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Endolysosomal defects in neurons are central to the pathogenesis of prion and other neurodegenerative disorders. In prion disease, prion oligomers traffic through the multivesicular body (MVB) and are routed for degradation in lysosomes or for release in exosomes, yet how prions impact proteostatic pathways is unclear. We found that prion-affected human and mouse brain showed a marked reduction in Hrs and STAM1 (ESCRT-0), which route ubiquitinated membrane proteins from early endosomes into MVBs. To determine how the reduction in ESCRT-0 impacts prion conversion and cellular toxicityin vivo, we prion-challenged conditional knockout mice (male and female) havingHrsdeleted from neurons, astrocytes, or microglia. The neuronal, but not astrocytic or microglial, Hrs-depleted mice showed a shortened survival and an acceleration in synaptic derangements, including an accumulation of ubiquitinated proteins, deregulation of phosphorylated AMPA and metabotropic glutamate receptors, and profoundly altered synaptic structure, all of which occurred later in the prion-infected control mice. Finally, we found that neuronal Hrs (nHrs) depletion increased surface levels of the cellular prion protein, PrPC, which may contribute to the rapidly advancing disease through neurotoxic signaling. Taken together, the reduced Hrs in the prion-affected brain hampers ubiquitinated protein clearance at the synapse, exacerbates postsynaptic glutamate receptor deregulation, and accelerates neurodegeneration.SIGNIFICANCE STATEMENTPrion diseases are rapidly progressive neurodegenerative disorders characterized by prion aggregate spread through the central nervous system. Early disease features include ubiquitinated protein accumulation and synapse loss. Here, we investigate how prion aggregates alter ubiquitinated protein clearance pathways (ESCRT) in mouse and human prion-infected brain, discovering a marked reduction in Hrs. Using a prion-infection mouse model with neuronal Hrs (nHrs) depleted, we show that low neuronal Hrs is detrimental and markedly shortens survival time while accelerating synaptic derangements, including ubiquitinated protein accumulation, indicating that Hrs loss exacerbates prion disease progression. Additionally, Hrs depletion increases the surface distribution of prion protein (PrPC), linked to aggregate-induced neurotoxic signaling, suggesting that Hrs loss in prion disease accelerates disease through enhancing PrPC-mediated neurotoxic signaling.

The present work demonstrates an experience in estimating the threshold value of journey distances travelled by transit passengers using generalized polynomial function. The threshold value of journey distances may be defined as that distance beyond which passengers might no more be interested to travel by their reported mode. A knowledge on this threshold value is realized to be useful to limit the upper-most slab of transit fare, while preparing of a length-based fare matrix table. Theoretically, the threshold value can be obtained at that point on the cumulative frequency distribution (CFD) curve of journey distances at which the maximum rate of change of the slope of curve occurs. In this work, the CFD curve of the journey distance values is empirically modelled using Newton’s Polynomial Interpolation method, which helps to overcome various challenges usually encountered while an assumption of a theoretical probability distribution is considered a priori for the CFD.

Phosphorylation of α-synuclein at the Serine-129 site (α-syn Ser129P) is an established pathologic hallmark of synucleinopathies, and also a therapeutic target. In physiologic states, only a small fraction of total α-syn is phosphorylated at this site, and consequently, almost all studies to date have focused on putative pathologic roles of this post-translational modification. We noticed that unlike native (total) α-syn that is widely expressed throughout the brain, the overall pattern of α-syn Ser129P is restricted, suggesting intrinsic regulation and putative physiologic roles. Surprisingly, preventing phosphorylation at the Ser-129 site blocked the ability of α-syn to attenuate activity-dependent synaptic vesicle (SV) recycling – widely thought to reflect its normal function. Exploring mechanisms, we found that neuronal activity augments α-syn Ser-129P, and this phosphorylation is required for α-syn binding to VAMP2 and synapsin – two functional binding-partners that are necessary for α-syn function. AlphaFold2-driven modeling suggests a scenario where Ser129P induces conformational changes in the C-terminus that stabilizes this region and facilitates protein-protein interactions. Our experiments indicate that the pathology-associated Ser129P is an unexpected physiologic trigger of α-syn function, which has broad implications for pathophysiology and drug-development.

