo experiments, castrated SCID mice were grafted with H660 cells (3 million) to generate NEPC murine models. Once tumors became palpable, the animals were divided into two groups. Group 1 received vehicle control and group 2 received GSNO treatment at the dosage of 10 mg/kg/day intraperitoneally (IP). Treatment was performed daily for two weeks and tumor growth was monitored until tumor reached 1000 mm3 size. Subsequently, tumors and blood were extracted; proteins were harvested from the tumors to study the expression of neuroendocrine markers and markers of cancer progression. Animal experiments were carried out in compliance with the Institutional Animal Care and Use Committee of University of Miami. Molecular analyses were performed using standard procedures. GraphPad Prism (GraphPad Software) was used for statistical analysis. All data were presented as the means±SEM. The statistical significance between two groups was estimated by unpaired two-tailed t test. RESULTS: Cell proliferation assay showed that increase NO levels suppresses H660 cell proliferation with increasing doses. In-vivo results showed that treatment with GSNO in castrated SCID mice treated with H660 cells showed a significant decrease (p<0.05) in tumor volume compared to the mice which received PBS. Proteins from the tumors showed that GSNO induces a reduction in chromogranin A and synaptophysin (neuroendocrine markers) compared to control. Moreover, immunohistochemistry results showed a significant reduction in synaptophysin, PSA and CD206 levels. CONCLUSIONS: This study shows that GSNO decreases NEPC tumor volume and H660 cell proliferation. GSNO is known to reduce tumor burden in CRPC through interaction with the TME. Further studies investigating the mechanism of effect of nitric oxide in NEPC through the targeting of the TME are in process. Source of Funding: Clinician Scientist Development award from American Cancer Society to RR and Research Scholar Award from American Urological Association to HA © 2021 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 206Issue Supplement 3September 2021Page: e741-e741 Advertisement Copyright & Permissions© 2021 by American Urological Association Education and Research, Inc.MetricsAuthor Information Fabio Frech More articles by this author Omar Rosete More articles by this author Khushi Shah More articles by this author Fakiha Firdaus More articles by this author Joshua Hare More articles by this author Ranjith Ramasamy More articles by this author Himanshu Arora More articles by this author Expand All Advertisement Loading ...

Depletion of acetate-producing bacteria from the gut microbiota facilitates cognitive impairment through the gut-brain neural mechanism in diabetic mice

BackgroundModification of the gut microbiota has been reported to reduce the incidence of type 1 diabetes mellitus (T1D). We hypothesized that the gut microbiota shifts might also have an effect on cognitive functions in T1D. Herein we used a non-absorbable antibiotic vancomycin to modify the gut microbiota in streptozotocin (STZ)-induced T1D mice and studied the impact of microbial changes on cognitive performances in T1D mice and its potential gut-brain neural mechanism.ResultsWe found that vancomycin exposure disrupted the gut microbiome, altered host metabolic phenotypes, and facilitated cognitive impairment in T1D mice. Long-term acetate deficiency due to depletion of acetate-producing bacteria resulted in the reduction of synaptophysin (SYP) in the hippocampus as well as learning and memory impairments. Exogenous acetate supplement or fecal microbiota transplant recovered hippocampal SYP level in vancomycin-treated T1D mice, and this effect was attenuated by vagal inhibition or vagotomy.ConclusionsOur results demonstrate the protective role of microbiota metabolite acetate in cognitive functions and suggest long-term acetate deficiency as a risk factor of cognitive decline.9cNWQAXYUcub27-EQKqGKfVideo

Inflammatory, synaptic, motor, and behavioral alterations induced by gestational sepsis on the offspring at different stages of life

