Tracey C. Dickson

The cause of neuronal degeneration in Alzheimer's disease

Alzheimers disease is associated with a specific pattern of pathological changes in the brain that result in neurodegeneration and the progressive development of dementia. Pathological hallmarks common to the disease include beta-amyloid plaques, dystrophic neurites associated with plaques and neurofibrillary tangles within nerve cell bodies. The exact relationship between these pathological features has been elusive, although it is clear that beta-amyloid plaques precede neurofibrillary tangles in neocortical areas. Examination of the brains of individuals in the preclinical stage of the disease have shown that the earliest form of neuronal pathology associated with beta-amyloid plaques resembles the cellular changes that follow structural injury to axons. Thus, the development of beta-amyloid plaques in the brain may cause physical damage to axons, and the abnormally prolonged stimulation of the neuronal response to this kind of injury ultimately results in the profound cytoskeletal alterations that underlie neurofibrillary pathology and neurodegeneration. Therapeutically, inhibition of the neuronal reaction to physical trauma may be a useful neuroprotective strategy in the earliest stages of Alzheimers disease.

Alpha-synuclein is upregulated in neurones in response to chronic oxidative stress and is associated with neuroprotection

Chronic oxidative stress has been linked to the neurodegenerative changes characteristic of Parkinsons disease, particularly alpha-synuclein accumulation and aggregation. However, it remains contentious whether these alpha-synuclein changes are cytotoxic or neuroprotective. The current study utilised long-term primary neural culture techniques with antioxidant free media to study the cellular response to chronic oxidative stress. Cells maintained in antioxidant free media were exquisitely more vulnerable to acute exposure to hydrogen peroxide, yet exposure of up to 10 days in antioxidant free media did not lead to morphological alterations in neurones or glia. However, a subpopulation of neurones demonstrated a significant increase in the level of alpha-synuclein expressed within the cell body and at synaptic sites. This subset of neurones was also more resistant to apoptotic changes following exposure to antioxidant free media relative to other neurones. These data indicate that increased alpha-synuclein content is associated with neuroprotection from relatively low levels of oxidative stress.

Focal demyelination in Alzheimer's disease and transgenic mouse models

We have investigated alterations in myelin associated with Aβ plaques, a major pathological hallmark of Alzheimer’s disease (AD), in human tissue and relevant transgenic mice models. Using quantitative morphological techniques, we determined that fibrillar Aβ pathology in the grey matter of the neocortex was associated with focal demyelination in human presenilin-1 familial, sporadic and preclinical AD cases, as well as in two mouse transgenic models of AD, compared with age-matched control tissue. This demyelination was most pronounced at the core of Aβ plaques. Furthermore, we found a focal loss of oligodendrocytes in sporadic and preclinical AD cases associated with Aβ plaque cores. In human and transgenic mice alike, plaque-free neocortical regions showed no significant demyelination or oligodendrocyte loss compared with controls. Dystrophic neurites associated with the plaques were also demyelinated. We suggest that such plaque-associated focal demyelination of the cortical grey matter might impair cortical processing, and may also be associated with aberrant axonal sprouting that underlies dystrophic neurite formation.

Altered synapses and gliotransmission in Alzheimer's disease and AD model mice.

Amyloid-β (Aβ) plaque accumulation in Alzheimers disease (AD) is associated with glutamatergic synapse loss, but less is known about its effect on inhibitory synapses. Here, we demonstrate that vesicular γ-aminobutyric acid (GABA) transporter (VGAT) presynaptic bouton density is unaffected in human preclinical and end-stage AD and in APP/PS1 transgenic (TG) mice. Conversely, excitatory vesicular glutamate transporter 1 (VGlut1) boutons are significantly reduced in end-stage AD cases and less reduced in preclinical AD cases and TGs. Aged TGs also show reduced protein levels of VGlut1 and synaptophysin but not VGAT or glutamate decarboxylase (GAD). These findings indicate that GABAergic synapses are preserved in human AD and mouse TGs. Synaptosomes isolated from plaque-rich TG cortex had significantly higher GAD activity than those from plaque-free cerebellum or the cortex of wild-type littermates. Using tissue fractionation, this increased activity was localized to glial synaptosomes, suggesting that Aβ plaques stimulate increased astrocyte GABA synthesis.

Neurofilament triplet proteins are restricted to a subset of neurons in the rat neocortex

The cellular localisation of neurofilament triplet subunits was investigated in the rat neocortex. A subset of mainly pyramidal neurons showed colocalisation of subunit immunolabelling throughout the neocortex, including labelling with the antibody SMI32, which has been used extensively in other studies of the primate cortex as a selective cellular marker. Neurofilament-labelled neurons were principally localised to two or three cell layers in most cortical regions, but dramatically reduced labelling was present in areas such as the perirhinal cortex, anterior cingulate and a strip of cortex extending from caudal motor regions through the medial parietal region to secondary visual areas. However, quantitative analysis demonstrated a similar proportion (10-20%) of cells with neurofilament triplet labelling in regions of high or low labelling. Combining retrograde tracing with immunolabelling showed that cellular content of the neurofilament proteins was not correlated with the length of projection. Double labelling immunohistochemistry demonstrated that neurofilament content in axons was closely associated with myelination. Analysis of SMI32 labelling in development indicated that content of this epitope within cell bodies was associated with relatively late maturation, between postnatal days 14 and 21. This study is further evidence of a cell type-specific regulation of neurofilament proteins within neocortical neurons. Neurofilament triplet content may be more closely related to the degree of myelination, rather than the absolute length, of the projecting axon.

