scholarly journals Nerve fibers in culture and their interactions with non-neural cells visualized by immunofluorescence.

1979 ◽  
Vol 80 (3) ◽  
pp. 629-641 ◽  
Author(s):  
H Jockusch ◽  
B M Jockusch ◽  
M M Burger
Keyword(s):  
Spinal Cord ◽  
Ganglion Cells ◽  
Cell Types ◽  
Nerve Fibers ◽  
Affinity Column ◽  
Neural Cells ◽  
Embryonic Mouse ◽  
Antibody Staining ◽  
Order Of Magnitude ◽  
Nerve Muscle

Cultures of embryonic mouse spinal cord explants, alone or in combination with rat myotubes, were stained by indirect immunofluorescence using antibodies against three structural proteins to: (a) reveal the distribution of these proteins among different cell types, and (b) test the usefulness of antibody staining to reveal the gross morphology of the neurite network in complex cultures. Affinity column purified antibodies were used against chicken gizzard actin, porcine brain tubulin, and skeletal muscle alpha-actinin. Neurites were stained intensely by anti-actin as was the stress fiber pattern of underlying fibroblasts. With anti-tubulin, the staining of neurites was an order of magnitude more intense than the staining of the microtubule pattern of background fibroblasts. Neurite cell bodies and astrocyte-like glia cells were stained with anti-tubulin and their nuclei remained unstained. Anti-tubulin could thus be used to trace even the finest extensions of nerve processes in spinal cord and spinal cord-muscle cultures. Furthermore, it could be combined with the histochemical reaction for acetylcholinesterase (AChE, EC 3.1.1.7) to demonstrate AChE-positive neurons and specialized nerve-muscle contact sites. The staining of neural elements with anti-alpha-actinin was generally much weaker than with anti-actin and anti-tubulin. Neurites were stained only moderately in comparison to myotube Z lines in the same culture. However, a distinct staining of the periphery of dorsal root ganglion cells was observed. Thus, a protein immunologically related to muscle alpha-actinin is present in the nervous system. In myotubes, Z lines were stained intensely with anti-alpha-actinin while I bands were only faintly stained with anti-actin. In isolated myofibrils, both structures were stained intensely with the same antibody preparations.

Directing Neurite Growth in 3D Collagen Scaffolds With Gradients of Mechanical Properties

2007 ◽  
Author(s):  
Harini G. Sundararaghavan ◽  
David I. Shreiber
Keyword(s):  
Mechanical Properties ◽  
Spinal Cord ◽  
Crosslinking Agent ◽  
Collagen Gel ◽  
Neurite Growth ◽  
Cell Types ◽  
Spinal Cord Regeneration ◽  
3D Scaffold ◽  
Directional Bias ◽  
Order Of Magnitude

Biomaterial scaffolds for nerve and spinal cord regeneration must not only promote neurite re-growth but also direct it. Several cell types, including neurons, respond to the mechanical properties of the substrate on which they are grown. We believe that gradients of mechanical properties can be used to direct neurons. To spatially control the mechanical properties, gradients of genipin — a naturally occurring, cell-tolerated crosslinking agent — are created in 3D through a compliant collagen gel using microfluidics. Gradients of mechanical properties are evaluated by measuring genipin-induced fluorescence, which we have previously correlated to mechanical properties. Growth of neurites was evaluated in gels of uniform stiffness and a gradient generated by incubation in 0 to 1 mM genipin for 12hrs to produce approximately an order of magnitude change in the shear modulus. Neurite growth was evaluated 5 days after gradient formation. Neurites demonstrated a directional bias against the gradient of stiffness. These results demonstrate that neurites can respond to subtle gradients of mechanical properties within a 3D scaffold and point to opportunities to manipulate properties for directed nerve and spinal cord regeneration.