We have studied the viability and possible patterns of a strong first-order electroweak phase transition (SFOEWPT) within the $Z_3$-symmetric Next-to-Minimal Supersymmetric Standard Model (NMSSM), in view of the latest experimental results from the dark matter (DM) sector, Higgs sector and the searches of the lighter chargino and neutralinos at the Large Hadron Collider (LHC). We show that the region of parameter space with relatively small $\mu_\mathrm{eff}$ that favors an SFOEWPT has started to get excluded from the searches at the LHC and various DM experiments. However, there still remain phenomenologically much involved and compatible regions for an SFOEWPT that are yet not sensitive to the latest LHC and DM searches. We further estimate the production of stochastic gravitational waves (GW) from an SFOEWPT within and without the bag model and the prospects of detecting such signals at various future/proposed GW experiments., Comment: 6 pages, 1 figure, 2 tables, Proceedings for 41st International Conference on High Energy Physics (ICHEP2022, Bologna, Italy) based on JHEP06(2022)108 (arXiv:2202.12476)

We study the prospects of a simultaneous hunt at the Large Hadron Collider (LHC) of relatively light electroweakinos and a singlet-like scalar of the $Z_3$-symmetric Next-to-Minimal Supersymmetric Standard Model ($Z_3$-NMSSM) in the cascade decays of not so heavy ($\lesssim 1.5$ TeV) top squarks that are produced in pairs at the LHC which characteristically involve the singlet-like states. We work in a scenario where the lightest (next-to-lightest) SUSY particle is bino (singlino)-like with a mass below 100 GeV ($\gtrsim 100$ GeV), whereas a pair of immediately heavier neutralinos and the lighter chargino are higgsino-like with masses in the range $\sim 500$ GeV -- 1 TeV. Further, the singlet-like scalar present in the spectrum provides a funnel for a rapid enough mutual annihilation of the LSP thus making the latter meet the experimental upper bound on its relic abundance. The scenario is motivated by its ability to offer a bino-like dark matter with such a mass unlike what is now disfavored in the MSSM while avoiding the stringent lower bounds from the LHC experiments on the masses of these involved particles and still remaining reasonably `natural'. We find that while a usual cut-based analysis (CBA) on LHC data worth 300 $\text{fb}^{-1}$ would be unable to discover such excitations, a multivariate analysis (MVA) can be reasonably sensitive to higgsino-like electroweakinos having masses $\gtrsim 650$ GeV when the lighter top squark has a mass $\gtrsim 1$ TeV. On the other hand, with 3000 $\text{fb}^{-1}$ of data these masses become accessible in a CBA while even an MVA on such a data set is unlikely to find these electroweakinos with masses around 1 TeV when the mass of the lighter top squark hits $\sim 1.5$ TeV., 35 pages, 3 figures, 6 tables

Genome editing of somatic cells via clustered regularly interspaced short palindromic repeats (CRISPR) offers promise for new therapeutics to treat a variety of genetic disorders, including neurological diseases. However, the dense and complex parenchyma of the brain and the post-mitotic state of neurons make efficient genome editing challenging. In vivo delivery systems for CRISPR-Cas proteins and single guide RNA (sgRNA) include both viral vectors and non-viral strategies, each presenting different advantages and disadvantages for clinical application. We developed non-viral and biodegradable PEGylated nanocapsules (NCs) that deliver preassembled Cas9-sgRNA ribonucleoproteins (RNPs). Here, we show that the RNP NCs led to robust genome editing in neurons following intracerebral injection into the mouse striatum. Genome editing was predominantly observed in medium spiny neurons (>80%), with occasional editing in cholinergic, calretinin, and parvalbumin interneurons. Glial activation was minimal and was localized along the needle tract. Our results demonstrate that the RNP NCs are capable of safe and efficient neuronal genome editing in vivo.SIGNIFICANCE STATEMENTModifying the DNA of cells in the brain could present opportunities for new treatments of neurological diseases. In this report, we describe a nanocapsule system designed to deliver the elements needed to modify the DNA of brain cells, also known as genome editing. These nanocapsules are created by chemically encapsulating the genome editing components, such that the nanocapsules are stable when prepared and biodegradable to release their payload upon entering cells. When injected into the mouse brain, our research shows that the nanocapsules lead to safe and efficient editing of DNA in neurons.

The move from reading to writing the human genome offers new opportunities to improve human health. The United States National Institutes of Health (NIH) Somatic Cell Genome Editing (SCGE) Consortium aims to accelerate the development of safer and more-effective methods to edit the genomes of disease-relevant somatic cells in patients, even in tissues that are difficult to reach. Here we discuss the consortium’s plans to develop and benchmark approaches to induce and measure genome modifications, and to define downstream functional consequences of genome editing within human cells. Central to this effort is a rigorous and innovative approach that requires validation of the technology through third-party testing in small and large animals. New genome editors, delivery technologies and methods for tracking edited cells in vivo, as well as newly developed animal models and human biological systems, will be assembled—along with validated datasets—into an SCGE Toolkit, which will be disseminated widely to the biomedical research community. We visualize this toolkit—and the knowledge generated by its applications—as a means to accelerate the clinical development of new therapies for a wide range of conditions., This Perspective discusses how the Somatic Cell Genome Editing Consortium aims to accelerate the implementation of safe and effective genome-editing therapies in the clinic.

