Optogenetic technologies paved the way to dissect complex neural circuits and monitor neural activity using light in animals. In retinal disease, optogenetics has been used as a therapeutic modality to reanimate the retina after the loss of photoreceptor outer segments. However, it is not clear today which ones of the great diversity of microbial opsins are best suited for therapeutic applications in human retinas as cell lines, primary cell cultures and animal models do not predict expression patterns of microbial opsins in human retinal cells. Therefore, we sought to generate retinal organoids derived from human induced pluripotent stem cells (hiPSCs) as a screening tool to explore the membrane trafficking efficacy of some recently described microbial opsins. We tested both depolarizing and hyperpolarizing microbial opsins including CatCh, ChrimsonR, ReaChR, eNpHR 3.0, and Jaws. The membrane localization of eNpHR 3.0, ReaChR, and Jaws was the highest, likely due to their additional endoplasmic reticulum (ER) release and membrane trafficking signals. In the case of opsins that were not engineered to improve trafficking efficiency in mammalian cells such as CatCh and ChrimsonR, membrane localization was less efficient. Protein accumulation in organelles such as ER and Golgi was observed at high doses with CatCh and ER retention lead to an unfolded protein response. Also, cytoplasmic localization was observed at high doses of ChrimsonR. Our results collectively suggest that retinal organoids derived from hiPSCs can be used to predict the subcellular fate of optogenetic proteins in a human retinal context. Such organoids are also versatile tools to validate other gene therapy products and drug molecules.

is an open access repository that collects the work of Arts et Métiers ParisTech researchers and makes it freely available over the web where possible. An analysis of high cycle multiaxial fatigue behaviour is conducted through the numerical simulation of polycrystalline aggregates using the finite element method. The metallic material chosen for investigation is pure copper, which has a Face Centred Cubic (FCC) crystalline microstructure. The elementary volumes are modelled in 2D using an hypothesis of generalised plane strain and consist of 300 equi-probability, randomly oriented grains with equiaxed geometry. The aggregates are loaded at levels equivalent to the average macroscopic fatigue strength at 10 7 cycles. The goal is to compute the mechanical quantities at the mesoscopic scale (i.e., average within the grain) after stabilization of the local cyclic behaviour. The results show that the mesoscopic mechanical variables are characterised by high dispersion. A statistical analysis of the response of the aggregates is undertaken for different loading modes: fully reversed tensile loads, torsion and combined in-phase tension-torsion. Via the calculation of the local mechanical quantities for a sufficiently large number of different microstructures, a critical analysis of certain multiaxial endurance criteria (Crossland, Dang Van and Matake) is conducted. In terms of material behaviour models, it is shown that elastic anisotropy strongly affects the scatter of the mechanical parameters used in the different criteria and that its role is predominant compared to that of crystal plasticity. The analysis of multiaxial endurance criteria at both the macroscopic and mesoscopic scales clearly show that the critical plane type criteria (Dang Van and Matake) give an adequate estimation of the shear stress but badly reflect the scatter of the normal stress or the hydrostatic stress.