BackgroundThe term sepsis is used to designate a systemic condition of infection and inflammation associated with hemodynamic changes that result in organic dysfunction. Gestational sepsis can impair the development of the central nervous system and may promote permanent behavior alterations in the offspring. The aim of our work was to evaluate the effects of maternal sepsis on inflammatory cytokine levels and synaptic proteins in the hippocampus, neocortex, frontal cortex, and cerebellum of neonatal, young, and adult mice. Additionally, we analyzed the motor development, behavioral features, and cognitive impairments in neonatal, young and adult offspring.MethodsPregnant mice at the 14th embryonic day (E14) were intratracheally instilled with saline 0.9% solution (control group) or Klebsiella spp. (3 × 108 CFU) (sepsis group) and started on meropenem after 5 h. The offspring was sacrificed at postnatal day (P) 2, P8, P30, and P60 and samples of liver, lung, and brain were collected for TNF-α, IL-1β, and IL-6 measurements by ELISA. Synaptophysin, PSD95, and β-tubulin levels were analyzed by Western blot. Motor tests were performed at all analyzed ages and behavioral assessments were performed in offspring at P30 and P60.ResultsGestational sepsis induces a systemic pro-inflammatory response in neonates at P2 and P8 characterized by an increase in cytokine levels. Maternal sepsis induced systemic downregulation of pro-inflammatory cytokines, while in the hippocampus, neocortex, frontal cortex, and cerebellum an inflammatory response was detected. These changes in the brain immunity were accompanied by a reduction of synaptophysin and PSD95 levels in the hippocampus, neocortex, frontal cortex, and cerebellum, in all ages. Behavioral tests demonstrated motor impairment in neonates, and depressive-like behavior, fear-conditioned memory, and learning impairments in animals at P30 and P60, while spatial memory abilities were affected only at P60, indicating that gestational sepsis not only induces an inflammatory response in neonatal mouse brains, but also affects neurodevelopment, and leads to a plethora of behavioral alterations and cognitive impairments in the offspring.ConclusionThese data suggest that maternal sepsis may be causatively related to the development of depression, learning, and memory impairments in the litter.

Nutritional Intervention for Developmental Brain Damage: Effects of Lactoferrin Supplementation in Hypocaloric Induced Intrauterine Growth Restriction Rat Pups.

Introduction: Intrauterine Growth Restriction (IUGR) refers to an impaired development of the fetus and hence results in adverse neurodevelopmental and psychiatric consequences later in life. Lactoferrin (Lf) is a glycoprotein present in milk that has already shown neuroprotective effects through its anti-inflammatory and antioxidant properties on impaired developing brains. The aim of this study was to characterize a rat model of IUGR and assess the neuroprotective effect of a nutritional supplementation with bovine Lf during pregnancy and lactation on this model.Methods: A model of 50% gestational caloric restriction (CR) was used. Three groups were designed, and pregnant rats had either ad libitum access to food (control group, CTL) or 50% of the controls' intake (restricted group, IUGR). The diet was isocaloric and supplemented with bovine Lf for the caloric restricted dams (restricted-Lf, IUGR_Lf). At postnatal day 7 and 21, advanced ex-vivo diffusion MRI techniques at 9.4T were used to investigate brain cortical and white matter microstructure. Further, genes and proteins involved in structure (synaptophysin, MBP), microglia (Iba-1), metabolism (MCT2, βCaMKII) and apoptosis (Bcl-2) were analyzed in the cortex and striatum. In the cortex, the number of parvalbumin immunoreactive interneurons and their perineuronal nets were quantified. Behavioral tests were performed at P31.Results: Effects of the CR were significant in the cortex and striatum with reduction of synaptophysin (marker of synaptogenesis) at P7 and MBP (marker of myelin) at P21 in the cortex. Indeed, MCT2 (energy metabolism), Bcl-2 (anti-apoptotic protein) and βCaMKII (synapse activity) expressions were reduced in IUGR groups at P7. In the striatum NG2 (marker of oligodendrocyte precursor cells) and Bcl-2 at P7 as well as βCaMKII at P21 were decreased following IUGR and restored by Lf. Cortical microstructure was impaired following CR with partial effect of Lf. Lf prevented oxidative stress induced parvalbumin interneurons impairments whereas striatum and external capsule showed alterations in microstructure depicted by diffusion MRI, which were also partially reversed by Lf.Discussion and Conclusion: The model of 50% caloric restriction induced mild impairment partially reversed by nutritional intervention using Lf during pregnancy and lactation.

