Imaging membrane potential in dendrites and axons of single neurons

This review focuses on the use of imaging techniques to record electrical signaling in the fine processes of neurons such as dendrites and axons. Voltage imaging began with the use and development of externally applied voltage-sensitive dyes. With the introduction of internally applied dyes and advances in detection technology, it is now possible to record supra-threshold action potential responses, as well as sub-threshold synaptic potentials, in fine neuronal processes including dendritic spines. The development of genetically coded sensors, as well as variants of laser scanning microscopy such as second harmonic generation, offers promise for further advances in this field. Through the use and further development of these methods, optical imaging of membrane potential will continue to be a valuable tool for investigators wishing to explore the electrical events underlying single neuronal computation.     

Mechanisms of translational control in dendrites

Synaptogenesis and synaptic plasticity demonstrate the ability of neurons to mature and respond to various stimuli. Both of these events require protein synthesis, in particular, localized translation within dendrites. Dendrites localize specific mRNAs in proximity to dendritic spines and have the capacity to actively translate these mRNAs within various ‘hotspots’ in the dendritic space. Rates of dendritic translation are stimulated and inhibited by various agents, suggesting that several signaling pathways that modulate localized protein synthesis may exist within the dendrite. This review will cover several suggested pathways for regulation of dendritic translation and propose a correlation between deregulated dendritic translation and disease.     

Genetic and age related models of neurodegeneration in mice: dystrophic axons.

Dystrophic axons (DA) are non-specific lesions that occur in a wide variety of human and animal diseases. In this paper we describe the distribution of these lesions in three newly discovered mouse neurological mutants. The distribution of DA in these mutants is defined by their names, lumbosacral neuroaxonal dystrophy (lnd), located on Chromosome 7, generalized neuroaxonal dystrophy (gnd) and vestibulomotor degeneration (vmd). The last mutant, which has degeneration as well as DA in lateral vestibular nucleus and vestibulo-spinal tracts, dies in the first weeks of life; the first two live for approximately one year. A previously described mutation, dystonia musculorum (dt), was found to produce generalized DA like gnd, but dt/dt mutants die at an early age. DA were also found to occur in the nuclei gracilis and cuneatus, in the area of Clark's column and in lumbo-sacral spinal cord in aging normal mice either fed ad libitum or at a level of 40% dietary restriction. The dietary regimen had little effect on the numbers of DA observed in susceptible areas of the neuroaxis. The mutant models of neuroaxonal dystrophy may prove useful in studies of the pathophysiology of DA in general and of specific inherited diseases of man, such as infantile neuroaxonal dystrophy and Hallervordin-Spatz disease.     

Differential effects of NgCAM and N-cadherin on the development of axons and dendrites by cultured hippocampal neurons

A fundamental step in neuronal development is the acquisition of a polarized form, with distinct axons and dendrites. Although the ability to develop a polarized form appears to be largely an intrinsic property of neurons, it can be influenced by environmental cues. For example, in cell cultures substrate and diffusible factors can enhance and orient axonal development. In this study we examine the effects of growth on each of two cell adhesion molecules (CAMs), NgCAM and N-cadherin, on the development of polarity by cultured hippocampal neurons. We find that although the same pattern of development occurs on control substrates and the CAMs, the CAMs greatly accelerate the rate and extent of development of axons—axons form sooner and grow longer on the CAMs than on the control substrate. In contrast, the CAMs have opposite effects on dendritic development—N-cadherin enhances, but NgCAM reduces dendritic growth compared to control. These results provide further evidence that the development of polarity is largely determined by a cell-autonomous program, but that environmental cues can independently regulate axonal and dendritic growth.     

Atypical features of rat dentate granule cells: recurrent basal dendrites and apical axons

The stereotyped morphology of dentate granule cells in rodents consists of apical dendrites arborizing in the molecular layer and an axon arising from the opposite pole of the soma. Recently, we showed that epilepsy induces the formation of basal dendrites on granule cells and that these dendrites extend into the hilus of the dentate gyrus. The present Golgi study of granule cells from adult rats shows two atypical features for granule cells in control rats. One is the occurrence of recurrent basal dendrites (RBDs) that are defined as basal dendrites arising at or near the hilar pole of the soma and then curving back to the molecular layer. The frequency of granule cells with RBDs was 3.8% in control rats. The second is apical axons of granule cells that were observed to originate from either the apical pole of the soma or an apical dendrite. The incidence of these "apical" axons was about 1%. These morphological findings in the present study suggest that rat granule cells are more heterogeneous than previously indicated. Furthermore, their frequency was not increased in epileptic rats.     

