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At least 19 records

Up-regulation of DRP-3 long isoform during the induction of neural progenitor cells by glutamate treatment in the ex vivo rat retina

Glutamate has been shown to induce neural progenitor cells in the adult vertebrate retina. However, protein dynamics during progenitor cell induction by glutamate are not fully understood. To identify specific proteins involved in the process, we employed two-dimensional electrophoresis-based proteomics on glutamate untreated and treated retinal ex vivo sections. Rat retinal tissues were incubated with 1 mM glutamate for 1 h, followed by incubation in glutamate-free media for a total of 24 h. Consistent with prior reports, it was found that mitotic cells appeared in the outer nuclear layer without any histological damage. Immunohistological evaluations and immunoblotting confirmed the emergence of neuronal progenitor cells in the mature retina treated with glutamate. Proteomic analysis revealed the up-regulation of dihydropyrimidinase-related protein 3 (DRP-3), DRP-2 and stress-induced-phosphoprotein 1 (STIP1) during neural progenitor cell induction by glutamate. Moreover, mRNA expression of DRP-3, especially, its long isoform, robustly increased in the treated retina compared to that in the untreated retina. These results may indicate that glutamate induces neural progenitor cells in the mature rat retina by up-regulating the proteins which mediate cell mitosis and neurite growth. - Highlights: • Glutamate induced neuronal progenitor cells in the mature rat retina. • Proteomic analysis revealed the up-regulation of DRP-3, DRP-2 and STIP1. • mRNA expression of DRP-3, especially, its long isoform, robustly increased.

60 APPLIED LIFE SCIENCES↗

The neurotoxic effects of N-methyl-N-nitrosourea on the electrophysiological property and visual signal transmission of rat's retina

The neurotoxic effects of N-methyl-N-nitrosourea (MNU) on the inner retinal neurons and related visual signal circuits have not been described in any animal models or human, despite ample morphological evidences about the MNU induced photoreceptor (PR) degeneration. With the helping of MEA (multielectrode array) recording system, we gained the opportunity to systemically explore the neural activities and visual signal pathways of MNU administrated rats. Our MEA research identified remarkable alterations in the electrophysiological properties and firstly provided instructive information about the neurotoxicity of MNU that affects the signal transmission in the inner retina. Moreover, the spatial electrophysiological functions of retina were monitored and found that the focal PRs had different vulnerabilities to the MNU. The MNU-induced PR dysfunction exhibited a distinct spatial- and time-dependent progression. In contrast, the spiking activities of both central and peripheral RGCs altered synchronously in response to the MNU administration. Pharmacological tests suggested that gap junctions played a pivotal role in this homogeneous response of RGCs. SNR analysis of MNU treated retina suggested that the signaling efficiency and fidelity of inner retinal circuits have been ruined by this toxicant, although the microstructure of the inner retina seemed relatively consolidated. The present study provided an appropriate example of MEA investigations on the toxicant induced pathological models and the effects of the pharmacological compounds on neuron activities. The positional MEA information would enrich our knowledge about the pathology of MNU induced RP models, and eventually be instrumental for elucidating the underlying mechanism of human RP. - Highlights: • We systemically explored the neural activities and visual signal pathways of MNU administrated retinas. • The focal photoreceptors had different vulnerabilities to the MNU administration. • Pharmacological tests suggested that gap junctions played a pivotal role in this homogeneous response of RGCs. • The present study provided an appropriate example of MEA investigations on the toxicant induced pathological models. • The MEA information would enrich our knowledge about the pathology of MNU induced RP models.

