How do neurons age? A focused review on the aging of the microtubular cytoskeleton.

Aging is the leading risk factor for Alzheimer's disease and other neurodegenerative diseases. We now understand that a breakdown in the neuronal cytoskeleton, mainly underpinned by protein modifications leading to the destabilization of microtubules, is central to the pathogenesis of Alzheimer's disease. This is accompanied by morphological defects across the somatodendritic compartment, axon, and synapse.

Functional effects of haemoglobin can be rescued by haptoglobin in an in vitro model of subarachnoid haemorrhage.

During subarachnoid haemorrhage, a blood clot forms in the subarachnoid space releasing extracellular haemoglobin (Hb), which causes oxidative damage and cell death in surrounding tissues. High rates of disability and cognitive decline in SAH survivors are attributed to loss of neurons and functional connections during secondary brain injury. Haptoglobin sequesters Hb for clearance, but this scavenging system is overwhelmed after a haemorrhage.

A luminescence-based reporter to study tau secretion reveals overlapping mechanisms for the release of healthy and pathological tau.

In Alzheimer's disease, tau pathology is thought to spread via a prion-like manner along connected neuronal networks. For this to occur, the usually cytosolic tau protein must be secreted via an unconventional mechanism prior to uptake into the connected neuron. While secretion of healthy and pathological tau has been documented, it remains under-investigated whether this occurs via overlapping or distinct processes.

Stable Isotope Labeling by Amino Acids and Bioorthogonal Noncanonical Amino Acid Tagging in Cultured Primary Neurons.

Cultured primary neurons are a well-established model for the study of neuronal function. Conventional stable isotope labeling with amino acids in cell culture (SILAC) requires nearly complete metabolic labeling of proteins and therefore is difficult to apply to cultured primary neurons, which do not divide in culture. In a multiplex SILAC strategy, two different sets of heavy amino acids are used for labeling cells for the different experimental conditions.

Molecular Investigation of the Unfolded Protein Response in Select Human Tauopathies.

Background: Tauopathies are a group of neurodegenerative diseases associated with the accumulation of misfolded tau protein. The mechanisms underpinning tau-dependent proteinopathy remain to be elucidated. A protein quality control pathway within the endoplasmic reticulum, the unfolded protein response (UPR), has been suggested as a possible pathway modulating cellular responses in a range of neurodegenerative diseases, including those associated with misfolded cytosolic tau.

Co-culture of Murine Neurons Using a Microfluidic Device for The Study of Tau Misfolding Propagation.

The deposition of misfolded, aggregated tau protein is a hallmark of several neurodegenerative diseases, collectively termed "tauopathies". Tau pathology spreads throughout the brain along connected pathways in a prion-like manner. The process of tau pathology propagation across circuits is a focus of intense research and has been investigated in human post-mortem brain and in mouse models of the diseases, in diverse cellular systems including primary neurons, and in cell free assays using purified recombinant tau protein.

A 5' UTR GGN repeat controls localisation and translation of a potassium leak channel mRNA through G-quadruplex formation.

RNA G-quadruplexes (G4s) are secondary structures proposed to function as regulators of post-transcriptional mRNA localisation and translation. G4s within some neuronal mRNAs are known to control distal localisation and local translation, contributing to distinct local proteomes that facilitate the synaptic remodelling attributed to normal cellular function. In this study, we characterise the G4 formation of a (GGN)13 repeat found within the 5' UTR of the potassium 2-pore domain leak channel Task3 mRNA.

Quantitative and correlative extreme ultraviolet coherent imaging of mouse hippocampal neurons at high resolution.

Microscopy with extreme ultraviolet (EUV) light can provide many advantages over optical, hard x-ray or electron-based techniques. However, traditional EUV sources and optics have large disadvantages of scale and cost. Here, we demonstrate the use of a laboratory-scale, coherent EUV source to image biological samples-mouse hippocampal neurons-providing quantitative phase and amplitude transmission information with a lateral resolution of 80 nm and an axial sensitivity of ~1 nm.

Age-Dependent Changes in Synaptic NMDA Receptor Composition in Adult Human Cortical Neurons.

The molecular processes underlying the aging-related decline in cognitive performance and memory observed in humans are poorly understood. Studies in rodents have shown a decrease in N-methyl-D-aspartate receptors (NMDARs) that contain the GluN2B subunit in aging synapses, and this decrease is correlated with impaired memory functions. However, the age-dependent contribution of GluN2B-containing receptors to synaptic transmission in human cortical synapses has not been previously studied.

Tau Misfolding Efficiently Propagates between Individual Intact Hippocampal Neurons.

Neurofibrillary tangles, formed of misfolded, hyperphosphorylated tau protein, are a pathological hallmark of several neurodegenerations, including Alzheimer's disease. Tau pathology spreads between neurons and propagates misfolding in a prion-like manner throughout connected neuronal circuits. Tauopathy is accompanied by significant neuronal death, but the relationships between initial tau misfolding, propagation across connected neurons and cytotoxicity remain unclear.

Proteomic Approaches to Dissect Neuronal Signalling Pathways.

With an increasing awareness of mental health issues and neurological disorders, "understanding the brain" is one of the biggest current challenges in biological research. This has been recognised by both governments and funding agencies, and it includes the need to understand connectivity of brain regions and coordinated network activity, as well as cellular and molecular mechanisms at play. In this chapter, we will describe how we have taken advantage of different proteomic techniques to unravel molecular mechanisms underlying two modulators of neuronal function: Neurotrophins and antipsychotics.

A primate-specific short GluN2A-NMDA receptor isoform is expressed in the human brain.