scholarly journals Melanopsin expression in the cornea

2018 ◽  
Vol 35 ◽  
Author(s):  
ANTON DELWIG ◽  
SHAWNTA Y. CHANEY ◽  
ANDREA S. BERTKE ◽  
JAN VERWEIJ ◽  
SUSANA QUIRCE ◽  
...  
Keyword(s):  
Ganglion Cells ◽  
Nerve Fibers ◽  
Visual Signals ◽  
Light Responses ◽  
Rods And Cones ◽  
Retinal Tissue ◽  
Antibody Staining ◽  
Electrophysiological Recordings ◽  
Pupil Constriction ◽  
Sensory Fibers

AbstractA unique class of intrinsically photosensitive retinal ganglion cells in mammalian retinae has been recently discovered and characterized. These neurons can generate visual signals in the absence of inputs from rods and cones, the conventional photoreceptors in the visual system. These light sensitive ganglion cells (mRGCs) express the non-rod, non-cone photopigment melanopsin and play well documented roles in modulating pupil responses to light, photoentrainment of circadian rhythms, mood, sleep and other adaptive light functions. While most research efforts in mammals have focused on mRGCs in retina, recent studies reveal that melanopsin is expressed in non-retinal tissues. For example, light-evoked melanopsin activation in extra retinal tissue regulates pupil constriction in the iris and vasodilation in the vasculature of the heart and tail. As another example of nonretinal melanopsin expression we report here the previously unrecognized localization of this photopigment in nerve fibers within the cornea. Surprisingly, we were unable to detect light responses in the melanopsin-expressing corneal fibers in spite of our histological evidence based on genetically driven markers and antibody staining. We tested further for melanopsin localization in cell bodies of the trigeminal ganglia (TG), the principal nuclei of the peripheral nervous system that project sensory fibers to the cornea, and found expression of melanopsin mRNA in a subset of TG neurons. However, neither electrophysiological recordings nor calcium imaging revealed any light responsiveness in the melanopsin positive TG neurons. Given that we found no light-evoked activation of melanopsin-expressing fibers in cornea or in cell bodies in the TG, we propose that melanopsin protein might serve other sensory functions in the cornea. One justification for this idea is that melanopsin expressed in Drosophila photoreceptors can serve as a temperature sensor.

scholarly journals Characterization and localization of actinogelin, a Ca2+ - sensitive actin accessory protein, in nonmuscle cells.

1982 ◽  
Vol 93 (3) ◽  
pp. 899-909 ◽  
Author(s):  
N Mimura ◽  
A Asano
Keyword(s):  
Intracellular Localization ◽  
Cell Types ◽  
Cell Extract ◽  
Affinity Column ◽  
Precipitation Line ◽  
Actin Bundles ◽  
Antibody Staining ◽  
Nonmuscle Cells ◽  
Tetramethylrhodamine Isothiocyanate ◽  
Antibody Preparations

Actinogelin, which induces gelation of F-actin at Ca2+ concentrations below micromolar concentrations but not at higher concentrations, was isolated in the pure state from Ehrlich tumor cells. The protein consists of subunits of 112,000-115,000 daltons and under physiological conditions is present mostly as a dimer. Up to 1 mol of actinogelin (dimer) binds to 10-12 mol of actin monomer. The binding was slightly decreased by the presence of 50 microM Ca2+ and almost completely inhibited by 300 mM KCl. Antibodies against actinogelin giving a single precipitation line with Ehrlich cell extract and with pure actinogelin were raised in rabbits. Antibody preparations were purified before use in an affinity column containing purified actinogelin. In mouse embryo fibroblasts and 3T3 cells, staining of actin bundles by the antiactinogelin antibody usually was discontinuous or gave a striated appearance. Most of the crossing points of the actin bundles were intensively stained. In epithelial cells from mouse small intestine, actinogelin was distributed throughout the cell, with the exception of the microvilli, which were devoid of staining. In mouse peritoneal cells, the antibody staining patterns were similar to those of tetramethylrhodamine isothiocyanate-labeled heavy meromyosin, but the former usually were sharper than the latter. Intracellular localization of actinogelin was drastically altered by cytochalasin D treatment at 10 microgram/ml. We conclude that actinogelin is present in a wide variety of cell types and discuss the possible participation of actinogelin in the Ca2+-dependent regulation of microfilament distribution.