60 APPLIED LIFE SCIENCES↗

Evolution of neuronal cell classes and types in the vertebrate retina

The basic plan of the retina is conserved across vertebrates, yet species differ profoundly in their visual needs1. Retinal cell types may have evolved to accommodate these varied needs, but this has not been systematically studied. Here we generated and integrated single-cell transcriptomic atlases of the retina from 17 species: humans, two non-human primates, four rodents, three ungulates, opossum, ferret, tree shrew, a bird, a reptile, a teleost fish and a lamprey. We found high molecular conservation of the six retinal cell classes (photoreceptors, horizontal cells, bipolar cells, amacrine cells, retinal ganglion cells (RGCs) and Müller glia), with transcriptomic variation across species related to evolutionary distance. Major subclasses were also conserved, whereas variation among cell types within classes or subclasses was more pronounced. However, an integrative analysis revealed that numerous cell types are shared across species, based on conserved gene expression programmes that are likely to trace back to an early ancestral vertebrate. The degree of variation among cell types increased from the outer retina (photoreceptors) to the inner retina (RGCs), suggesting that evolution acts preferentially to shape the retinal output. Finally, we identified rodent orthologues of midget RGCs, which comprise more than 80% of RGCs in the human retina, subserve high-acuity vision, and were previously believed to be restricted to primates. By contrast, the mouse orthologues have large receptive fields and comprise around 2% of mouse RGCs. Projections of both primate and mouse orthologous types are overrepresented in the thalamus, which supplies the primary visual cortex. We suggest that midget RGCs are not primate innovations, but are descendants of evolutionarily ancient types that decreased in size and increased in number as primates evolved, thereby facilitating high visual acuity and increased cortical processing of visual information.

59 BASIC BIOLOGICAL SCIENCES↗

Exposure to As, Cd and Pb-mixture impairs myelin and axon development in rat brain, optic nerve and retina

Arsenic (As), lead (Pb) and cadmium (Cd) are the major metal contaminants of ground water in India. We have reported the toxic effect of their mixture (metal mixture, MM), at human relevant doses, on developing rat astrocytes. Astrocyte damage has been shown to be associated with myelin disintegration in CNS. We, therefore, hypothesized that the MM would perturb myelinating white matter in cerebral cortex, optic nerve (O.N.) and retina. We observed modulation in the levels of myelin and axon proteins, such as myelin basic protein (MBP), proteolipid protein, 2′-, 3′-cyclic-nucleotide-3′-phosphodiesterase, myelin-associated glycoprotein and neurofilament (NF) in the brain of developing rats. Dose and time-dependent synergistic toxic effect was noted. The MBP- and NF-immunolabeling, as well as luxol-fast blue (LFB) staining demonstrated a reduction in the area of intact myelin-fiber, and an increase in vacuolated axons, especially in the corpus-callosum. Transmission electron microscopy (TEM) of O.N. revealed a reduction in myelin thickness and axon-density. The immunolabeling with MBP, NF, and LFB staining in O.N. supported the TEM data. The hematoxylin and eosin staining of retina displayed a decrease in the thickness of nerve-fiber, plexiform-layer, and retinal ganglion cell (RGC) count. Investigating the mechanism revealed a loss in glutamine synthetase activity in the cerebral cortex and O.N., and a fall in the brain derived neurotrophic factor in retina. An enhanced apoptosis in MBP, NF and Brn3b-containing cells justified the diminution in myelinating axons in CNS. Our findings for the first time indicate white matter damage by MM, which may have significance in neurodevelopmental-pediatrics, neurotoxicology and retinal-cell biology. - Highlights: • As, Cd and Pb-mixture, at human relevant dose, demyelinate developing rat CNS. • The attenuation in myelin and axon is synergistic. • The optic nerve and brain demonstrate reduced glutamine synthetase. • The retina exhibits diminished neurotrophin levels and cellular differentiation. • The toxic effect is apoptotic.

60 APPLIED LIFE SCIENCES↗

Restricted expression of Neuroglobin in the mouse retina and co-localization with Melanopsin and Tyrosine Hydroxylase

Highlights: Black-Right-Pointing-Pointer Restricted Neuroglobin expression in the mouse retina. Black-Right-Pointing-Pointer Antibody validation using Neuroglobin-null mice. Black-Right-Pointing-Pointer Co-expression of Neuroglobin with Melanopsin and tyrosine hydroxylase. Black-Right-Pointing-Pointer No effect of Neuroglobin deficiency on neuronal survival. -- Abstract: Neuroglobin (Ngb), a neuronal specific oxygen binding heme-globin, reported to be expressed at high levels in most layers of the murine retina. Ngb's function is presently unknown, but based on its high expression level and oxygen binding capabilities Ngb was proposed to function as an oxygen reservoir facilitating oxygen metabolism in highly active neurons or to function as a neuroprotectant. In the present study, we re-examined the expression pattern of Ngb in the retina using a highly validated antibody. Furthermore, intactness of retino-hypothalamic projections and the retinal expression level of Melanopsin and Tyrosine Hydroxylase were investigated in Ngb-null mice. Ngb-immunoreactivity was found in a few neurons of the ganglion cell and inner nuclear layers co-expressing Melanopsin and Tyrosine Hydroxylase, respectively. Ngb deficiency neither affected the level of Melanopsin and Tyrosine Hydroxylase proteins nor the intactness of PACAP-positive retinohypothalamic projections in the suprachiasmatic nucleus. Based on the present results, it seems unlikely that Ngb could have a major role in retinal oxygen homeostasis and neuronal survival under normal conditions. The present study suggests that a number of previously published reports have relied on antibodies with dubious specificity.