The role of motor proteins in establishing the microtubule arrays of axons and dendrites

Neurons generate two distinct types of processes called axons and dendrites, both of which rely on highly organized arrays of microtubules for their growth and maintenance. Axonal microtubules are uniformly oriented with their plus-ends distal to the cell body, while dendritic microtubules are nonuniformly oriented. In neither case are the microtubules attached to the centrosome or any detectable structure that could establish their distinct patterns of polarity orientation. Here I describe several lines of evidence from my laboratory that support a model for the establishment of these microtubule arrays based on microtubule transport by motor proteins. Microtubules destined for axons and dendrites are nucleated at the centrosome within the cell body of the neuron, and rapidly released. The released microtubules are then transported into the developing processes. Early in neuronal development, the microtubules are transported with their plus-ends-leading into the developing axon and into the immature processes that will develop into dendrites. In the case of the developing dendrites, the plus-end-distal microtubules are later joined by a population of microtubules that are transported into these processes with their minus-ends-leading. Implicit in this model is that there are molecular motor proteins that transport microtubules with the appropriate orientation into each type of process. There is precedent for molecular motor proteins transporting microtubules during mitosis, and our results suggest that the same or similar motors are utilized during the development of axons and dendrites after a neuroblast becomes terminally postmitotic.     

Mechanisms of neurodegeneration in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is the most common variant of motor neurone disease affecting adults that usually strikes during mid to late life. Its aetiology is still poorly understood, although a major breakthrough came with the discovery that mutations in the Cu/Zn superoxide dismutase (SOD1) gene affect approximately 20% of patients with familial ALS. Experiments using both transgenic mice and ALS tissues have been useful in delineating other genetic defects in ALS. However, because only a subset of cases can be attributed to one particular molecular defect (such as mutation of SOD1 or the gene encoding neurofilament H), the aetiology of ALS is likely to be multifactorial. This review discusses the major mechanisms of neurodegeneration in ALS, such as oxidative stress, glutaminergic excitotoxicity, damage to vital organelles, and aberrant protein aggregation.     

Mechanisms of cardiac potassium channel trafficking

The regulation of ion channels involves more than just modulation of their synthesis and kinetics, as controls on their trafficking and localization are also important. Although the body of knowledge is fairly large, the entire trafficking pathway is not known for any one channel. This review summarizes current knowledge on the trafficking of potassium channels that are expressed in the heart. Our knowledge of channel assembly, trafficking through the Golgi apparatus and on to the surface is covered, as are controls on channel surface retention and endocytosis.     

External and internal inputs affecting plasticity of dendrites and axons of the fly's neurons

Neurons and glial cells in the fly's visual system exhibit circadian rhythms through changes in shape and size. Moreover, the number of synaptic contacts between these cells changes during the day and night and in the case of one type of synapses, feedback synapses, is maintained under constant conditions indicating an endogenous origin of this rhythm. The structural changes described above, involving the oscillations in the number of synapses and the size of interneurons and glial cells, are examples of plasticity in the central nervous system driven by internal inputs from a circadian clock and by external stimuli such as light. They are also modulated by visual and other sensory stimuli and by motor activity.     

The role of genetics for biomarker development in neurodegeneration

Research on biomarkers and genetics shares a number of objectives, including the identification of novel disease mechanisms, optimization of therapeutic studies, and improvement of diagnosis and prognosis. The latter is of particular relevance in neurodegenerative diseases where the underlying molecular processes often go on for decades until the first clinical symptoms appear. In this commentary we review the potential contribution that insight gained from genetic research may have on biomarker development in neurodegeneration. We argue that future progress will largely depend on a widespread application of novel high-throughput technologies now becoming available in both fields.     

The role of genetics for biomarker development in neurodegeneration

Research on biomarkers and genetics shares a number of objectives, including the identification of novel disease mechanisms, optimization of therapeutic studies, and improvement of diagnosis and prognosis. The latter is of particular relevance in neurodegenerative diseases where the underlying molecular processes often go on for decades until the first clinical symptoms appear. In this commentary we review the potential contribution that insight gained from genetic research may have on biomarker development in neurodegeneration. We argue that future progress will largely depend on a widespread application of novel high-throughput technologies now becoming available in both fields.     

Hurdles to lymphocyte trafficking in the tumor microenvironment: implications for effective immunotherapy

An important consideration in the development of T cell-based cancer immunotherapy is that effector T cells must efficiently traffic to the tumor microenvironment in order to control malignant progression. T cell trafficking to target tissues is orchestrated by dynamic interactions between circulating lymphocytes and endothelial cells lining blood vessels. It is informative, in this regard, to compare and contrast the molecular mechanisms governing lymphocyte extravasation at distinct vascular sites: (1) high endothelial venules (HEV) of secondary lymphoid organs, which are portals for efficient trafficking of naive and central memory T lymphocytes; (2) non-activated endothelium of normal tissues that mediate relatively low basal levels of trafficking but are rapidly transformed into HEV-like vessels in response to local inflammatory stimuli; and (3) vessels within the intratumoral region and the surrounding peritumoral areas. These vessels can be distinguished by differential expression of hallmark trafficking molecules that function as molecular beacons directing lymphocyte migration across vascular barriers. This article reviews evidence that recruitment of effector T cells to the intratumoral microenvironment is impeded by sub-threshold expression of trafficking molecules on tumor microvessels. Emerging data support the thesis that when considered from the perspective of extravasation, vessels embedded within the intratumoral microenvironment of established tumors do not exhibit stereotypical characteristics of a chronic inflammatory state. A major challenge will be to develop therapeutic approaches to improve trafficking of effector T lymphocytes to tumor sites without skewing the balance in favor of a chronic inflammatory milieu that facilitates tumor maintenance and progression.     

