Foveolar Müller Cells of the Pied Flycatcher: Morphology and Distribution of Intermediate Filaments Regarding Cell Transparency.

Specialized intermediate filaments (IFs) have critical importance for the clearness and uncommon transparency of vertebrate lens fiber cells, although the physical mechanisms involved are poorly understood. Recently, an unusual low-scattering light transport was also described in retinal Müller cells. Exploring the function of IFs in Müller cells, we have studied the morphology and distribution pattern of IFs and other cytoskeletal filaments inside the Müller cell main processes in the foveolar part of the avian (pied flycatcher) retina. We found that some IFs surrounded by globular nanoparticles (that we suggest are crystallines) are present in almost every part of the Müller cells that span the retina, including the microvilli. Unlike IFs implicated in the mechanical architecture of the cell, these IFs are not connected to any specific cellular membranes. Instead, they are organized into bundles, passing inside the cell from the endfeet to the photoreceptor, following the geometry of the processes, and repeatedly circumventing numerous obstacles. We believe that the presently reported data effectively confirm that the model of nanooptical channels built of the IFs may provide a viable explanation of Müller cell transparency.

Social Health and Psychological Safety of Students Involved in Online Education during the COVID-19 Pandemic.

Our paper focuses on the issues of social health and psychological safety of university students involved in digital sustainable education during the COVID-19 pandemic. Currently, modern education is becoming inclusive due to the advancements in information and communication technologies (ICT), and it is important not only to stress the relevance of sustainable development and the use of digital technologies, but also their impact on students at schools and universities worldwide. Digital literacy is a newly emerging feature that results from the attitude of team members in the field of digital technologies. This paper explores the impacts of the COVID-19 pandemic on students' learning and well-being and outlines the potential considerations for educational systems as they support students through the recovery period and beyond. Our study is based on the results of our own survey that was administered using a snowball and convenient sample of 1524 respondents (aged 19-26 years; 56.2% females and 43.8% males) from the Czech Republic (N = 804) and Russia (N = 720). We employed the ANOVA and Dirichlet Process mixtures of Generalized Linear Models (DP-GLM) in order to explain the causes of stress and anxiety after grouping variables represented by gender and the study specializations. Our results demonstrate that more than 87% of the students in the sample expressed a medium to high vulnerability to stress, while 58% of the respondents were affected by severe anxiety during their online education engagement. The most important factors that emerged as significant were the fear of getting infected and social distancing, while the best strategy to cope with the stress was self-control. These results allow us to provide practical recommendations for effectively coping with and controlling stress and anxiety among students in the post-pandemic era. In addition, our findings might contribute considerably to the study of the overall long-term effect of the COVID-19 pandemic on the university students, in general, and the use of digital technologies in higher education, as well as on the public health.

Novel noncoding antisense RNA transcribed from human anti-NOS2A locus is differentially regulated during neuronal differentiation of embryonic stem cells.

Here, we report on the discovery of a locus in the human genome, which evolved by gene duplication followed by an internal DNA inversion. This locus exhibits high sequence similarity to the gene for the inducible isoform of NOS protein (NOS2A) and is transcribed into a noncoding RNA containing a region of significant antisense homology with the NOS2A mRNA. We show that this antisense transcript (anti-NOS2A RNA) is expressed in different types of brain tumors, including meningiomas and glioblastomas. More importantly, we demonstrate that the expression profiles of the anti-NOS2A RNA and the NOS2A mRNA exhibit concurrent reciprocal changes in undifferentiated human embryonic stem cells (hESCs) and in hESCs induced to differentiate into neurogenic precursors such as neurospheres. As NOS2A has a role in neurogenesis, our results suggest that the anti-NOS2A RNA is involved in the regulation of neuronal differentiation of hESCs through the modulation of NOS2A gene expression.

Electron microscopy study of the central retinal fovea in Pied flycatcher: evidence of a mechanism of light energy transmission through the retina.

We present unique ultrastructural data on avian retinal cells. Presently and earlier (Zueva et al., 2016) we explored distribution of intermediate filaments (IFs) in retinal cells of the Pied flycatcher (Ficedula hypoleuca, Passeriformes, Aves) in the central foveolar zone. This retinal zone only contains single and double cone photoreceptors. Previously we found that continuous IFs span Müller cells (MC) lengthwise from the retinal inner limiting membrane (ILM) layer up to the outer limiting membrane (OLM) layer. Here we describe long cylindrical bundles of IFs (IFBs) inside the cone inner segments (CIS) adjoining the cone plasma membrane, with these IFBs following along the cone lengthwise, and surrounding the cone at equal spacing one from the other. Double cones form a combined unit, wherein they are separated by their respective plasma membranes. Double cones thus have a common external ring of IFBs, surrounding both cone components. In the layer of cilia, the IFBs that continue into the cone outer segment (COS) follow on to the cone apical tip along the direction of incident light, with single IFs separating from the IFB, touching, and sometimes passing in-between the light-sensitive lamellae of the COS. These new data support our previous hypothesis on the quantum mechanism of light energy propagation through the vertebrate retina (Zueva et al., 2016, 2019).