Localization of glutamate receptors in developing cortical neurons in culture and relationship to susceptibility to excitotoxicity

Overactivation of glutamate receptors leading to excitotoxicity has been implicated in the neurodegenerative alterations of a range of central nervous system (CNS) disorders. We have investigated the cell‐type‐specific changes in glutamate receptor localization in developing cortical neurons in culture, as well as the relationship between glutamate receptor subunit distribution with synapse formation and susceptibility to excitotoxicity. Glutamate receptor subunit clustering was present prior to the formation of synapses. However, different receptor types showed distinctive temporal patterns of subunit clustering, localization to spines, and apposition to presynaptic terminals. N‐methyl‐D‐aspartate (NMDA) receptor subunit immunolabelling was present in puncta along dendrites prior to the formation of synapses, with relatively little localization to spines. Vulnerability to NMDA receptor‐mediated excitotoxicity occurred before receptor subunits became localized in apposition to presynaptic terminals. Clustering of α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionic acid (AMPA) receptors occurred concurrently with development of vulnerability to excitotoxicity and was related to localization of AMPA receptors at synapses and in spines. Different AMPA receptor subunits demonstrated cell‐type‐specific localization as well as distribution to spines, dendrites, and extrasynaptic subunit clusters. A subclass of neurons demonstrated substantial perineuronal synaptic innervation, and these neurons expressed relatively high levels of GluR1 and/or GluR4 at receptor puncta, indicating the presence of calcium‐permeable AMPA receptors and suggesting alternative synaptic signalling mechanisms and vulnerability to excitotoxicity. These data demonstrate the relationship between glutamate receptor subunit expression and localization with synaptogenesis and development of neuronal susceptibility to excitotoxicity. These data also suggest that excitotoxicity can be mediated through extrasynaptic receptor subunit complexes along dendrites. J. Comp. Neurol. 498:277–294, 2006.

Initial calcium release from intracellular stores followed by calcium dysregulation is linked to secondary axotomy following transient axonal stretch injury.

J. Neurochem. (2010) 112, 1147–1155.

Axonopathy and cytoskeletal disruption in degenerative diseases of the central nervous system

There has been growing interest in the axon as the initial focus of pathological change in a number of neurodegenerative diseases of the central nervous system. This review concentrates on three major neurodegenerative conditions--amyotrophic lateral sclerosis, multiple sclerosis and Alzheimers disease--with emphasis on key cellular changes that may underlie early axonal dysfunction and pathology and, potentially, the degeneration of neurons. In particular, this review will address recent data that indicate that the main pathological stimuli for these conditions, though often not definitively determined, result in an initial perturbation of the axon and its cytoskeleton, which then results in slow neuronal degeneration and loss of connectivity. The identification of a degenerative process initiated in the axon may provide new therapeutic targets for early intervention to inhibit the grim outcomes related to the progression of these diseases.

Rho kinase activates ezrin-radixin-moesin (ERM) proteins and mediates their function in cortical neuron growth, morphology and motility in vitro

The ezrin‐radixin‐moesin (ERM) family of proteins contribute to cytoskeletal processes underlying many vital cellular functions. Their previously elucidated roles in non‐neuronal cells are an indication of their potential importance in CNS neurons. The specific mechanisms of their activation are unknown, but are likely to depend on factors such as the cell type and biological context. For ERM proteins to become active they must be phosphorylated at a specific C‐terminal threonine residue. In non‐neuronal cells, several kinases, including the Rho GTPase family member Rho kinase, have been identified as capable of phosphorylating the C‐terminal threonine. In these experiments we have investigated specifically the potential role of Rho kinase mediated ERM activation in cortical neurons, utilizing a new pharmacologic inhibitor of Rho kinase and quantitative analysis of aspects of neuronal functions potentially mediated by ERM proteins. Rho kinase inhibition significantly suppressed aspects of neuronal development including neurite initiation and outgrowth, as well as growth cone morphology, with a concomitant loss of phosphorylated ERM immunolabeling in areas associated with neuronal growth. The ability of the Rho kinase inhibitor to decrease the amount of pERM protein was shown by immunoblotting. Post‐injury responses were negatively affected by Rho kinase inhibition, namely by a significant decrease in the number of regenerative neurites. We investigated a novel role for ERM proteins in neuron migration using a post‐injury motility assay, where Rho kinase inhibition resulted in significant and drastic reduction in neuron motility and phosphorylated ERM immunolabeling. Thus, Rho kinase is an important activator of ERMs in mediating specific neuronal functions.

α-Internexin immunoreactivity reflects variable neuronal vulnerability in Alzheimer's disease and supports the role of the β-amyloid plaques in inducing neuronal injury

This study investigated the role of α-internexin in the neuronal alterations associated with β-amyloid plaque formation in Alzheimers disease (AD). Cortical neurons could be defined by their variable content of neurofilament (NF) triplet and α-internexin proteins, with a distinct population of supragranular pyramidal cells containing α-internexin alone. Both NF triplet and α-internexin were localized to reactive axonal structures in physically damaged neurons in experimental trauma models. Similarly, NF triplet and α-internexin immunoreactive neurites were localized to plaques densely packed with β-amyloid fibrils in preclinical AD cases, indicating that certain plaques may cause structural injury or impediment of local axonal transport. However, α-internexin, and not NF triplet, ring-like reactive neurites were present in end-stage AD cases, indicating the relatively late involvement of neurons that selectively contain α-internexin. These results implicate the expression of specific intermediate filament proteins in a distinct hierarchy of differential neuronal vulnerability to AD.