Gene therapy is making a comeback. With its twin promise of targeting disease etiology and 'long-term correction', gene-based therapies (defined here as all forms of genome manipulation) are particularly appealing for neurodegenerative diseases, for which conventional pharmacologic approaches have been largely disappointing. The recent success of a viral-vector-based gene therapy in spinal muscular atrophy-promoting survival and motor function with a single intravenous injection-offers a paradigm for such therapeutic intervention and a platform to build on. Although challenges remain, the newfound optimism largely stems from advances in the development of viral vectors that can diffusely deliver genes throughout the CNS, as well as genome-engineering tools that can manipulate disease pathways in ways that were previously impossible. Surely spinal muscular atrophy cannot be the only neurodegenerative disease amenable to gene therapy, and one can imagine a future in which the toolkit of a clinician will include gene-based therapeutics. The goal of this Review is to highlight advances in the development and application of gene-based therapies for neurodegenerative diseases and offer a prospective look into this emerging arena.

Genome editing of somatic cells via clustered regularly interspaced short palindromic repeats (CRISPR) offers promise for new therapeutics to treat a variety of genetic disorders, including neurological diseases. However, the dense and complex parenchyma of the brain and the post-mitotic state of neurons make efficient genome editing challenging. In vivo delivery systems for CRISPR-Cas proteins and single guide RNA (sgRNA) include both viral vectors and non-viral strategies, each presenting different advantages and disadvantages for clinical application. We developed non-viral and biodegradable PEGylated nanocapsules (NCs) that deliver preassembled Cas9-sgRNA ribonucleoproteins (RNPs). Here, we show that the RNP NCs led to robust genome editing in neurons following intracerebral injection into the healthy mouse striatum. Genome editing was predominantly observed in medium spiny neurons (80%), with occasional editing in cholinergic, calretinin, and parvalbumin interneurons. Glial activation was minimal and was localized along the needle tract. Our results demonstrate that the RNP NCs are capable of safe and efficient neuronal genome editing in vivo.

The polarized morphology of neurons necessitates the delivery of proteins synthesized in the soma along the length of the axon to distal synapses; critical for sustaining communication between neurons. This constitutive and dynamic process of protein transport along axons termed "axonal transport" was initially characterized by classic pulse-chase radiolabeling studies which identified two major rate components: a fast component and a slow component. Early radiolabeling studies indicated "cohesive co-transport" of slow transport cargos. However, this approach could not be used to visualize or provide mechanistic insights on this highly dynamic process. The advent of fluorescent and photoactivatable imaging probes have now enabled real-time imaging of axonal transport. Conventional fluorescent probes have helped visualize and characterize the molecular mechanisms of transport of vesicular proteins. These proteins typically move in the fast component of axonal transport and appear as "punctate structures" along axons. However, a large majority of transported proteins that move in the slow component of transport, typically show a "uniform diffusive glow" along axons when tagged to conventional fluorescent probes. This makes it challenging to unequivocally track them in real time. Our lab has used photoactivatable fluorescent probes to tag three individual cytosolic proteins moving in the slow component of axonal transport, and identified three distinct modes of transport along axons. Our data from these experiments argue against the prevailing hypothesis based on classic radiolabeling studies, which suggested that all slow-transport proteins may move along the axon as one large macromolecular protein complex. Although other labs have started using photoactivation to study axonal transport of cytosolic proteins, this technique remains largely under-utilized. Here, we describe the detailed protocols to image and analyze axonal transport of three typical slow-component cargoes along axons of cultured hippocampal neurons.

The article presents an investigation into selection of priority areas of intervention for improving travel condition of walk-accessed bus users within a bus stop catchment area. The work considers that propensity to use bus service by walk-accessed bus users is bi-directionally elastic to both the quality of walk-access facility, and the service characteristics of bus operation. In view of this, the current work attempts to consider two lists of attributes each from walk-accessed facility and from service characteristics of bus operation, and then users’ opinions on them are recorded in terms of perceived importance and satisfaction. The intervention areas are then identified by employing a two-dimensional analysis called Revised Importance Performance Analysis (R-IPA) using fuzzy-C means clustering. In this process, the factor structure and management scheme of the attributes are identified, and the list of intervention areas are finalized by comparing them. The attributes related to walk-access facility such as surface quality of sidewalk, interval between crosswalks, presence of dedicated sidewalk facility with adequate width, provision of adequate street lights and road surveillance are found to warrant immediate intervention. Similarly, the attributes related to service characteristics of bus operation such as frequency of bus service, in-vehicle journey time, and bus fare are also found to warrant immediate intervention.