Melatonin As a Modulator of Degenerative and Regenerative Signaling Pathways in Injured Retinal Ganglion Cells

2019 ◽  
Vol 25 (28) ◽  
pp. 3057-3073 ◽  
Author(s):  
Kobra B. Juybari ◽  
Azam Hosseinzadeh ◽  
Habib Ghaznavi ◽  
Mahboobeh Kamali ◽  
Ahad Sedaghat ◽  
...  
Keyword(s):  
Optic Nerve ◽  
Mitochondrial Dysfunction ◽  
Signaling Pathways ◽  
Retinal Ganglion Cells ◽  
Molecular Mechanisms ◽  
Nitrosative Stress ◽  
Ganglion Cells ◽  
Axon Growth ◽  
Nerve Fibers ◽  
Retinal Ganglion

Optic neuropathies refer to the dysfunction or degeneration of optic nerve fibers caused by any reasons including ischemia, inflammation, trauma, tumor, mitochondrial dysfunction, toxins, nutritional deficiency, inheritance, etc. Post-mitotic CNS neurons, including retinal ganglion cells (RGCs) intrinsically have a limited capacity for axon growth after either trauma or disease, leading to irreversible vision loss. In recent years, an increasing number of laboratory evidence has evaluated optic nerve injuries, focusing on molecular signaling pathways involved in RGC death. Trophic factor deprivation (TFD), inflammation, oxidative stress, mitochondrial dysfunction, glutamate-induced excitotoxicity, ischemia, hypoxia, etc. have been recognized as important molecular mechanisms leading to RGC apoptosis. Understanding these obstacles provides a better view to find out new strategies against retinal cell damage. Melatonin, as a wide-spectrum antioxidant and powerful freeradical scavenger, has the ability to protect RGCs or other cells against a variety of deleterious conditions such as oxidative/nitrosative stress, hypoxia/ischemia, inflammatory processes, and apoptosis. In this review, we primarily highlight the molecular regenerative and degenerative mechanisms involved in RGC survival/death and then summarize the possible protective effects of melatonin in the process of RGC death in some ocular diseases including optic neuropathies. Based on the information provided in this review, melatonin may act as a promising agent to reduce RGC death in various retinal pathologic conditions.

scholarly journals In-depth transcriptomic analysis of human retina reveals molecular mechanisms underlying diabetic retinopathy

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Kolja Becker ◽  
Holger Klein ◽  
Eric Simon ◽  
Coralie Viollet ◽  
Christian Haslinger ◽  
...  
Keyword(s):  
Diabetic Retinopathy ◽  
Rna Sequencing ◽  
Molecular Mechanisms ◽  
Vision Loss ◽  
Ganglion Cells ◽  
Expression Profiles ◽  
Cell Types ◽  
Sequencing Data ◽  
Disease Stages ◽  
Post Transcriptional Regulation

AbstractDiabetic Retinopathy (DR) is among the major global causes for vision loss. With the rise in diabetes prevalence, an increase in DR incidence is expected. Current understanding of both the molecular etiology and pathways involved in the initiation and progression of DR is limited. Via RNA-Sequencing, we analyzed mRNA and miRNA expression profiles of 80 human post-mortem retinal samples from 43 patients diagnosed with various stages of DR. We found differentially expressed transcripts to be predominantly associated with late stage DR and pathways such as hippo and gap junction signaling. A multivariate regression model identified transcripts with progressive changes throughout disease stages, which in turn displayed significant overlap with sphingolipid and cGMP–PKG signaling. Combined analysis of miRNA and mRNA expression further uncovered disease-relevant miRNA/mRNA associations as potential mechanisms of post-transcriptional regulation. Finally, integrating human retinal single cell RNA-Sequencing data revealed a continuous loss of retinal ganglion cells, and Müller cell mediated changes in histidine and β-alanine signaling. While previously considered primarily a vascular disease, attention in DR has shifted to additional mechanisms and cell-types. Our findings offer an unprecedented and unbiased insight into molecular pathways and cell-specific changes in the development of DR, and provide potential avenues for future therapeutic intervention.