Glutamate receptors of the N-methyl-D-aspartate (NMDA) family are coincident detectors of pre- and postsynaptic activity, allowing Ca influx into neurons. These properties are central to neurological disease mechanisms and are proposed to be the basis of associative learning and memory. In addition to the well-characterised canonical GluN2A NMDAR isoform, large-scale open reading frames in human tissues had suggested the expression of a primate-specific short GluN2A isoform referred to as GluN2A-S.

Pathogenic tau does not drive activation of the unfolded protein response.

The unfolded protein response (UPR) is commonly associated with a range of neurodegenerative diseases, and targeting UPR components has been suggested as a therapeutic strategy. The UPR surveys protein folding within the endoplasmic reticulum. However, many of the misfolded proteins that accumulate in neurodegeneration are localized so that they do not directly cause endoplasmic reticulum triggers that activate this pathway.

Asymmetric confinement for defining outgrowth directionality.

Directional connectivity is required to develop accurate in vitro models of the nervous system. This research investigated the interaction of murine neuronal outgrowths with asymmetric microstructured geometries to provide insights into the mechanisms governing unidirectional outgrowth bias. The structures were designed using edge-guidance and critical turning angle principles to study different prohibitive to permissive edge-guidance ratios.

Emergence of synaptic and cognitive impairment in a mature-onset APP mouse model of Alzheimer's disease.

The synaptic changes underlying the onset of cognitive impairment in Alzheimer's disease (AD) are poorly understood. In contrast to the well documented inhibition of long-term potentiation (LTP) in CA3-CA1 synapses by acute Aβ application in adult neurons from rodents, young amyloid precursor protein (APP) transgenic mouse models often, surprisingly, show normal LTP. This suggests that there may be important differences between mature-onset and developmental-onset APP expression/ Aβ accumulation and the ensuing synaptic and behavioural phenotype.

Minimalistic in vitro systems for investigating tau pathology.

Neurofibrillary tangles, formed of hyperphosphorylated, misfolded tau accumulations, are a pathological hallmark of neurodegenerative diseases such as Alzheimer's disease (AD) and frontotemporal dementia. The neuroanatomical localisation of tau pathology in AD brains of different disease stages suggests that tau tangle pathology is spreading throughout the brain along connected neuronal circuits. Pathogenic tau can act as a prion-like seed, inducing the misfolding of native tau and leading to disease propagation throughout the brain.

Mechano-sensitization of mammalian neuronal networks through expression of the bacterial large-conductance mechanosensitive ion channel.

Development of remote stimulation techniques for neuronal tissues represents a challenging goal. Among the potential methods, mechanical stimuli are the most promising vectors to convey information non-invasively into intact brain tissue. In this context, selective mechano-sensitization of neuronal circuits would pave the way to develop a new cell-type-specific stimulation approach.

Modulation of nonsense mediated decay by rapamycin.

Rapamycin is a naturally occurring macrolide whose target is at the core of nutrient and stress regulation in a wide range of species. Despite well-established roles as an inhibitor of cap-dependent mRNA translation, relatively little is known about its effects on other modes of RNA processing. Here, we characterize the landscape of rapamycin-induced post-transcriptional gene regulation.

Akt Regulates Axon Wrapping and Myelin Sheath Thickness in the PNS.

Unlabelled: The signaling pathways that regulate myelination in the PNS remain poorly understood. Phosphatidylinositol-4,5-bisphosphate 3-kinase 1A, activated in Schwann cells by neuregulin and the extracellular matrix, has an essential role in the early events of myelination. Akt/PKB, a key effector of phosphatidylinositol-4,5-bisphosphate 3-kinase 1A, was previously implicated in CNS, but not PNS myelination.

Stable isotope labeling by amino acids in cultured primary neurons.

Cultured primary neurons are a well-established model for the study of neuronal function. Conventional stable isotope labeling with amino acids in cell culture (SILAC) requires nearly complete metabolic labeling of proteins and therefore is difficult to apply to cultured primary neurons, which do not divide in culture. Here we describe a protocol that utilizes a multiplex SILAC labeling strategy for primary cultured neurons.

Investigating a novel protein using mass spectrometry: the example of tumor differentiation factor (TDF).

Better understanding of central nervous system (CNS) molecules can include the identification of new molecules and their receptor systems. Discovery of novel proteins and elucidation of receptor targets can be accomplished using mass spectrometry (MS). We describe a case study of such a molecule, which our lab has studied using MS in combination with other protein identification techniques, such as immunohistochemistry (IHC) and Western blotting.

Proteomic approaches to dissect neuronal signaling pathways.

With an increasing awareness of mental health issues and neurological disorders, "understanding the brain" is one of the biggest current challenges in biological research. This has been recognized by both governments and funding agencies, and includes the need to understand connectivity of brain regions and coordinated network activity, as well as cellular and molecular mechanisms at play. In this chapter, we will describe how we have taken advantage of different proteomic techniques to unravel molecular mechanisms underlying two modulators of neuronal function: Neurotrophins and antipsychotics.

Trk receptors.

The tropomyosin-related tyrosine kinase (Trk) receptors were initially described as a family of growth factor receptors required for neuronal survival. They have since been shown to influence many aspects of neuronal development and function, including differentiation, outgrowth, and synaptic plasticity. This chapter will give an overview on the biology of Trk receptors within the nervous system.

Trkb signaling in pericytes is required for cardiac microvessel stabilization.

Pericyte and vascular smooth muscle cell (SMC) recruitment to the developing vasculature is an important step in blood vessel maturation. Brain-derived neurotrophic factor (BDNF), expressed by endothelial cells, activates the receptor tyrosine kinase TrkB to stabilize the cardiac microvasculature in the perinatal period. However, the effects of the BDNF/TrkB signaling on pericytes/SMCs and the mechanisms downstream of TrkB that promote vessel maturation are unknown.