Slow axonal transport conveys cytosolic and cytoskeletal proteins into axons and synapses at overall velocities that are several orders of magnitude slower than the fast transport of membranous organelles such as vesicles and mitochondria. The phenomenon of slow transport was characterized by in vivo pulse-chase radiolabeling studies done decades ago, and proposed models emphasized an orderly cargo-movement, with apparent cohesive transport of multiple proteins and subcellular structures along axons over weeks to months. However, visualization of cytosolic and cytoskeletal cargoes in cultured neurons at much higher temporal and spatial resolution has revealed an unexpected diversity in movement - ranging from a diffusion-like biased motion, to intermittent cargo dynamics and unusual polymerization-based transport paradigms. This review provides an updated view of slow axonal transport and explores emergent mechanistic themes in this enigmatic rate-class.

The article presents a work on the assessment of urban bus stop infrastructure in Bhubaneswar, and identifies improvement needs for the same as per bus passengers’ requirement and satisfaction. The work proposes a rationally appealing assessment framework, which uses perceived information of the bus passengers on various infrastructural items collected in scales of their importance and satisfaction level. In wake of this assessment, a set of key-factors are first identified using Exploratory Factor Analysis (EFA) on the stated importance ratings on an array of items, and then those key-factors are ordered by using Relative to an Identified Distribution Integral Transformation (RIDIT) analysis on stated satisfaction rating of the same survey items. The assessment framework takes an attempt to exploit the strength of these two types of dataset, while preparing the policy framework for improvement. The ordering of key-factors is carried out for two types of bus commuter such as captive users and choice users. The work documents new evidences in terms of improvement requirement of bus stop infrastructure in any urban area of emerging economies like India.

The work documents an experience in evaluation of the in-service infrastructural facilities of walk-access feeder paths to urban local bus stops in the city of Bhubaneswar, India. The work follows a rational approach, whereby both the in-service condition and relative importance of the walk-access infrastructures are considered, by taking into account the perception of those bus commuters, who usually undertake walk-mode to access bus stops. This approach allows determination of the relative priority among improvement areas of the existing walk-access infrastructural facilities, by focusing on those aspects where the perceived in-service condition is poor, but the importance is high. In view of this, a set of walk-access infrastructural items are identified existing in the study area, and bus commuters’ perception on their in-service condition and their importance is recorded. Thereafter, a two-dimensional fuzzy C-means algorithm is employed in order to identify the improvement areas. The work identifies a set of 5 major improvement areas of walk-access infrastructure, such as presence of dedicated sidewalk facility, width of sidewalk, presence of crosswalks, quality of road surface, and presence of street lights on access path. Overall, the work documents valuable insights in the context of infrastructural facility improvements in walk-access feeder paths with regard to a mid-sized urban area.

Loss of midbrain dopamine neurons causes the cardinal symptoms of Parkinson’s disease. However, not all dopamine neurons are equally vulnerable and a better understanding of the cell-type specific properties relating to selective dopamine neuron degeneration is needed. Most midbrain dopamine neurons express the vesicular glutamate transporter VGLUT2 during development and a subset continue to express low levels of VGLUT2 in adulthood, enabling the co-release of glutamate. Moreover, VGLUT2 expression in dopamine neurons can be neuroprotective since its genetic disruption was shown to sensitize dopamine neurons to neurotoxins. Here, we show that in response to toxic insult, and in two distinct models of alpha-synuclein stress, VGLUT2 dopamine neurons were resilient to degeneration. Dopamine neurons expressing VGLUT2 were enriched whether or not insult induced dopamine neuron loss, suggesting that while VGLUT2 dopamine neurons are more resilient, VGLUT2 expression can also be transcriptionally upregulated by injury. Finally, we observed that VGLUT2 expression was enhanced in surviving dopamine neurons from post-mortem Parkinson’s disease individuals. These data indicate that emergence of a glutamatergic identity in dopamine neurons may be part of a neuroprotective response in Parkinson’s disease.

International audience; In non-neuronal cells, clathrin has established roles in endocytosis, with clathrin cages enclosing plasma membrane infoldings, followed by rapid disassembly and reuse of monomers. However, in neurons, clathrin is conveyed in slow axonal transport over days to weeks, and the underlying transport/targeting mechanisms, mobile cargo structures, and even its precise presynaptic localization and physiologic role are unclear. Combining live imaging, photobleaching/conversion, mass spectrometry, electron microscopy, and super-resolution imaging, we found that unlike in dendrites, where clathrin cages rapidly assemble and disassemble, in axons, clathrin and related proteins organize into stable “transport packets” that are unrelated to endocytosis and move intermittently on microtubules, generating an overall slow anterograde flow. At synapses, multiple clathrin packets abut synaptic vesicle (SV) clusters, and clathrin packets also exchange between synaptic boutons in a microtubule-dependent “superpool.” Within synaptic boundaries, clathrin is surprisingly dynamic, continuously exchanging between local clathrin assemblies, and its depletion impairs SV recycling. Our data provide a conceptual framework for understanding clathrin trafficking and presynaptic targeting that has functional implications.

