These findings establish minocycline as a promising therapeutic for the treatment of fragile X mental retardation.

Background: An ELISA was developed to determine the role of apoE/A␤ on soluble A␤ accumulation. Results: In AD transgenic mouse brain and human synaptosomes and CSF, levels of soluble apoE/A␤ are lower and oligomeric A␤ levels are higher with APOE4 and AD. Conclusion: Isoform-specific apoE/A␤ levels modulate soluble oligomeric A␤ levels. Significance: ApoE/A␤ and oligomeric A␤ represent a mechanistic approach to AD biomarkers.

SummaryMicroglial activation and macrophage infiltration into the CNS are common features of CNS autoimmune disease and of chronic neurodegenerative diseases. Because these cells largely express an overlapping set of common macrophage markers, it has been difficult to separate their respective contributions to disease onset and progression. This problem is further confounded by the many types of macrophages that have been termed microglia. Several approaches, ranging from molecular profiling of isolated cells to the generation of irradiation chimeric rodent models, are now beginning to generate rudimentary definitions distinguishing the various types of microglia and macrophages found within the CNS and the potential roles that these cells may play in health and disease.

The microtubule-associated protein tau has primarily been associated with axonal location and function; however, recent work shows tau release from neurons and suggests an important role for tau in synaptic plasticity. In our study, we measured synaptic levels of total tau using synaptosomes prepared from cryopreserved human postmortem Alzheimer's disease (AD) and control samples. Flow cytometry data show that a majority of synaptic terminals are highly immunolabeled with the total tau antibody (HT7) in both AD and control samples. Immunoblots of synaptosomal fractions reveal increases in a 20 kDa tau fragment and in tau dimers in AD synapses, and terminal-specific antibodies show that in many synaptosome samples tau lacks a C-terminus. Flow cytometry experiments to quantify the extent of C-terminal truncation reveal that only 15-25% of synaptosomes are positive for intact C-terminal tau. Potassium-induced depolarization demonstrates release of tau and tau fragments from presynaptic terminals, with increased release from AD compared to control samples. This study indicates that tau is normally highly localized to synaptic terminals in cortex where it is well-positioned to affect synaptic plasticity. Tau cleavage may facilitate tau aggregation as well as tau secretion and propagation of tau pathology from the presynaptic compartment in AD.

Amyloid-b (Ab) and hyperphosphorylated tau (p-tau) aggregates form the two discrete pathologies of Alzheimer disease (AD), and oligomeric assemblies of each protein are localized to synapses. To determine the sequence by which pathology appears in synapses, Ab and p-tau were quantified across AD disease stages in parietal cortex. Nondemented cases with high levels of AD-related pathology were included to determine factors that confer protection from clinical symptoms. Flow cytometric analysis of synaptosome preparations was used to quantify Ab and p-tau in large populations of individual synaptic terminals. Soluble Ab oligomers were assayed by a single antibody sandwich enzyme-linked immunosorbent assay. Total in situ Ab was elevated in patients with early-and late-stage AD dementia, but not in high pathology nondemented controls compared with age-matched normal controls. However, soluble Ab oligomers were highest in early AD synapses, and this assay distinguished early AD cases from high pathology controls. Overall, synapse-associated p-tau did not increase until late-stage disease in human and transgenic rat cortex, and p-tau was elevated in individual Ab-positive synaptosomes in early AD. These results suggest that soluble oligomers in surviving neocortical synaptic terminals are associated with dementia onset and suggest an amyloid cascade hypothesis in which oligomeric Ab drives phosphorylated tau accumulation and synaptic spread. These results indicate that antiamyloid therapies will be less effective once p-tau pathology is developed. (Am J Pathol 2016, 186: 185e198; http://dx.doi.org/10.1016/j.ajpath.2015 A large body of evidence indicates that soluble oligomers of amyloid-b (Ab) are the primary toxic peptides that initiate downstream tau pathology in the amyloid cascade hypothesis of Alzheimer disease (AD). 1,2 However, the time course and severity of AD dementia have been generally found to correlate with neurofibrillary tangle development rather than plaque appearance, 3e8 although a few studies have linked plaques with early cognitive decline. 9e12 Soluble oligomeric

