The development of functional retinal imaging is of great interest to clinical and experimental ophthalmology because it should improve current clinical practice that relies on a simple evaluation of disease-related changes in retinal morphology. In this presentation, we will review our recent progress in the measurements and interpretation of Optoretinograms (ORG). The ORG comprises light-dependent changes in photoreceptor outer segment length and scattering measured in vivo with submicrometer-resolution by Optical Coherence Tomography (OCT). Our recent data confirm that the mechanisms underlying “slow” ORG response is that extreme phototransduction increases osmotic pressure in the outer segment, driving water entry and swelling to restore osmotic balance. Water movement between different retinal layers was quantified by OCT measurements, and analysis of retinal layer thickness dynamics provided estimates of the water permeability of key barriers between the Choriocapillaris (CC) and photoreceptors, viz., of the CC cell membrane, Bruch’s Membrane (BrM), the RPE layer, and the rod plasma membrane. A model proposed to link the changes in refractive index caused by the osmotically driven water movement between the CC lumen and the outer segment during the ORG predicts the observed BrM scattering changes. Overall, the results and modeling provide new insights into critical water barriers of the posterior eye and an experimental and theoretical toolkit for measuring their permeabilities.
In vivo longitudinal monitoring of the inner retinal cellular morphology is of great importance in both clinical and experimental ophthalmology due to its importance in many blinding diseases, including glaucoma, the second leading cause of blindness in the adult population worldwide. At the cellular level, Ganglion Cells (GCs) damage and, ultimately, death often plays a central role in inner retinal disease progression. Recently, advances in OCT-based observation of highly translucent cell somas in the Ganglion Cell Layer (GCL) in both the living human and experimental animal eyes opened the door to noninvasive, label-free monitoring of RGC in vivo. However, a longitudinal validation of OCT's ability to follow individual RGC in experimental animals was still needed. To address this, we will report on our quantitative longitudinal studies in mice lines with fluorescently labeled ganglion cells. Here we used our custom-built mouse retinal Scanning Laser Ophthalmoscopy / Optical Coherence Tomography (SLO/OCT) system to acquire serial OCT volumes (with corresponding SLO intensity and fluorescence data) to provide input for Temporal Speckle Averaging (TSA) OCT volume processing method. To allow in vivo validation of TSA-OCT-based RGC quantification and monitoring, two mouse lines with fluorescently labeled RGC based on RGCs transcription factor (Brn3b-mCherry and Isl2-GFP) have been used and imaged simultaneously with fluorescence SLO (fSLO).
Development of high resolution functional retinal imaging tools is of great interest to clinical and experimental ophthalmology because the alterations in retinal function, at cellular resolution, hold the promise of being more sensitive for disease diagnostic then the purely retinal morphology-based assays. In this work we present our initial design and implementation of mouse retinal imaging system that incorporates full-field (FF) swept- source (SS) optical coherence tomography (OCT) with dedicated light stimulation channel for high-speed measurements of light evoked responses in photoreceptors of mice. The Optoretinograpahy (ORG) results acquired with our FF-SS-OCT system are compared with those acquired with our standard raster scanning mouse ORG-OCT system.
The development of functional retinal imaging is of great interest to clinical and experimental ophthalmology, because it should provide more sensitive tools for ocular diseases diagnostic that would go beyond current gold standard of simple evaluation of the static retinal morphology. In this presentation we will review our recent progress in measurements and interpretation of OCT-based optoretinograms (ORG) i.e., the paradigm of using NIR OCT to measure in vivo bleaching-induced changes in retinal morphology (transient changes in volume of individual neurons, or thickness of retinal layers). Specifically, comparison between different instrumentations used to acquire ORGs and between results acquired using clinical (human) and experimental (animal) systems will be presented. Additionally, intensity-based and phase-based ORG extraction framework will be presented. Finally, we will discuss our findings in the context of current understanding of measured process, being a result osmotically driven water movements between the photoreceptors, and other retinal neurons and its surroundings.
The development of functional retinal imaging is of great interest to clinical and experimental ophthalmology. Recent progress in OCT-based functional imaging of rods and/or cones outer segments elongations in response to stimuli that bleach fraction of their rhodopsin promises to address the need for probing photoreceptor function. Building on this model, we developed the framework to monitor changes in the gross morphology of the whole retina of mice in response to light stimulation. In our current submission, we will discuss our findings in the context of known retinal water barriers restricting water movements between inner and outer retina.
A combined scattering and autofluorescence (AF) measurement from the retinal pigment epithelium (RPE) was investigated to study the changes in the AF granules in an Abca4-/- mouse (murine model of Stargardt disease). The directional scattering was measured with an optical coherence tomography (OCT), whereas the multi-color fundus AF spectra was measured with a spectrometer-integrated scanning laser ophthalmoscope. Increased scattering from the RPE in Abca4-/- mouse relative to control was well correlated with the elevated AF spectra and indicated the changes in RPE fluorophores. Ex vivo studies based on scanning confocal and electron microscopy were performed to validate the in vivo findings.
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