Chakrabarty et al. propose a model in which slow axonal transport of actin can occur by a biased polymerization of actin filaments along the axon shaft without the involvement of microtubules (MTs) or MT-based motors. These dynamics are distinct from polymer sliding—the canonical mechanism thought to convey cytoskeletal cargoes in slow transport., Classic pulse-chase studies have shown that actin is conveyed in slow axonal transport, but the mechanistic basis for this movement is unknown. Recently, we reported that axonal actin was surprisingly dynamic, with focal assembly/disassembly events (“actin hotspots”) and elongating polymers along the axon shaft (“actin trails”). Using a combination of live imaging, superresolution microscopy, and modeling, in this study, we explore how these dynamic structures can lead to processive transport of actin. We found relatively more actin trails elongated anterogradely as well as an overall slow, anterogradely biased flow of actin in axon shafts. Starting with first principles of monomer/filament assembly and incorporating imaging data, we generated a quantitative model simulating axonal hotspots and trails. Our simulations predict that the axonal actin dynamics indeed lead to a slow anterogradely biased flow of the population. Collectively, the data point to a surprising scenario where local assembly and biased polymerization generate the slow axonal transport of actin without involvement of microtubules (MTs) or MT-based motors. Mechanistically distinct from polymer sliding, this might be a general strategy to convey highly dynamic cytoskeletal cargoes.

A highly bino-like Dark Matter (DM), which is the Lightest Supersymmetric Particle (LSP), could be motivated by the stringent upper bounds on the DM direct detection rates. This is especially so when its mass is around or below 100 GeV for which such a bound tends to get most severe. Requiring not so large a higgsino mass parameter, that would render the scenario reasonably natural, prompts such a bino-like state to be relatively light. In the Minimal Supersymmetric Standard Model (MSSM), in the absence of comparably light scalars, such an excitation, if it has to be a thermal relic, is unable to meet the stringent experimental upper bound on its abundance unless its self-annihilation hits a funnel involving either the $Z$-boson or the Standard Model (SM)-like Higgs boson. We demonstrate that, in such a realistic situation, a highly bino-like DM of the popular $Z_3$-symmetric Next-to-Minimal Supersymmetric Standard Model (NMSSM) is viable over an extended range of its mass, from our targeted maximum in the vicinity of the mass of the top quark down to about 30 GeV. This is facilitated by the presence of comparably light singlet-like states that could serve as funnel (scalars) and/or coannihilating (singlino) states even as the bino-like LSP receives a minimal (but optimal) tempering triggered by suitably light higgsino states that, in the first place, evade stringent lower bounds on their masses that can be derived from the Large Hadron Collider (LHC) experiments only in the presence of a lighter singlino-like state. An involved set of blind spot conditions is derived for the DM direct detection rates by considering for the very first time the augmented system of neutralinos comprising of the bino, the higgsinos and the singlino which highlights the important roles played by the NMSSM parameters $\lambda$ and $\tan\beta$ in delivering a richer phenomenology., Comment: 70 pages, 16 figures and 3 tables

Animal identification is necessary for records, registration, and proof of ownership. The owner of few Black Bengal Goats can identify his goats by sight but it will create a problem for a larger number of goats as they are looking almost similar. A number of identification tools have been used for Black Bengal Goats like ear tag, tattoo, branding, RFID, etc. The Tattoos are permanent identification marking but inconvenient to read after a few months or years. Most of the farmers and breeders have used ear tags, which contain a number for identification of particular goat but may be lost at the time of grazing. Some organized farmers have placed RFID chips in tags but RFID reader is necessary to read the content of chips. In this paper, an effort has been made to identify individual Black Bengal Goat using their iris image like a human. The eye images have been captured preprocessed, enhanced, and irises have been segmented. The template has been generated from each segmented iris and stored in the database. The matching has been performed among different segmented iris images from the same goat and also been performed among iris images captured from different goats. It has been observed that the average Hamming distance among iris images captured at different times from the same goat are different from the average hamming distances among iris images from other goats. Finally, the matching threshold has been decided for the identification of Black Bengal Goat.