60 APPLIED LIFE SCIENCES↗

Dll4-Notch1 signaling but not VEGF-A is essential for hyperoxia induced vessel regression in retina

Highlights: • Our study find that VEGF-A is not an indispensable factor in hyperoxia induced vessel regression. • Hyperoxia exposure inactivated Dll4-Notch1 signaling in vessel regression region. • Inhibition of Dll4-Notch1 signaling by short-time administration of Dll4 neutralizing antibodies or DAPT exaggerates hyperoxia induced vascular regression. It is well recognized that decreased vascular endothelial growth factor A (VEGF-A) mRNA plays an important role in retinal vessel regression induced by hyperoxia. However, this concept has been challenged by increasing new evidence. Furthermore, VEGF-A strongly enhances Dll4 expression and inhibition of Dll4-Notch signaling leads to excessive sprouting angiogenesis. Recently, it is shown that inactivation of Dll4-Notch1 signaling reduce hyperoxia induced vessel regression. It is unknown whether sprouting angiogenesis contributes to the protective effect or not and further investigations are needed. Moreover, the expression of Dll4 or Notch1 activation in the regressing plexus remains elucidated. To determine the role of VEGF-A and Dll4-Notch1 signaling in hyperoxia induced vascular regression in the retina, we used mice at postnatal day 5 (P5) - P7. Hyperoxia induced massive vascular regression in the central plexus but not in the angiogenic plexus and had no effect on sprouting angiogenesis. Immunostaining showed that VEGF-A was significantly repressed in the angiogenic front region after hyperoxia exposure but not detectable in the central area of both normoxia and hyperoxia treated retinas. In contrast, Notch ligand Delta-like 4 (Dll4) and Notch1 intracellular domain (N1-ICD) expression were inhibited in the regressing capillaries of central retina but comparable in the angiogenic plexus after high oxygen treatment. Moreover, administration of Dll4 neutralizing antibody or γ-Secretase inhibitor DAPT significantly aggravated vessel regression induced by short-time hyperoxia administration. Our data show that repressed Dll4-Notch1 signaling pathway but not downregulation of VEGF-A expression are responsible for hyperoxia induced pervasive vessel regression.

60 APPLIED LIFE SCIENCES↗

A biomimetic eye with a hemispherical perovskite nanowire array retina

Human eyes possess exceptional image-sensing characteristics such as an extremely wide field of view, high resolution and sensitivity with low aberration1. Biomimetic eyes with such characteristics are highly desirable, especially in robotics and visual prostheses. However, the spherical shape and the retina of the biological eye pose an enormous fabrication challenge for biomimetic devices2,3. In this paper, we present an electrochemical eye with a hemispherical retina made of a high-density array of nanowires mimicking the photoreceptors on a human retina. The device design has a high degree of structural similarity to a human eye with the potential to achieve high imaging resolution when individual nanowires are electrically addressed. Additionally, we demonstrate the image-sensing function of our biomimetic device by reconstructing the optical patterns projected onto the device. This work may lead to biomimetic photosensing devices that could find use in a wide spectrum of technological applications.