Quantum mechanism of light energy propagation through an avian retina.

Taking into account the ultrastructure of the Pied Flycatcher foveal retina reported earlier and the earlier reported properties of Müller cell (MC) intermediate filaments (IFs) isolated from vertebrate retina, we proposed a quantum mechanism (QM) of light energy transfer from the inner limiting membrane level to visual pigments in the photoreceptor cells. This mechanism involves electronic excitation energy transfer in a donor-acceptor system, with the IFs excited by photons acting as energy donors, and visual pigments in the photoreceptor cells acting as energy acceptors. It was shown earlier that IFs with diameter 10 nm and length 117 µm isolated from vertebrate eye retina demonstrate properties of light energy guide, where exciton propagates along such IFs from MC endfeet area to photoreceptor cell area. The energy is mostly transferred via the contact exchange quantum mechanism. Our estimates demonstrate that energy transfer efficiencies in such systems may exceed 80-90%. Thus, the presently developed quantum mechanism of light energy transfer in the inverted retina complements the generally accepted classic optical mechanism and the mechanism whereby Müller cells transmit light like optical fibers. The proposed QM of light energy transfer in the inverted retina explains the high image contrast achieved in photopic conditions by an avian eye, being probably also active in other vertebrates.

Errata: Quantum mechanism of light transmission by the intermediate filaments in some specialized optically transparent cells.

[This corrects the article DOI: 10.1117/1.NPh.4.1.011005.].

Quantum mechanism of light transmission by the intermediate filaments in some specialized optically transparent cells.

Some very transparent cells in the optical tract of vertebrates, such as the lens fiber cells, possess certain types of specialized intermediate filaments (IFs) that have essential significance for their transparency. The exact mechanism describing why the IFs are so important for transparency is unknown. Recently, transparency was described also in the retinal Müller cells (MCs). We report that the main processes of the MCs contain bundles of long specialized IFs, each about 10 nm in diameter; most likely, these filaments are the channels providing light transmission to the photoreceptor cells in mammalian and avian retinas. We interpret the transmission of light in such channels using the notions of quantum confinement, describing energy transport in structures with electroconductive walls and diameter much smaller than the wavelength of the respective photons. Model calculations produce photon transmission efficiency in such channels exceeding 0.8, in optimized geometry. We infer that protein molecules make up the channels, proposing a qualitative mechanism of light transmission by such structures. The developed model may be used to describe light transmission by the IFs in any transparent cells.

Spectral selectivity model for light transmission by the intermediate filaments in Müller cells.

Presently we continue our studies of the quantum mechanism of light energy transmission in the form of excitons by axisymmetric nanostructures with electrically conductive walls. Using our theoretical model, we analyzed the light energy transmission by biopolymers forming optical channels within retinal Müller cells. There are specialized intermediate filaments (IF) 10-18nm in diameter, built of electrically conductive polypeptides. Presently, we analyzed the spectral selectivity of these nanostructures. We found that their transmission spectrum depends on their diameter and wall thickness. We also considered the classical approach, comparing the results with those predicted by the quantum mechanism. We performed experimental measurements on model quantum waveguides, made of rectangular nanometer-thick chromium (Cr) tracks. The optical spectrum of such waveguides varied with their thickness. We compared the experimental absorption/transmission spectra with those predicted by our model, with good agreement between the two. We report that the observed spectra may be explained by the same mechanisms as operating in metal nanolayers. Both the models and the experiment show that Cr nanotracks have high light transmission efficiency in a narrow spectral range, with the spectral maximum dependent on the layer thickness. Therefore, a set of intermediate filaments with different geometries may provide light transmission over the entire visible spectrum with a very high (~90%) efficiency. Thus, we believe that high contrast and visual resolution in daylight are provided by the quantum mechanism of energy transfer in the form of excitons, whereas the ultimate retinal sensitivity of the night vision is provided by the classical mechanism of photons transmitted by the Müller cell light-guides.

Timed and targeted differential regulation of nitric oxide synthase (NOS) and anti-NOS genes by reward conditioning leading to long-term memory formation.

In a number of neuronal models of learning, signaling by the neurotransmitter nitric oxide (NO), synthesized by the enzyme neuronal NO synthase (nNOS), is essential for the formation of long-term memory (LTM). Using the molluscan model system Lymnaea, we investigate here whether LTM formation is associated with specific changes in the activity of members of the NOS gene family: Lym-nNOS1, Lym-nNOS2, and the antisense RNA-producing pseudogene (anti-NOS). We show that expression of the Lym-nNOS1 gene is transiently upregulated in cerebral ganglia after conditioning. The activation of the gene is precisely timed and occurs at the end of a critical period during which NO is required for memory consolidation. Moreover, we demonstrate that this induction of the Lym-nNOS1 gene is targeted to an identified modulatory neuron called the cerebral giant cell (CGC). This neuron gates the conditioned feeding response and is an essential part of the neural network involved in LTM formation. We also show that the expression of the anti-NOS gene, which functions as a negative regulator of nNOS expression, is downregulated in the CGC by training at 4 h after conditioning, during the critical period of NO requirement. This appears to be the first report of the timed and targeted differential regulation of the activity of a group of related genes involved in the production of a neurotransmitter that is necessary for learning, measured in an identified neuron of known function. We also provide the first example of the behavioral regulation of a pseudogene.