In this paper, individual light coloured pig (Yorkshire) has been recognized based on their ear images captured using mobile phones. The ears have been kept parallel to the light source and the images have been captured from the opposite side of the light source. The auricular venation pattern from each captured ear image has been extracted, the template has been generated and stored in a database. The templates of recaptured ear images have been matched with the stored templates of the same pig using average Euclidean distance. The pig has been verified if average Euclidean distances of matching are \({

Convolutional Neural Network (CNN) is widely used as a strong framework for object classification. This framework is suitable for classification of group of objects which carry similar features like animal breeds, plant varieties, etc. In this paper, an attempt has been made to build a model of identifying individual Ghungroo pig (one of the most popular registered pig breeds in India) based on its face image using CNN. Individual Ghungroo pigs look very similar, and it is a very challenging task even for humans to identify them from a group of Ghungroo pigs. Ten Ghungroo pigs from ICAR-Indian Veterinary Research Institute (IVRI) Animal Farm Kalyani (Block B, Kanchrapara, West Bengal 741235) have been captured using a DSLR camera. The pigs have been captured in natural environment without any restriction. The Inception-v3 model with transfer learning has been used to build the individual pig identification model. The captured images have been separated with 90%:10% into training images and test images sets, respectively. It has been observed that the model has predicted individual pigs with 100% accuracy at 0.87 confidence level.

Now days electricity has become the most demandable thing for the human beings. Due to continuous depletion of the conventional energy resources the demand for the use of nonconventional energy resources has increased. In the present paper we have used non-conventional energy resources i.e. solar energy and wind energy for generating uninterrupted power supply for the consumers. This paper comprises of combination of two sources of energy that will provide uninterrupted power supply to the system. Solar panels and wind turbines together have been used for converting the respective energies to the electrical energy. In this paper generation of electrical energy by combining two non-renewable sources at minimal cost and by not affecting the natural environment.

The normal function of α-synuclein (α-syn) remains elusive. Although recent studies suggest α-syn as a physiologic attenuator of synaptic vesicle (SV) recycling, mechanisms are unclear. Here, we show that synapsin—a cytosolic protein with known roles in SV mobilization and clustering—is required for presynaptic functions of α-syn. Our data offer a critical missing link and advocate a model where α-syn and synapsin cooperate to cluster SVs and attenuate recycling.

Alzheimer's disease (AD) is characterized by the accumulation of insoluble deposits of Amyloid β (Aβ) in brains. Aβ is derived by sequential cleavage of the amyloid precursor protein (APP) by β-site secretase enzyme (BACE-1) and γ-secretase. Proteolytic processing of APP by BACE-1 is the rate-limiting step in Aβ production, and this pathway is a prime target for AD drug development. Both APP and BACE-1 are membrane-spanning proteins, transported via secretory and endocytic pathways; and the physical interaction of APP and BACE-1 during trafficking is a key cell biological event initiating the amyloidogenic pathway. Here, we highlight recent research on intracellular trafficking/sorting of APP and BACE-1, and discuss how dysregulation of these pathways might lead to enhanced convergence of APP and BACE-1, and subsequent β-cleavage of APP. © 2018 Wiley Periodicals, Inc. Develop Neurobiol 78: 340-347, 2018.

High levels of amyloid-β peptide (Aβ) have been related to Alzheimer9s disease pathogenesis. However, in the healthy brain, low physiologically relevant concentrations of Aβ are necessary for long-term potentiation (LTP) and memory. Because cGMP plays a key role in these processes, here we investigated whether the cyclic nucleotide cGMP influences Aβ levels and function during LTP and memory. We demonstrate that the increase of cGMP levels by the phosphodiesterase-5 inhibitors sildenafil and vardenafil induces a parallel release of Aβ due to a change in the approximation of amyloid precursor protein (APP) and the β-site APP cleaving enzyme 1. Moreover, electrophysiological and behavioral studies performed on animals of both sexes showed that blocking Aβ function, by using anti-murine Aβ antibodies or APP knock-out mice, prevents the cGMP-dependent enhancement of LTP and memory. Our data suggest that cGMP positively regulates Aβ levels in the healthy brain which, in turn, boosts synaptic plasticity and memory. SIGNIFICANCE STATEMENT Amyloid-β (Aβ) is a key pathogenetic factor in Alzheimer9s disease. However, low concentrations of endogenous Aβ, mimicking levels of the peptide in the healthy brain, enhance hippocampal long-term potentiation (LTP) and memory. Because the second messenger cGMP exerts a central role in LTP mechanisms, here we studied whether cGMP affects Aβ levels and function during LTP. We show that cGMP enhances Aβ production by increasing the APP/BACE-1 convergence in endolysosomal compartments. Moreover, the cGMP-induced enhancement of LTP and memory was disrupted by blockade of Aβ, suggesting that the physiological effect of the cyclic nucleotide on LTP and memory is dependent upon Aβ.