Sex-dependent behavioral impairments in the HdhQ350/+ mouse line

Huntington’s Disease (HD) is an autosomal dominant neurodegenerative disease characterized by gradual deterioration of motor and cognitive functions and development of psychiatric deficits. Animal models provide powerful means to study the pathological processes, molecular dysfunctions and symptoms associated with HD. We performed a longitudinal behavioral study of the newly developed HdhQ350/+ mouse line, a knock-in model that expresses a repeat of 350 glutamines. We found remarkable sex-dependent differences on symptom onset and severity. While both sexes lose weight and grip strength, only HdhQ350/+ males have impaired motor coordination as measured by the rotarod and alterations in gait as measured by the catwalk assay. While HdhQ350/+ females do not exhibit impairment in motor coordination, we found a reduction in dark phase locomotor activity. Male and female HdhQ350/+ mice do not show anxiety as measured by the elevated plus maze or changes in exploration as measured by the open field test. To investigate these sex-dependent differences, we performed western blot analyses of striatal tissue. We measured equal mutant huntingtin protein expression in both sexes and found evidence of aggregation. We found the expected decrease of DARPP-32 expression only in female HdhQ350/+ mice. Remarkably, we found no evidence of reduction in synaptophysin or CB1 receptors in HdhQ350/+ tissue of either sex. Our study indicates that male and female HdhQ350/+ mice differentially recapitulate select behavioral impairments commonly measured in other HD mouse models with limited sex-dependent changes in recognized histopathological markers. We conclude that expanded polyglutamine repeats influence HD pathogenesis in a sex-dependent manner.

Molecular evidence of synaptic pathology in the CA1 region in schizophrenia.

Alterations of postsynaptic density (PSD)95-complex proteins in schizophrenia ostensibly induce deficits in synaptic plasticity, the molecular process underlying cognitive functions. Although some PSD95-complex proteins have been previously examined in the hippocampus in schizophrenia, the status of other equally important molecules is unclear. This is especially true in the cornu ammonis (CA)1 hippocampal subfield, a region that is critically involved in the pathophysiology of the illness. We thus performed a quantitative immunoblot experiment to examine PSD95 and several of its associated proteins in the CA1 region, using post mortem brain samples derived from schizophrenia subjects with age-, sex-, and post mortem interval-matched controls (n=20/group). Our results indicate a substantial reduction in PSD95 protein expression (−61.8%). Further analysis showed additional alterations to the scaffold protein Homer1 (Homer1a: +42.9%, Homer1b/c: −24.6%), with a twofold reduction in the ratio of Homer1b/c:Homer1a isoforms (P=0.011). Metabotropic glutamate receptor 1 (mGluR1) protein levels were significantly reduced (−32.7%), and Preso, a protein that supports interactions between Homer1 or PSD95 with mGluR1, was elevated (+83.3%). Significant reduction in synaptophysin (−27.8%) was also detected, which is a validated marker of synaptic density. These findings support the presence of extensive molecular abnormalities to PSD95 and several of its associated proteins in the CA1 region in schizophrenia, offering a small but significant step toward understanding how proteins in the PSD are altered in the schizophrenia brain, and their relevance to overall hippocampal and cognitive dysfunction in the illness.

Synaptic protein levels altered in vascular dementia.

IntroductionCerebral ischaemia is the defining pathophysiological abnormality in most forms of vascular dementia (VAD), but the pathogenesis of the dementia remains poorly understood. In Alzheimer's disease (AD), there is early loss of synaptic proteins, but these have been little studied in VAD.Materials and MethodsWe measured synaptophysin, postsynaptic density protein 95 (PSD-95), drebrin, synaptosomal-associated protein 25 (SNAP-25) and vascular endothelial growth factor (VEGF) by enzyme-linked immunosorbent assays in superior temporal cortex from 11 patients with VAD and, initially, 11 non-dementia controls. We corrected for neuronal content by measurement of neuron-specific enolase. A further 11 controls were subsequently used in a validation study. Simulation of post-mortem delay found that PSD-95 was stable at 4°C but declined slightly at RT. SNAP-25 and drebrin showed good post-mortem stability. Previous studies had shown good post-mortem preservation of synaptophysin and VEGF.ResultsThe VAD cases had lower synaptophysin (but P > 0.05 in initial study), significantly lower SNAP-25 (P = 0.024) and significantly higher drebrin (P = 0.020). On comparison with the second control group, the reduction in synaptophysin was significant (P = 0.008), and the other results were confirmed.ConclusionThere is probably a reduction in presynaptic proteins in the temporal cortex in VAD, although not as marked as in AD. In VAD, there is also an increase in drebrin, which may be a response to reduced synaptic input.

Perforant path synaptic loss correlates with cognitive impairment and Alzheimer's disease in the oldest-old.