Long-term potentiation (LTP) is an activity-dependent and persistent increase in synaptic transmission. Currently available techniques to measure LTP are time-intensive and require highly specialized expertise and equipment, and thus are not well suited for screening of multiple candidate treatments, even in animal models. To expand and facilitate the analysis of LTP, here we use a flow cytometry-based method to track chemically induced LTP by detecting surface AMPA receptors in isolated synaptosomes: fluorescence analysis of single-synapse long-term potentiation (FASS-LTP). First, we demonstrate that FASS-LTP is simple, sensitive, and models electrically induced LTP recorded in intact circuitries. Second, we conducted FASS-LTP analysis in two well-characterized Alzheimer's disease (AD) mouse models (3xTg and Tg2576) and, importantly, in cryopreserved human AD brain samples. By profiling hundreds of synaptosomes, our data provide the first direct evidence to support the idea that synapses from AD brain are intrinsically defective in LTP. Third, we used FASS-LTP for drug evaluation in human synaptosomes. Testing a panel of modulators of cAMP and cGMP signaling pathways, FASS-LTP identified vardenafil and Bay-73-6691 (phosphodiesterase-5 and -9 inhibitors, respectively) as potent enhancers of LTP in synaptosomes from AD cases. These results indicate that our approach could provide the basis for protocols to study LTP in both healthy and diseased human brains, a previously unattainable goal.

Dendritic spines are protrusions from the dendritic shaft that host most excitatory synapses in the brain. Although they first emerge during neuronal maturation, dendritic spines remain plastic through adulthood, and recent advances in the molecular mechanisms governing spine morphology have shown them to be exquisitely sensitive to changes in the micro-environment. Among the many factors affecting spine morphology are components and regulators of the extracellular matrix (ECM). Modification of the ECM is critical to the repair of injuries throughout the body, including the CNS. Matrix metalloproteinase (MMP)-7/matrilysin is a key regulator of the ECM during pathogen infection, after nerve crush and in encephalitogenic disorders. We have investigated the effects of MMP-7 on dendritic spines in hippocampal neuron cultures and found that it induces the transformation of mature, short mushroom-shaped spines into long, thin filopodia reminiscent of immature spines. These changes were accompanied by a dramatic redistribution of F-actin from spine heads into thick, rope-like structures in the dendritic shaft. Strikingly, MMP-7 effects on dendritic spines were similar to those of NMDA treatment, and both could be blocked by channel-specific antagonists. These findings are the first direct evidence that MMPs can influence the morphology of mature dendritic spines, and hence synaptic stability.

Much evidence indicates that soluble amyloid beta (Aβ) oligomers are key mediators of early cognitive loss, but the localization and key peptide species remain unclear. We have used flow cytometry analysis to demonstrate that surviving Alzheimer's disease (AD) synapses accumulate both Aβ and p-tau. The present experiments use peptide-specific xMAP assays and Western blotting to identify the Aβ peptide species in synaptosome-enriched samples from normal human subjects, neurologic controls, and AD cases. Aβ40 peptide levels did not vary, but both Aβ42 and Aβ oligomers were increased in soluble AD extracts, with oligomer levels 20-fold higher in aqueous compared to detergent extracts. In Western blots, a ladder of SDS-stable oligomers was observed in AD cases, varying in size from monomer, the major peptide observed, to larger assemblies up to about 200 kD and larger. Multiple oligomers, including monomer, small oligomers, a 56 kD assembly, and APP were correlated with the Aβ level measured in flow cytomety-purified synaptosomes. These results suggest that multiple APP processing pathways are active in AD synapses and multiple soluble oligomeric assemblies may contribute to synaptic dysfunction.