Using proteomics, live microscopy, and superresolution microscopy, Ganguly et al. offer insight into the molecular composition of cytosolic cargo complexes conveyed in slow axonal transport, identifying the heat shock protein Hsc70 as a major regulator of this transport., Soluble cytosolic proteins vital to axonal and presynaptic function are synthesized in the neuronal soma and conveyed via slow axonal transport. Our previous studies suggest that the overall slow transport of synapsin is mediated by dynamic assembly/disassembly of cargo complexes followed by short-range vectorial transit (the “dynamic recruitment” model). However, neither the composition of these complexes nor the mechanistic basis for the dynamic behavior is understood. In this study, we first examined putative cargo complexes associated with synapsin using coimmunoprecipitation and multidimensional protein identification technology mass spectrometry (MS). MS data indicate that synapsin is part of a multiprotein complex enriched in chaperones/cochaperones including Hsc70. Axonal synapsin–Hsc70 coclusters are also visualized by two-color superresolution microscopy. Inhibition of Hsc70 ATPase activity blocked the slow transport of synapsin, disrupted axonal synapsin organization, and attenuated Hsc70–synapsin associations, advocating a model where Hsc70 activity dynamically clusters cytosolic proteins into cargo complexes, allowing transport. Collectively, our study offers insight into the molecular organization of cytosolic transport complexes and identifies a novel regulator of slow transport.

The function of α-synuclein (α-syn) has been long debated, and two seemingly divergent views have emerged. In one, α-syn binds to VAMP2, acting as a SNARE chaperone—but with no effect on neurotransmission—while another posits that α-syn attenuates neurotransmitter release by restricting synaptic vesicle mobilization and recycling. Here, we show that α-syn–VAMP2 interactions are necessary for α-syn–induced synaptic attenuation. Our data connect divergent views and suggest a unified model of α-syn function.

Bubble column reactors are common in many industries such as biochemical, energy generation, process industry etc. The performance of the reactor is dependent on the flow regime inside the column. Identification of the flow regimes inside the reactor is significant and a challenging task. In the present paper, non-intrusive, real time approaches for regime identification are developed, based on the image processing technique and Symbolic Dynamic Filtering (SDF) in MATLAB. Flow images are captured using a Photron high speed (1000 fps) camera. The bubble detection algorithm results in the calculation of bubble diameters and the distribution of the bubble diameters for different flow rates, leading to regime identification. The algorithm includes the method of calculation of the optimum threshold luminescence value and converting the input image into binary using that optimum threshold value. From the binary image, boundaries are traced and the average bubble diameter is calculated. The deviation from one regime to another regime is detected by SDF algorithm. Flow measure (μ) is a parameter obtained from SDF, which indicates the deviation of a particular flow from the reference flow. The variation of flow measure is plotted against the air flow rate. It is found that change in the slope of this curve is significant during regime transition. This parameter can be used as a marker of flow regime identification.

The article presents a method for ranking various catchment areas of an urban area according to their current service condition of the walk environment and their improvement needs. The stud...

The brain represents one of the most divergent and critical organs in the human body. Yet, it can be afflicted by a variety of neurodegenerative diseases specifically linked to aging, about which we lack a full biomolecular understanding of onset and progression, such as Alzheimer's disease (AD). Here we provide a proteomic resource comprising nine anatomically distinct sections from three aged individuals, across a spectrum of disease progression, categorized by quantity of neurofibrillary tangles. Using state-of-the-art mass spectrometry, we identify a core brain proteome that exhibits only small variance in expression, accompanied by a group of proteins that are highly differentially expressed in individual sections and broader regions. AD affected tissue exhibited slightly elevated levels of tau protein with similar relative expression to factors associated with the AD pathology. Substantial differences were identified between previous proteomic studies of mature adult brains and our aged cohort. Our findings suggest considerable value in examining specifically the brain proteome of aged human populations from a multiregional perspective. This resource can serve as a guide, as well as a point of reference for how specific regions of the brain are affected by aging and neurodegeneration.

