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Jesse Schallek, Aby Joseph, Vigneshwar Subramanian, Andres Guevara-Torres; Imaging invisible cells: new advances in adaptive optics reveal structure of the translucent retinal cells of the inner retina. Journal of Vision 2017;17(7):2. doi: https://doi.org/10.1167/17.7.2.
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© ARVO (1962-2015); The Authors (2016-present)
While the advent of adaptive optics ophthalmoscopy has provided microscopic resolution in the living eye, the majority of neurons in the mammalian retina have evaded detection. This is because most retinal cells are highly translucent, allowing photons to travel through the neural retina with minimal scatter and absorption before reaching the photoreceptors. While retinal translucency is beneficial for vision, it poses a challenge for imaging because cells provide weak optical contrast.
Recent advances in non-confocal ophthalmoscopy now reveal cells that were once hidden by their translucency. By comparing directional cell scatter, we provide detailed images of: ganglion cells, horizontal cells, multi-laminar photoreceptor somata, red blood cells, platelets, white blood cells and putative sub-cellular organelles in the living eye without contrast agents.
We optimized the non-confocal contrast by integrating “split-detection” capabilities (Scoles et al. 2014) into an adaptive optics scanning light ophthalmoscope (AOSLO) custom built for the mouse eye (Guevara-Torres et al. 2015). The split-detection approach compares directional light scatter in the retina. The imaged point spread function of the AOSLO was bisected by a knife-edge prism and relayed into two, phase-locked photomultiplier tubes. The left and right half of the imaged retinal point spread function was digitally subtracted to remove the mutual light information common in the two channels. C57BL/6J mice were anesthetized and imaged with and without contrast agents applied to validate cell type.
Imaged ganglion cells were confirmed by imaging tagged Thy-1 fluorescent cells. Leukocytes were confirmed with fluorescent labeling of nuclei with acridine orange. Red blood cells, platelets, photoreceptor somata and horizontal cells matched the known morphology, density and/or topography in the mouse retina. This new catalog of cells has greatly expanded the number of cell types that may be studied in the living mammalian retina in health and disease.
Meeting abstract presented at the 2016 OSA Fall Vision Meeting
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