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Light-sheet microscopy (LSM), in combination with intrinsically transparent zebrafish larvae, is a method of choice to observe brain function with high frame rates at cellular resolution. Inherently to LSM, however, residual opaque objects cause stripe artifacts, which obscure features of interest and, during functional imaging, modulate fluorescence variations related to neuronal activity. Here, we report how Bessel beams reduce streaking artifacts and produce high-fidelity quantitative data demonstrating a fivefold increase in sensitivity to calcium transients and a 20-fold increase in accuracy in the detection of activity correlations in functional imaging. Furthermore, using principal component analysis, we show that measurements obtained with Bessel beams are clean enough to reveal in one-shot experiments correlations that can not be averaged over trials after stimuli as is the case when studying spontaneous activity. Our results not only demonstrate the contamination of data by systematic and random errors through conventional Gaussian illumination and but,furthermore, quantify the increase in fidelity of such data when using Bessel beams.

[This corrects the article DOI: 10.3389/fncel.2018.00315.].

Age-related macular degeneration (AMD) is a leading cause for visual impairment in aging populations with limited established therapeutic interventions available. Oxidative stress plays an essential role in the pathogenesis of AMD, damaging the retinal pigment epithelium (RPE), which is essential for the function and maintenance of the light-sensing photoreceptors. This study aimed to evaluate the effects of crocetin, one of the main components of Saffron, on an in vitro RPE model of tert-butyl hydroperoxide (TBHP) induced oxidative stress using ARPE19 cells. The effects of crocetin were assessed using lactate de-hydrogenase (LDH) and ATP assays, as well as immunocytochemistry for cell morphology, junctional integrity, and nuclear morphology. The mechanism of crocetin action was determined via assessment of energy production pathways, including mitochondrial respiration and glycolysis in real-time as well as investigation of extracellular signal-regulated kinase 1/2 (ERK1/2) activation and distribution. Our results show that crocetin pre-treatment protects ARPE19 cells from TBHP-induced LDH release, intracellular ATP depletion, nuclear condensation, and disturbance of junctional integrity and cytoskeleton. The protective effect of crocetin is mediated via the preservation of energy production pathways and activation of ERK1/2 in the first minutes of TBHP exposure to potentiate survival pathways. The combined data suggest that a natural antioxidant, such as crocetin, represents a promising candidate to prevent oxidative stress in RPE cells and might halt or delay disease progression in AMD.

The authors wish to make the following correction to Figure 6B of this article [...]

This experiment was conducted to evaluate the effect of zinc, manganese, and copper sources (inorganic vs. organic) in the diet on laying performance and eggshell quality characteristics. One hundred and eighty Hy-Line W-36 layers at 38 weeks of age were allocated to 36-layer cages of five hens each. Each six cages were randomly assigned to one of the six experimental diets fed from 38 to 53 week of age. In three experimental treatments, the basal diet was supplemented with 65–75–7 or 65–75–7 or 40–40–7 mg/kg of Zn, Mn, and Cu, respectively, from their oxide or sulfate sources. Three other groups were fed diets supplemented with 20–20–3.5 or 40–40–7.5 or 60–60–10.5 mg/kg of organic forms of Zn, Mn, and Cu, respectively. Dietary treatments significantly did affect feed intake (P < 0.001), feed conversion ratio (P < 0.001) and percentage of broken eggs (P < 0.05). Substitution of Zn and Mn oxides (65 and 75 mg kg−1, respectively) with equal amounts of their sulfate forms significantly improved feed intake, feed conversion ratio, percentage of broken eggs, and Haugh Unit (P < 0.05). In addition, laying hens maintained their performance when substitution of Zn and Mn oxides and Cu sulfate (65, 75, and 7 mg kg−1, respectively) reduced up to 20, 20, and 3.5 mg kg−1 by amino acid complexes of the microelements. The results showed that a corn–soybean diet supplemented with the organic forms of Zn, Mn, and Cu at a dosage 50% to 75% lower than NRC recommendation is sufficient to maintain laying performance and can improve eggshell and albumen qualities of the egg in laying hens.