Neutrophil trafficking to lymphoid tissues: physiological and pathological implications

Recent advances have provided evidence for the involvement of neutrophils in both innate and adaptive immunity, robustly challenging the old dogma that neutrophils are short-lived prototypical innate immune cells solely involved in acute responses to microbes and exerting collateral tissue damage. There is now ample evidence showing that neutrophils can migrate into different compartments of the lymphoid system where they contribute to the orchestration of the activation and/or suppression of lymphocyte effector functions in homeostasis and during chronic inflammation, such as autoimmune disorders and cancer. In support of this notion, neutrophils can generate a wide range of cytokines and other mediators capable of regulating the survival, proliferation and functions of both T and B cells. In addition, neutrophils can directly engage with lymphocytes and promote antigen presentation. Furthermore, there is emerging evidence of the existence of distinct and diverse neutrophil phenotypes with immunomodulatory functions that characterise different pathological conditions, including chronic and autoimmune inflammatory conditions. The aim of this review is to discuss the mechanisms implicated in neutrophil trafficking into the lymphoid system and to provide an overview of the immuno-regulatory functions of neutrophils in health and disease in the context of adaptive immunity. Copyright © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.     

Molecular approaches to cerebral laterality: development and neurodegeneration

Little is understood about the genetic or epigenetic mechanisms that underlie brain asymmetry. Because higher cognitive functions such as language, constructional and spatial abilities, and attention are organized along the left/right axis, understanding the underpinnings of this process has significant implications for both developmental biology and cognitive neuroscience. However, scientists have begun to explore, in only the most preliminary manner, the influences of subtle biologically inherited brain asymmetries on human behavior and disease. Because brain asymmetry develops prenatally, the recognition of asymmetry in neurodegeneration implies a possible relationship between the development of cerebral laterality and regional vulnerability in neurodegenerative diseases. This suggests that the study of cerebral asymmetry and laterality is likely to be relevant to a number of degenerative conditions that were previously considered to be only diseases of aging. In this article, I will outline our perspective and some of the approaches that my laboratory has begun to take to characterize the molecular basis of cerebral asymmetry. Most of these data are preliminary and the models presented are highly speculative, reflecting the primitive stage of work defining the molecular basis of cerebral asymmetry.     

The endoplasmic reticulum-associated VP7 of rotavirus is targeted to axons and dendrites in polarized neurons

Summary Rotavirus, which matures and is retained in the endoplasmic reticulum, was used to examine how polarized dorsal root ganglion and spinal cord neurons distributed cytoplasmic and endoplasmic reticulum-associated proteins. A remarkable observation was that NS28, a trans-endoplasmic reticulum-membrane protein which functions as a receptor for budding particles, remained in the cell body during the whole course of infection (48 h) while the VP7 glycoprotein, which is endoplasmic reticulum associated and usually retained in the endoplasmic reticulum, was targeted to axons already 4 h post infection. VP7 was furthermore transported in an endo--N-acetylglucosaminidase H sensitive form through the secretory pathway. The segregated appearances of NS28 and the endo--N-acetylglucosaminidase H sensitive VP7 indicate that VP7 enters a transport compartment separate from NS28. Brefeldin A treatment rapidly disintegrated the Golgi apparatus of the neurons and rapidly blocked axonal transport of Sendai virus glycoproteins, while axonal transport of rotavirus VP7 was not blocked, suggesting that VP7 uses an intracellular pathway in neurons which does not involve the Golgi apparatus.     

Mechanisms of hyperpolarization in regenerated mature motor axons in cat

We found persistent abnormalities in the recovery of membrane excitability in long-term regenerated motor nerve fibres in the cat as indicated in the companion paper. These abnormalities could partly be explained by membrane hyperpolarization. To further investigate this possibility, we compared the changes in excitability in control nerves and long-term regenerated cat nerves (3–5 years after tibial nerve crush) during manoeuvres known to alter axonal membrane Na⁺–K⁺ pump function: polarization, cooling to 20°C, reperfusion after 10 min ischaemia, and up to 60 s of repetitive stimulation at 200 Hz. The abnormalities in excitability of regenerated nerves were reduced by depolarization and cooling and increased by hyperpolarization and during postischaemia. Moreover, the time course of recovery of excitability from repetitive stimulation and ischaemia was prolonged in regenerated nerves. Our data are consistent with an increased demand for electrogenic Na⁺–K⁺ pumping in regenerated nerves leading to membrane hyperpolarization. Such persistent hyperpolarization may influence the ability of the axon to compensate for changes in membrane potential following normal repetitive activity.