In vertebrate eyes, vision begins when the photoreceptor's outer segment absorbs photons and generates a neural signal destined for the brain. The extreme optical and metabolic demands of this process of phototransduction necessitate continual renewal of the outer segment. Outer segment renewal has been long studied in post-mortem rods using autoradiography, but has been observed neither in living photoreceptors nor directly in cones. Using adaptive optics, which permits the resolution of cones, and temporally coherent illumination, which transforms the outer segment into a “biological interferometer,” we observed cone renewal in three subjects, manifesting as elongation of the cone outer segment, with rates ranging from 93 to 113 nm/hour (2.2 to 2.7 μm/day). In one subject we observed renewal occurring over 24 hours, with small but significant changes in renewal rate over the day. We determined that this novel method is sensitive to changes in outer segment length of 139 nm, more than 20 times better than the axial resolution of ultra-high resolution optical coherence tomography, the best existing method for depth imaging of the living retina.
Ultrahigh speed line scan detectors based on CMOS technology have been recently demonstrated in ultrahigh resolution spectral-domain optical coherence tomography (UHR-SD-OCT) for retinal imaging. While successful, fundamental tradeoffs exist been image acquisition time, image sampling density, and sensitivity, all of which impact the extent of motion artifacts, visualization of fine spatial detail, and detection of faint reflections. Here we investigate these tradeoffs for imaging retinal nerve fiber bundles (RNFBs) using UHR-SD-OCT with adaptive optics (AO). Volume scans of 3°x3° and 1.5°x1.5° were acquired at retinal locations of 3° nasal and 6° superior to the fovea on a healthy subject. Dynamic AO compensation across a 6 mm pupil provided near-diffraction-limited performance. The acquisition rates were 22.5k lines/s and 125k lines/s with A-lines spaced at 0.9 μm and 1.8 μm and B-scans at 1.8 μm and 9 μm. Focus was optimized for visualizing the retinal nerve fiber bundles (RNFBs). En face projection and crosssectional views of the RNFBs were extracted from the volumes and compared to images acquired with established conventional CCD-based line-scan camera. The projection view was found highly sensitive to eye motion artifacts, yet could only be partially compensated with coarser sampling, since fine sampling was necessary to observe the microscopic features in the RNFBs. For the cross-sectional view, speckle noise rather than eye motion artifacts limited bundle clarity. The highest B-scan density (1.8 μm spacing) coupled with B-scan averaging proved the best combination. Regardless of view, the higher line rate provided better RNFB clarity.
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