Morphogen gradients impart positional information to cells in a homogenous tissue field. Fgf8a, a highly conserved growth factor, has been proposed to act as a morphogen during zebrafish gastrulation. However, technical limitations have so far prevented direct visualization of the endogenous Fgf8a gradient and confirmation of its morphogenic activity. Here, we monitored Fgf8a propagation in the developing neural plate using a CRISPR/Cas9-mediated EGFP knock-in at the endogenous fgf8a locus. By combining sensitive imaging platforms with single-molecule Fluorescence Correlation Spectroscopy (FCS), we demonstrate that Fgf8a, produced at the embryonic margin, propagates by free diffusion through the extracellular space and forms a graded distribution towards the animal pole. Overlaying the Fgf8a gradient curve with expression profiles of its downstream targets determines the precise input-output relationship of Fgf8a mediated patterning. Manipulation of the Fgf8a input alters the signaling outcome, thereby establishing Fgf8a as a bona fide morphogen during zebrafish gastrulation. Furthermore, using diffusion-hindered versions of Fgf8a, we demonstrate that extracellular diffusion of the protein from the source is critical for it to achieve its morphogenic potential.

One of the most exciting challenges of neurosciences in the last few years is the real-time recording of neuronal activity with single cell resolution across the entire brain. Thanks to the use of optical methods, together with animal models in which the whole encephalon is optically accessible, this goal is getting within reach. In this work, we use a transgenic zebrafish line expressing the genetically encoded calcium indicator GCaMP6s in which binding of calcium ions leads to an increase in the emitted fluorescence of the reporter. GCaMP6s is the most sensitive calcium reporter within the genetically encoded calcium indicator family and allows us to record zebrafish larva neuronal activity with a high signal-to-noise ratio and single neuron resolution. To record the fluorescence emitted by the GCaMP6s reporter we use a custom-made confocal light-sheet microscope (LSM), in which the sample is illuminated with a thin sheet of light and the detection optical axis is perpendicular to the illumination axis. Owing to its intrinsic optical sectioning, this technique provides cellular resolution with high frame-rates and low photobleaching, allowing us to record the neuronal activity of zebrafish larvae with high spatio-temporal resolution. Conventional one-sided illumination LSM can suffer from limitations arising from even low levels of pigmentation in the sample or the presence of other obstructions reducing the quality of the incoming excitation light sheet. This can lead to shadowing in the image and possibly dynamic artefacts when the components responsible for the optical perturbations move. We have therefore implemented an illumination system based on a Bessel beam to overcome these limitations. Due to their nondiffractive and “self-healing” properties Bessel beams improve the quality of the images obtained from zebrafish larvae, reducing shadowing effects and increasing image homogeneity.

Light-sheet microscopy (LSM) has proven a useful tool in neuroscience to image whole brains with high frame rates at cellular resolution. LSM is employed either in combination with tissue clearing to reconstruct the cyto-architecture over the entire mouse brain or with intrinsically transparent samples like zebrafish larvae for functional imaging. Inherently to LSM, however, residual opaque objects cause stripe artifacts, which obscure features of interest and, during functional imaging, modulate fluorescence variations related to neuronal activity. Here, we report how Bessel beams reduce streaking artifacts and produce high-fidelity structural data. Furthermore, using Bessel beams, we demonstrate a fivefold increase in sensitivity to calcium transients and a 20-fold increase in accuracy in the detection of activity correlations in functional imaging. Our results demonstrate the contamination of data by systematic and random errors through Gaussian illumination and furthermore quantify the increase in fidelity of such data when using Bessel beams.

Artificial intelligence techniques are excessively used in computing for training, forecasting and evaluation purposes. Among these techniques, artificial neural network (ANN) is widely used for developing prediction models. ANNs use various Meta-heuristic algorithms including approximation methods for training the neural networks. ANN plays a significant role in this area and can be helpful in determining the neural network input coefficient. The main goal of presented study is to train the neural network using meta-heuristic approaches and to enhance the perceptron neural network precision. In this article, we used an integrated algorithm to determine the neural network input coefficients. Later, the proposed algorithm was compared with other algorithms such as ant colony and invasive weed optimization for performance evaluation. The results reveal that the proposed algorithm results in more convergence with neural network coefficient as compared to existing algorithms. However the proposed method resulted in reduction of prediction error in the neural network.