Alzheimer's disease, which is defined pathologically by abundant amyloid plaques and neurofibrillary tangles concurrent with synaptic and neuronal loss, is the most common underlying cause of dementia in the elderly. Among the oldest-old, those aged 90 and older, other ageing-related brain pathologies are prevalent in addition to Alzheimer's disease, including cerebrovascular disease and hippocampal sclerosis. Although definite Alzheimer's disease pathology can distinguish dementia from normal individuals, the pathologies underlying cognitive impairment, especially in the oldest-old, remain poorly understood. We therefore conducted studies to determine the relative contributions of Alzheimer's disease pathology, cerebrovascular disease, hippocampal sclerosis and the altered expression of three synaptic proteins to cognitive status and global cognitive function. Relative immunohistochemistry intensity measures were obtained for synaptophysin, Synaptic vesicle transporter Sv2 (now known as SV2A) and Vesicular glutamate transporter 1 in the outer molecular layer of the hippocampal dentate gyrus on the first 157 participants of 'The 90+ Study' who came to autopsy, including participants with dementia (n = 84), those with cognitive impairment but no dementia (n = 37) and those with normal cognition (n = 36). Thal phase, Braak stage, cerebrovascular disease, hippocampal sclerosis and Pathological 43-kDa transactive response sequence DNA-binding protein (TDP-43) were also analysed. All measures were obtained blind to cognitive diagnosis. Global cognition was tested by the Mini-Mental State Examinaton. Logistic regression analysis explored the association between the pathological measures and the odds of being in the different cognitive groups whereas multiple regression analyses explored the association between pathological measures and global cognition scores. No measure clearly distinguished the control and cognitive impairment groups. Comparing the cognitive impairment and dementia groups, synaptophysin and SV2 were reduced, whereas Braak stage, TDP-43 and hippocampal sclerosis frequency increased. Thal phase and VGLUT1 did not distinguish the cognitive impairment and dementia groups. All measures distinguished the dementia and control groups and all markers associated with the cognitive test scores. When all markers were analysed simultaneously, a reduction in synaptophysin, a high Braak stage and the presence of TDP-43 and hippocampal sclerosis associated with global cognitive function. These findings suggest that tangle pathology, hippocampal sclerosis, TDP-43 and perforant pathway synaptic loss are the major contributors to dementia in the oldest-old. Although an increase in plaque pathology and glutamatergic synaptic loss may be early events associated with cognitive impairment, we conclude that those with cognitive impairment, but no dementia, are indistinguishable from cognitively normal subjects based on the measures reported here.

The enduring effects of an adolescent social stressor on synaptic density, part II: Poststress reversal of synaptic loss in the cortex by adinazolam and MK‐801

Experience programs synaptic development to match the needs of the environment. This process depends on the nature and timing of the experience. Exposure to stress during adolescence selectively reduces synaptic density in the prefrontal cortex (a later maturing region), while sparing hippocampal synapses (an earlier maturing region). To determine whether the anatomical effects of an adolescent social stressor in rats endures into adulthood and are reversible, male subjects were isolation or group housed between days 30 and 35 and then treated with vehicle, adinazolam, MK-801, or tianeptine between days 40 and 55. At day 60, immunohistochemistry revealed a 13.5% +/- 5.3% reduction in synaptophysin in the infralimbic cortex and cingulate gyrus in isolation-housed subjects. MK-801 and adinazolam restored cortical synaptic density to within 2% of group-housed values, suggesting that the synaptic loss induced by stress during adolescence is modulated through reduced glutamatergic activity directly by NMDA antagonism or indirectly by enhancing GABAergic activity. Tianeptine did not modulate adolescent stress effects in the prefrontal cortex. None of these drugs increased cortical synaptophysin in group-housed controls. Increased synaptophysin was observed in the group-housed condition in the hippocampus, striatum, and nucleus accumbens following drug exposure. Although stress did not decrease synaptic density in these regions, drug exposure failed to increase synaptic density when compared with the controls. Taken together, stress-induced changes in cortical, but not hippocampal, synaptic density initiated during adolescence endure into adulthood. These cortical changes can be reversed through a reduction of glutamatergic activity, but not serotonin augmentation.

Down‐regulation of vesicular glutamate transporters precedes cell loss and pathology in Alzheimer's disease

Alzheimer's disease (AD) is characterized pathologically by plaques, tangles, and cell and synapse loss. As glutamate is the principle excitatory neurotransmitter of the CNS, the glutamatergic system may play an important role in AD. An essential step in glutamate neurotransmission is the concentration of glutamate into synaptic vesicles before release from the presynaptic terminal. Recently a group of proteins responsible for uptake has been identified - the vesicular glutamate transporters (VGLUTs). The generation of antibodies has facilitated the study of glutamatergic neurones. Here, we used antibodies to the VGLUTs together with immunohistochemistry and western blotting to investigate the status of glutamatergic neurones in temporal, parietal and occipital cortices of patients with AD; these regions were chosen to represent severely, moderately and mildly affected regions at the end stage of the disease. There was no change in expression of the synaptic markers in relation to total protein in the temporal cortex, but a significant reduction in synaptophysin and VGLUT1 was found in both the parietal and occipital cortices. These changes were found to relate to the number of tangles in the temporal cortex. There were no correlations with either mental test score or behaviour syndromes, with the exception of depression.