42 ENGINEERING↗

Simulation and performance of an artificial retina for 40 MHz track reconstruction

We present the results of a detailed simulation of the artificial retina pattern-recognition algorithm, designed to reconstruct events with hundreds of charged-particle tracks in pixel and silicon detectors at LHCb with LHC crossing frequency of 40 MHz. Performances of the artificial retina algorithm are assessed using the official Monte Carlo samples of the LHCb experiment. We found performances for the retina pattern-recognition algorithm comparable with the full LHCb reconstruction algorithm.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗

Retina Image Analysis and Ocular Telehealth: The Oak Ridge National Laboratory-Hamilton Eye Institute Case Study

Automated retina image analysis has reached a high level of maturity in recent years, and thus the question of how validation is performed in these systems is beginning to grow in importance. One application of retina image analysis is in telemedicine, where an automated system could enable the automated detection of diabetic retinopathy and other eye diseases as a low-cost method for broad-based screening. In this work we discuss our experiences in developing a telemedical network for retina image analysis, including our progression from a manual diagnosis network to a more fully automated one. We pay special attention to how validations of our algorithm steps are performed, both using data from the telemedicine network and other public databases.

Karnowski, Thomas↗

Retina-inspired narrowband perovskite sensor array for panchromatic imaging

The retina is the essential part of the human visual system that receives light, converts it to neural signal, and transmits to brain for visual recognition. The red, green, and blue (R/G/B) cone retina cells are natural narrowband photodetectors (PDs) sensitive to R/G/B lights. Connecting with these cone cells, a multilayer neuro-network in the retina provides neuromorphic preprocessing before transmitting to brain. Inspired by this sophistication, we develop the narrowband (NB) imaging sensor combining R/G/B perovskite NB sensor array (mimicking the R/G/B photoreceptors) with a neuromorphic algorithm (mimicking the intermediate neural network) for high-fidelity panchromatic imaging. Compared to commercial sensors, we use perovskite “intrinsic” NB PD to exempt the complex optical filter array. In addition, we use an asymmetric device configuration to collect photocurrent without external bias, enabling a power-free photodetection feature. These results display a promising design for efficient and intelligent panchromatic imaging.

42 ENGINEERING↗

Morphological Changes in the Retina Under Conditions of Experimental In Vivo Regional Ischemia/Reperfusion

Two models of retinal ischemia/reperfusion were developed in an experiment on rats and structural changes in eye tissues in the early and late postischemic periods were studied. Ischemia/reperfusion was modeled by elevation of intraocular pressure to 110 mm Hg for 30 min with air injection into the anterior chamber of the eye with a special device or subconjunctival administration of 0.2 ml 4×10{sup –6} M endothelin-1. Morphological studies of the retina of enucleated eyes were performed in 3, 7, and 30 days after ischemia/reperfusion. In 3 days, signs of retinal ischemia were seen (retinal edema and ganglion cell damage). In the late post-ischemic period (30 days), atrophy of the outer nuclear and outer plexiform layer of the retina was observed in animals with retinal ischemia/reperfusion caused by intraocular pressure elevation and complete destruction of neuronal cells was found after administration of endothelin-1.

60 APPLIED LIFE SCIENCES↗

Staurosporine induces ganglion cell differentiation in part by stimulating urokinase-type plasminogen activator expression and activation in the developing chick retina

Highlights: Black-Right-Pointing-Pointer Staurosporine mediates stimulation of RGC differentiation in vitro cultured retinal neuroblasts. Black-Right-Pointing-Pointer Staurosporine mediates uPA activation during RGC differentiation in vitro. Black-Right-Pointing-Pointer Inhibition of uPA blocks the staurosporine mediated RGC differentiation both in vitro and in ovo. Black-Right-Pointing-Pointer Thus, uPA may play a role in the staurosporine-mediated stimulation of RGC differentiation. -- Abstract: Here, we investigated whether staurosporine-mediated urokinase-type plasminogen activator (uPA) activation is involved in retinal ganglion cell (RGC) differentiation. Retinal cells were isolated from developing chick retinas at embryonic day 6 (E6). Relatively few control cells grown in serum-free medium started to form processes by 12 h. In contrast, staurosporine-treated cells had processes within 3 h, and processes were evident at 8 h. Immunofluorescence staining showed that Tuj-1-positive cells with shorter neurites could be detected in control cultures at 18 h, whereas numerous Tuj-1 positive ganglion cells with longer neuritic extensions were seen in staurosporine-treated cultures. BrdU-positive proliferating cells were more numerous in control cultures than in staurosporine-treated cultures, and the BrdU staining was not detected in post-mitotic Tuj-1 positive ganglion cells. Western blotting of cell lysates showed that staurosporine induced high levels of the active form of uPA. The staurosporine-induced uPA signal was localized predominantly in the soma, neurites and axons of Tuj-1-positive ganglion cells. Amiloride, an inhibitor of uPA, markedly reduced staurosporine-induced Tuj-1 staining, neurite length, neurite number, and uPA staining versus controls. In developing retinas in ovo, amiloride administration remarkably reduced the staurosporine-induced uPA staining and RGC differentiation. Taken together, our in vitro and in vivo data collectively indicate that uPA plays a role in the staurosporine-mediated stimulation of RGC differentiation.