Suppression of nitric oxide (NO)-dependent behavior by double-stranded RNA-mediated silencing of a neuronal NO synthase gene.

We have used double-stranded RNA (dsRNA)-mediated RNA interference (RNAi) to disrupt neuronal nitric oxide (NO) synthase (nNOS) gene function in the snail Lymnaea stagnalis and have detected a specific behavioral phenotype. The injection of whole animals with synthetic dsRNA molecules targeted to the nNOS-encoding mRNA reduces feeding behavior in vivo and fictive feeding in vitro and interferes with NO synthesis by the CNS. By showing that synthetic dsRNA targeted to the nNOS mRNA causes a significant and long-lasting reduction in the levels of Lym-nNOS mRNA, we verify that specific RNAi has occurred. Importantly, our results establish that the expression of nNOS gene is essential for normal feeding behavior. They also show that dsRNA can be used in the investigation of functional gene expression in the context of whole animal behavior, regardless of the availability of targeted mutation technologies.

A novel long non-coding natural antisense RNA is a negative regulator of Nos1 gene expression.

Long non-coding natural antisense transcripts (NATs) are widespread in eukaryotic species. Although recent studies indicate that long NATs are engaged in the regulation of gene expression, the precise functional roles of the vast majority of them are unknown. Here we report that a long NAT (Mm-antiNos1 RNA) complementary to mRNA encoding the neuronal isoform of nitric oxide synthase (Nos1) is expressed in the mouse brain and is transcribed from the non-template strand of the Nos1 locus. Nos1 produces nitric oxide (NO), a major signaling molecule in the CNS implicated in many important functions including neuronal differentiation and memory formation. We show that the newly discovered NAT negatively regulates Nos1 gene expression. Moreover, our quantitative studies of the temporal expression profiles of Mm-antiNos1 RNA in the mouse brain during embryonic development and postnatal life indicate that it may be involved in the regulation of NO-dependent neurogenesis.

Evaluation of Severe Combined Immunodeficiency and Combined Immunodeficiency Pediatric Patients on the Basis of Cellular Radiosensitivity.

Pediatric patients with severe or nonsevere combined immunodeficiency have increased susceptibility to severe, life-threatening infections and, without hematopoietic stem cell transplantation, may fail to thrive. A subset of these patients have the radiosensitive (RS) phenotype, which may necessitate conditioning before hematopoietic stem cell transplantation, and this conditioning includes radiomimetic drugs, which may significantly affect treatment response. To provide statistical criteria for classifying cellular response to ionizing radiation as the measure of functional RS screening, we analyzed the repair capacity and survival of ex vivo irradiated primary skin fibroblasts from five dysmorphic and/or developmentally delayed pediatric patients with severe combined immunodeficiency and combined immunodeficiency. We developed a mathematical framework for the analysis of γ histone 2A isoform X foci kinetics to quantitate DNA-repair capacity, thus establishing crucial criteria for identifying RS. The results, presented in a diagram showing each patient as a point in a 2D RS map, were in agreement with findings from the assessment of cellular RS by clonogenic survival and from the genetic analysis of factors involved in the nonhomologous end-joining repair pathway. We provide recommendations for incorporating into clinical practice the functional assays and genetic analysis used for establishing RS status before conditioning. This knowledge would enable the selection of the most appropriate treatment regimen, reducing the risk for severe therapy-related adverse effects.

Müller cell alignment in bird fovea: possible role in vision.

Birds which possess high visual acuity, such as eagles and falcons, are known to have retinas with a deep conically curved central foveal pit. There have been different attempts to explain the importance of this particular shape of the fovea in visual resolution. Recently, the function of Müller cells as "light fibers" was discovered, showing how the endfeet of Müller cells trap the light and then transfer it to a single cone photoreceptor. Here we describe how the endfeet of Müller cells line the walls of the foveal pit in the Pied Flycatcher, and how the Müller cell body extends its processes towards individual cones, forming machinery that could allow for light transfer from the pit wall to the photoreceptor layer alongside the pit. We describe how this construction may send an image from the fovea to the cones, and also, how the angular positioning of Müller cells, being optical extensions of the cones, has the advantage of being much denser than on a flat or slightly curved fovea. We, therefore, suggest that this type of optic fiber alignment can be used as a novel type of "amplifying array" that simply increases the amount of megapixels at the photoreceptor cell layer.