Expression of Vascular Endothelial Growth Factor and Its Receptors in the Central Nervous System in Amyotrophic Lateral Sclerosis

Vascular endothelial growth factor (VEGF) prolongs survival in the mutant SOD1 transgenic mouse model of amyotrophic lateral sclerosis (ALS), whereas dysregulation of VEGF through deletion of its hypoxia-regulatory element causes motor neuron degeneration in mice. We investigated the expression of VEGF and its major agonist receptors in the normal central nervous system and in patients with ALS. Immunohistochemistry demonstrated similar expression patterns of VEGF and VEGF receptor 2 (VEGFR2) in the spinal cord with finely punctate staining of the neuropil and strong expression in anterior horn cells (AHCs). Granular staining on the surface of some AHCs, similar to that seen with synaptic markers, suggested synaptic labeling. VEGFR2 staining was reduced in the neuropil of ALS cases (p=0.018) associated with a reduction of synaptophysin but not SNAP25 expression. A greater proportion of AHCs in ALS cases showed low expression of VEGF (p=0.006) and VEGFR2 (p=0.009) compared with controls. Expression of VEGF and VEGFR2 was confirmed by Western blotting and quantitative reverse transcriptase-polymerase chain reaction (QPCR). The similar expression patterns of VEGF and VEGFR2 suggests autocrine/paracrine effects on spinal motor neurons, and the reduction in their expression seen in ALS cases would support the hypothesis that, as in mouse models of the disease, reduced VEGF signaling may play a role in the pathogenesis of ALS.

Staying Connected: Synapses in Alzheimer Disease

Careful dissection is continuing to reveal the complexity and importance of synapse alterations in Alzheimer disease (AD). Synaptic alterations are the best biological correlate to the extent of cognitive loss1,2 and have become the gold standard of meaningful biomarkers of AD. Not neurofibrillary tangles (NFT), senile plaques, or even neuronal loss show such strong statistical correlation with dementia.2–4 Connections are the hallmark of neurons and without functional synapses, neurons are in “solitary.” Recent findings showing that amyloid-β (Aβ) affects and accumulates in synapses5 and that amyloid-β protein precursor (AβPP) over-expressing mice show synaptic alterations6,7 have provided additional evidence that synapses are a critical readout of the biology of AD. In this issue, Gylys and co-workers8 provide an elegant and precise dissection of synaptic alterations in AD using synaptosomal preparations. The results are both consistent with the synaptic alteration hypothesis proposed by Terry and colleagues2–4 and refine it. In previous studies synapses have been quantitated by synaptophysin immunoreactivity,1,9 a protein belonging to a subclass of synaptic vesicles. However, when synapses were quantitated by precise ultrastructural morphometric/stereological approaches, reduction in synapses was much less striking than the reduction in synaptophysin.10 Furthermore, synapse alterations included major remodeling of morphology11 as noted in some of the earliest studies of AD.12 These established findings lead one to consider whether the alterations in synapses in AD are limited to global reduction in vesicles rather than synapse changes. Gylys and collaborators8 used a broad range of pre- and postsynaptic markers to precisely analyze synaptosomes by flow cytometry. They found that the synaptosomes in AD show no reduction in synaptophysin and only a slight reduction in the overall number of synaptosomes. To account for the undoubted major reduction in synaptophysin while synapse number and synaptic vesicle number are only slightly reduced, brings to mind the seminal findings of Suzuki and Terry13 who reported substantial axonal transport abnormalities in AD. Consistently, one finds vesicle pile-up in cell bodies similar to transport blockage models.14 Recent findings that microtubules are greatly reduced in AD15 and that AβPP/Aβ play important roles in vesicular transport16–20 further link Terry’s transport and Gylys’ synaptic hypotheses. Not answered is where the extrasynaptic synaptophysin is located. One of the most attractive possibilities is that many of the synaptophysin vesicles are located in axons; for that reason, Terry’s initial insight to focus on vesicles was particularly fruitful because it marked the reduction most specific to AD. Additional work either by cell fractionation studies, as Gylys and collaborators8 have so elegantly performed, or by immunoelectron microscopy is called for to dissect this issue more clearly.