60 APPLIED LIFE SCIENCES↗

Cholesterol enhances amyloid {beta} deposition in mouse retina by modulating the activities of A{beta}-regulating enzymes in retinal pigment epithelial cells

Highlights: Black-Right-Pointing-Pointer Cholesterol-treated RPE produces more A{beta} than non-treated RPE. Black-Right-Pointing-Pointer Neprilysin expression and activity decreased in cholesterol-treated RPE. Black-Right-Pointing-Pointer {alpha}-Secretase expression and activity decreased in cholesterol-treated RPE. Black-Right-Pointing-Pointer Cholesterol-enriched diet induced subRPE deposits in aged mice. Black-Right-Pointing-Pointer A{beta} were present in cholesterol-enriched-diet-induced subRPE deposits in aged mice. -- Abstract: Subretinally-deposited amyloid {beta} (A{beta}) is a main contributor of developing age-related macular degeneration (AMD). However, the mechanism causing A{beta} deposition in AMD eyes is unknown. Hypercholesterolemia is a significant risk for developing AMD. Thus, we investigated the effects of cholesterol on A{beta} production in retinal pigment epithelial (RPE) cells in vitro and in the mouse retina in vivo. RPE cells isolated from senescent (12-month-old) C57BL/6 mice were treated with 10 {mu}g/ml cholesterol for 48 h. A{beta} amounts in culture supernatants were measured by ELISA. Activity and expression of enzymes and proteins that regulate A{beta} production were examined by activity assay and real time PCR. The retina of mice fed cholesterol-enriched diet was examined by transmission electron microscopy. Cholesterol significantly increased A{beta} production in cultured RPE cells. Activities of A{beta} degradation enzyme; neprilysin (NEP) and anti-amyloidogenic secretase; {alpha}-secretase were significantly decreased in cell lysates of cholesterol-treated RPE cells compared to non-treated cells, but there was no change in the activities of {beta}- or {gamma}-secretase. mRNA levels of NEP and {alpha}-secretase (ADAM10 and ADAM17) were significantly lower in cholesterol-treated RPE cells than non-treated cells. Senescent (12-month-old) mice fed cholesterol-enriched chow developed subRPE deposits containing A{beta}, whereas age-matched mice fed standard rodent chow diet did not. Activities and mRNA levels of NEP and {alpha}-secretase were significantly lower in native RPE cells freshly isolated from cholesterol-enriched chow fed mice compared to standard rodent chow fed mice. These findings suggest that cholesterol enhances subretinal A{beta} accumulation by modulating the activities of enzymes degrading and processing A{beta} in RPE cells in senescent subjects.

60 APPLIED LIFE SCIENCES↗

Light induces translocation of NF-κB p65 to the mitochondria and suppresses expression of cytochrome c oxidase subunit III (COX III) in the rat retina

The transcription factor nuclear factor kappaB (NF-κB) plays various roles in cell survival, apoptosis, and inflammation. In the rat retina, NF-κB activity increases after exposure to damaging light, resulting in degeneration of photoreceptors. Here, we report that in dark-adapted rats exposed for 6 h to bright white light, the p65 subunit of retinal NF-κB translocates to the mitochondria, an event associated with a decrease in expression of cytochrome c oxidase subunit III (COX III). However, sustained exposure for 12 h depleted p65 from the mitochondria, and enhanced COX III expression. Treatment with the protective antioxidant PBN prior to light exposure prevents p65 depletion in the mitochondria and COX III upregulation during prolonged exposure, and apoptosis in photoreceptor cells. These results indicate that COX III expression is sensitive to the abundance of NF-κB p65 in the mitochondria, which, in turn, is affected by exposure to damaging light. - Highlights: • Damaging light exposure of the retina induces NF-κB p65 mitochondrial translocation. • NF-κB p65 mitochondrial translocation is associated with the decrease of COX III expression. • Prolonged light exposure depletes mitochondrial p65 resulting in the increase in COX III expression. • NF-κB p65 and COX III expression play an important role in the light-induced photoreceptor degeneration.