scholarly journals Zika Virus Infection Leads to Demyelination and Axonal Injury in Mature CNS Cultures

Viruses ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 91
Author(s):  
Verena Schultz ◽  
Stephanie L. Cumberworth ◽  
Quan Gu ◽  
Natasha Johnson ◽  
Claire L. Donald ◽  
...  
Keyword(s):  
Nervous System ◽  
Neural Development ◽  
Zika Virus ◽  
Developmental Stages ◽  
Cell Types ◽  
Neural Cells ◽  
Zikv Infection ◽  
Axonal Pathology ◽  
Time Frames

Understanding how Zika virus (Flaviviridae; ZIKV) affects neural cells is paramount in comprehending pathologies associated with infection. Whilst the effects of ZIKV in neural development are well documented, impact on the adult nervous system remains obscure. Here, we investigated the effects of ZIKV infection in established mature myelinated central nervous system (CNS) cultures. Infection incurred damage to myelinated fibers, with ZIKV-positive cells appearing when myelin damage was first detected as well as axonal pathology, suggesting the latter was a consequence of oligodendroglia infection. Transcriptome analysis revealed host factors that were upregulated during ZIKV infection. One such factor, CCL5, was validated in vitro as inhibiting myelination. Transferred UV-inactivated media from infected cultures did not damage myelin and axons, suggesting that viral replication is necessary to induce the observed effects. These data show that ZIKV infection affects CNS cells even after myelination—which is critical for saltatory conduction and neuronal function—has taken place. Understanding the targets of this virus across developmental stages including the mature CNS, and the subsequent effects of infection of cell types, is necessary to understand effective time frames for therapeutic intervention.

scholarly journals Spatial receptive field properties of rat retinal ganglion cells

2011 ◽  
Vol 28 (5) ◽  
pp. 403-417 ◽  
Author(s):  
WALTER F. HEINE ◽  
CHRISTOPHER L. PASSAGLIA
Keyword(s):  
Receptive Field ◽  
Ganglion Cell ◽  
Retinal Ganglion Cells ◽  
Ganglion Cells ◽  
Cell Types ◽  
Quantitative Information ◽  
Retinal Ganglion ◽  
X And Y Cells ◽  
Y Cells

AbstractThe rat is a popular animal model for vision research, yet there is little quantitative information about the physiological properties of the cells that provide its brain with visual input, the retinal ganglion cells. It is not clear whether rats even possess the full complement of ganglion cell types found in other mammals. Since such information is important for evaluating rodent models of visual disease and elucidating the function of homologous and heterologous cells in different animals, we recorded from rat ganglion cells in vivo and systematically measured their spatial receptive field (RF) properties using spot, annulus, and grating patterns. Most of the recorded cells bore likeness to cat X and Y cells, exhibiting brisk responses, center-surround RFs, and linear or nonlinear spatial summation. The others resembled various types of mammalian W cell, including local-edge-detector cells, suppressed-by-contrast cells, and an unusual type with an ON–OFF surround. They generally exhibited sluggish responses, larger RFs, and lower responsiveness. The peak responsivity of brisk-nonlinear (Y-type) cells was around twice that of brisk-linear (X-type) cells and several fold that of sluggish cells. The RF size of brisk-linear and brisk-nonlinear cells was indistinguishable, with average center and surround diameters of 5.6 ± 1.3 and 26.4 ± 11.3 deg, respectively. In contrast, the center diameter of recorded sluggish cells averaged 12.8 ± 7.9 deg. The homogeneous RF size of rat brisk cells is unlike that of cat X and Y cells, and its implication regarding the putative roles of these two ganglion cell types in visual signaling is discussed.

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