α-Synuclein overexpression and aggregation are linked to Parkinson’s disease (PD), dementia with Lewy bodies (DLB), and several other neurodegenerative disorders. In addition to effects in the cell body, α-synuclein accumulation occurs at presynapses where the protein is normally localized. While it is generally agreed that excess α-synuclein impairs synaptic vesicle trafficking, the underlying mechanisms are unknown. We show here that acute introduction of excess human α-synuclein at a classic vertebrate synapse, the lamprey reticulospinal synapse, selectively impaired the uncoating of clathrin-coated vesicles (CCVs) during synaptic vesicle recycling, leading to a severe depletion of synaptic vesicles. Furthermore, human α-synuclein and lamprey γ-synuclein both interact in vitro with Hsc70, the chaperone protein that uncoats CCVs at synapses. After introducing excess α-synuclein to lamprey axons, Hsc70 availability was reduced at the synapses, suggesting Hsc70 sequestration as a possible mechanism underlying the synaptic vesicle trafficking defects. In support of this hypothesis, increasing the levels of exogenous Hsc70 together with α-synuclein ameliorated the CCV uncoating and vesicle recycling defects. These experiments identify a reduction in Hsc70 availability at synapses, and consequently its function, as the mechanism by which α-synuclein induces synaptic vesicle recycling defects. To our knowledge, this is the first report of a viable chaperone-based strategy for reversing the toxic impacts of excess α-synuclein at synapses, which may be of value for ameliorating synaptic defects in PD and other synuclein-linked diseases.SIGNIFICANCE STATEMENTSynaptic defects caused by α-synuclein overexpression are linked to cognitive deficits in PD and other diseases. However, the mechanisms by which excess α-synuclein impairs synaptic vesicle trafficking are unknown. Data presented here demonstrate that acute introduction of excess α-synuclein at a classical vertebrate synapse selectively inhibits CCV uncoating, leading to impaired vesicle recycling. Furthermore, increasing α-synuclein reduced synaptic levels of Hsc70, the clathrin uncoating ATPase. Subsequently increasing Hsc70 restored CCV uncoating and improved vesicle recycling. This study identifies a novel molecular mechanism underlying the α-synuclein-induced synaptic defects and presents one viable strategy for reversing them.

Cleavage of amyloid precursor protein (APP) by BACE-1 (β-site APP cleaving enzyme-1) is the rate-limiting step in amyloid-β (Aβ) production and a neuropathologic hallmark of Alzheimer's disease; thus, physical approximation of this substrate-enzyme pair is a crucial event with broad biological and therapeutic implications. Despite much research, neuronal locales of APP and BACE-1 convergence and APP cleavage remain unclear. Here we report an optical assay, based on fluorescence complementation, for visualizing in cellulo APP-BACE-1 interactions as a simple on/off signal. Combining this with other assays tracking the fate of internalized APP in hippocampal neurons, we found that APP and BACE-1 interacted in both biosynthetic and endocytic compartments, particularly along recycling microdomains such as dendritic spines and presynaptic boutons. In axons, APP and BACE-1 were cotransported, and they also interacted during transit. Finally, our assay revealed that the Alzheimer's disease-protective 'Icelandic' mutation greatly attenuates APP-BACE-1 interactions, suggesting a mechanistic basis for protection. Collectively, the data challenge canonical models and provide concrete insights into long-standing controversies in the field.

CRISPR/Cas9 guided gene-editing is a potential therapeutic tool, however application to neurodegenerative disease models has been limited. Moreover, conventional mutation correction by gene-editing would only be relevant for the small fraction of neurodegenerative cases that are inherited. Here we introduce a CRISPR/Cas9-based strategy in cell and animal models to edit endogenous amyloid precursor protein (APP) at the extreme C-terminus and reciprocally manipulate the amyloid pathway, attenuating APP-β-cleavage and Aβ production, while up-regulating neuroprotective APP-α-cleavage. APP N-terminus and compensatory APP-homologues remain intact, with no apparent effects on neurophysiology in vitro. Robust APP-editing is seen in human iPSC-derived neurons and mouse brains with no detectable off-target effects. Our strategy likely works by limiting APP and BACE-1 approximation, and we also delineate mechanistic events that abrogates APP/BACE-1 convergence in this setting. Our work offers conceptual proof for a selective APP silencing strategy., Gene editing strategies are typically designed to correct mutant genes, but most neurodegenerative diseases are sporadic. Here the authors describe a strategy to selectively edit the C-terminus of APP and attenuate amyloid-β production, while upregulating neuroprotective α-cleavage.

The actin cytoskeleton in neurons plays critical roles in axonal growth and synaptic organization. Until recently, most studies on axonal actin were limited to terminal growth cones or synapses, whereas the organization of actin along the shaft of the axon was relatively ignored. However, experiments using super-resolution microscopy and live imaging have revealed previously unknown actin structures along the axonal shaft, such as periodic ‘actin rings’ circumferentially wrapping underneath the plasma membrane and dynamic actin pools deeper within the axon shaft (termed actin ‘hotspots’ and ‘trails’). In this short review, we highlight some open questions that have surfaced as a direct result of these discoveries.