60 APPLIED LIFE SCIENCES↗

An Artificial Retina Processor for Track Reconstruction at the LHC Crossing Rate

The goal of the INFN-RETINA R&D project is to develop and implement a computational methodology that allows to reconstruct events with a large number (> 100) of charged-particle tracks in pixel and silicon strip detectors at 40 MHz, thus matching the requirements for processing LHC events at the full bunch-crossing frequency. Our approach relies on a parallel pattern-recognition algorithm, dubbed artificial retina, inspired by the early stages of image processing by the brain. In order to demonstrate that a track-processing system based on this algorithm is feasible, we built a sizable prototype of a tracking processor tuned to 3 000 patterns, based on already existing readout boards equipped with Altera Stratix III FPGAs. The detailed geometry and charged-particle activity of a large tracking detector currently in operation are used to assess its performances. We report on the test results with such a prototype.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗

First results of the silicon telescope using an 'artificial retina' for fast track finding

We present the first results of the prototype of a silicon tracker with trigger capabilities based on a novel approach for fast track finding. The working principle of the 'artificial retina' is inspired by the processing of visual images by the brain and it is based on extensive parallelization of data distribution and pattern recognition. The algorithm has been implemented in commercial FPGAs in three main logic modules: a switch for the routing of the detector hits, a pool of engines for the digital processing of the hits, and a block for the calculation of the track parameters. The architecture is fully pipelined and allows the reconstruction of real-time tracks with a latency less then 100 clock cycles, corresponding to 0.25 microsecond at 400 MHz clock. The silicon telescope consists of 8 layers of single-sided silicon strip detectors with 512 strips each. The detector size is about 10 cm x 10 cm and the strip pitch is 183 μm. The detectors are read out by the Beetle chip, a custom ASICs developed for LHCb, which provides the measurement of the hit position and pulse height of 128 channels. The 'artificial retina' algorithm has been implemented on custom data acquisition boards based on FPGAs Xilinx Kintex 7 lx160. The parameters of the tracks detected are finally transferred to host PC via USB 3.0. The boards manage the read-out ASICs and the sampling of the analog channels. The read-out is performed at 40 MHz on 4 channels for each ASIC that corresponds to a decoding of the telescope information at 1.1 MHz. We report on the first results of the fast tracking device and compare with simulations. (authors)

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗

Infrared retina

Exemplary embodiments provide an infrared (IR) retinal system and method for making and using the IR retinal system. The IR retinal system can include adaptive sensor elements, whose properties including, e.g., spectral response, signal-to-noise ratio, polarization, or amplitude can be tailored at pixel level by changing the applied bias voltage across the detector. "Color" imagery can be obtained from the IR retinal system by using a single focal plane array. The IR sensor elements can be spectrally, spatially and temporally adaptive using quantum-confined transitions in nanoscale quantum dots. The IR sensor elements can be used as building blocks of an infrared retina, similar to cones of human retina, and can be designed to work in the long-wave infrared portion of the electromagnetic spectrum ranging from about 8 .mu.m to about 12 .mu.m as well as the mid-wave portion ranging from about 3 .mu.m to about 5 .mu.m.

47 OTHER INSTRUMENTATION↗

First Results of an “Artificial Retina” Processor Prototype

We report on the performance of a specialized processor capable of reconstructing charged particle tracks in a realistic LHC silicon tracker detector, at the same speed of the readout and with sub-microsecond latency. The processor is based on an innovative pattern-recognition algorithm, called “artificial retina algorithm”, inspired from the vision system of mammals. A prototype of the processor has been designed, simulated, and implemented on Tel62 boards equipped with high-bandwidth Altera Stratix III FPGA devices. The prototype is the first step towards a real-time track reconstruction device aimed at processing complex events of high-luminosity LHC experiments at 40 MHz crossing rate.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