Austin Roorda

The arrangement of the three cone classes in the living human eye

Human colour vision depends on three classes of receptor, the short- (S), medium- (M), and long- (L) wavelength-sensitive cones. These cone classes are interleaved in a single mosaic so that, at each point in the retina, only a single class of cone samples the retinal image. As a consequence, observers with normal trichromatic colour vision are necessarily colour blind on a local spatial scale. The limits this places on vision depend on the relative numbers and arrangement of cones. Although the topography of human S cones is known,, the human L- and M-cone submosaics have resisted analysis. Adaptive optics, a technique used to overcome blur in ground-based telescopes, can also overcome blur in the eye, allowing the sharpest images ever taken of the living retina. Here we combine adaptive optics and retinal densitometry to obtain what are, to our knowledge, the first images of the arrangement of S, M and L cones in the living human eye. The proportion of L to M cones is strikingly different in two male subjects, each of whom has normal colour vision. The mosaics of both subjects have large patches in which either M or L cones are missing. This arrangement reduces the eyes ability to recover colour variations of high spatial frequency in the environment but may improve the recovery of luminance variations of high spatial frequency.

Adaptive optics scanning laser ophthalmoscopy.

We present the first scanning laser ophthalmoscope that uses adaptive optics to measure and correct the high order aberrations of the human eye. Adaptive optics increases both lateral and axial resolution, permitting axial sectioning of retinal tissue in vivo. The instrument is used to visualize photoreceptors, nerve fibers and flow of white blood cells in retinal capillaries.

A population study on changes in wave aberrations with accomodation

Wave aberrations were measured with a Shack-Hartmann wavefront sensor (SHWS) in the right eye of a large young adult population when accommodative demands of 0, 3, and 6 D were presented to the tested eye through a Badal system. Three SHWS images were recorded at each accommodative demand and wave aberrations were computed over a 5-mm pupil (through 6th order Zernike polynomials). The accommodative response was calculated from the Zernike defocus over the central 3-mm diameter zone. Among all individual Zernike terms, spherical aberration showed the greatest change with accommodation. The change of spherical aberration was always negative, and was proportional to the change in accommodative response. Coma and astigmatism also changed with accommodation, but the direction of the change was variable. Despite the large inter-subject variability, the population average of the root mean square for all aberrations (excluding defocus) remained constant for accommodative levels up to 3.0 D. Even though aberrations change with accommodation, the magnitude of the aberration change remains less than the magnitude of the uncorrected aberrations, even at high accommodative levels. Therefore, a typical eye will benefit over the entire accommodative range (0-6 D) if aberrations are corrected for distance viewing.

Longitudinal Study of Cone Photoreceptors during Retinal Degeneration and in Response to Ciliary Neurotrophic Factor Treatment

PURPOSE To study cone photoreceptor structure and function in patients with inherited retinal degenerations treated with sustained-release ciliary neurotrophic factor (CNTF). METHODS Two patients with retinitis pigmentosa and one with Usher syndrome type 2 who participated in a phase 2 clinical trial received CNTF delivered by an encapsulated cell technology implant in one eye and sham surgery in the contralateral eye. Patients were followed longitudinally over 30 to 35 months. Adaptive optics scanning laser ophthalmoscopy (AOSLO) provided high-resolution images at baseline and at 3, 6, 12, 18, and 24 months. AOSLO measures of cone spacing and density and optical coherence tomography measures of retinal thickness were correlated with visual function, including visual acuity (VA), visual field sensitivity, and full-field electroretinography (ERG). RESULTS No significant changes in VA, visual field sensitivity, or ERG responses were observed in either eye of the three patients over 24 months. Outer retinal layers were significantly thicker in CNTF-treated eyes than in sham-treated eyes (P < 0.005). Cone spacing increased by 2.9% more per year in sham-treated eyes than in CNTF-treated eyes (P < 0.001, linear mixed model), and cone density decreased by 9.1%, or 223 cones/degree(2) more per year in sham-treated than in CNTF-treated eyes (P = 0.002, linear mixed model). CONCLUSIONS AOSLO images provided a sensitive measure of disease progression and treatment response in patients with inherited retinal degenerations. Larger studies of cone structure using high-resolution imaging techniques are urgently needed to evaluate the effect of CNTF treatment in patients with inherited retinal degenerations.

Packing arrangement of the three cone classes in primate retina

We describe a detailed analysis of the spatial arrangement of L, M and S cones in the living eyes of two humans and one monkey. We analyze the cone mosaics near 1 degrees eccentricity using statistical methods that characterize the arrangement of each type of cone in the mosaic of photoreceptors. In all eyes, the M and L cones are arranged randomly. This gives rise to patches containing cones of a single type. In human, but not in monkey, the arrangement of S-cones cannot be distinguished from random.

Revealing Henle's Fiber Layer Using Spectral Domain Optical Coherence Tomography

PURPOSE Spectral domain optical coherence tomography (SD-OCT) uses infrared light to visualize the reflectivity of structures of differing optical properties within the retina. Despite their presence on histologic studies, traditionally acquired SD-OCT images are unable to delineate the axons of photoreceptor nuclei, Henles fiber layer (HFL). The authors present a new method to reliably identify HFL by varying the entry position of the SD-OCT beam through the pupil. METHODS Fifteen eyes from 11 subjects with normal vision were prospectively imaged using 1 of 2 commercial SD-OCT systems. For each eye, the entry position of the SD-OCT beam through the pupil was varied horizontally and vertically. The reflectivity of outer retinal layers was measured as a function of beam position, and thicknesses were recorded. RESULTS The reflectivity of HFL was directionally dependent and increased with eccentricity on the side of the fovea opposite the entry position. When HFL was included in the measurement, the thickness of the outer nuclear layer (ONL) of central horizontal B-scans increased by an average of 52% in three subjects quantified. Four cases of pathology, in which alterations to the normal macular geometry affected HFL intensity, were identified. CONCLUSIONS The authors demonstrated a novel method to distinguish HFL from true ONL. An accurate measurement of the ONL is critical to clinical studies measuring photoreceptor layer thickness using any SD-OCT system. Recognition of the optical properties of HFL can explain reflectivity changes imaged in this layer in association with macular pathology.

Automated identification of cone photoreceptors in adaptive optics retinal images

In making noninvasive measurements of the human cone mosaic, the task of labeling each individual cone is unavoidable. Manual labeling is a time-consuming process, setting the motivation for the development of an automated method. An automated algorithm for labeling cones in adaptive optics (AO) retinal images is implemented and tested on real data. The optical fiber properties of cones aided the design of the algorithm. Out of 2153 manually labeled cones from six different images, the automated method correctly identified 94.1% of them. The agreement between the automated and the manual labeling methods varied from 92.7% to 96.2% across the six images. Results between the two methods disagreed for 1.2% to 9.1% of the cones. Voronoi analysis of large montages of AO retinal images confirmed the general hexagonal-packing structure of retinal cones as well as the general cone density variability across portions of the retina. The consistency of our measurements demonstrates the reliability and practicality of having an automated solution to this problem.

Optical fiber properties of individual human cones

The tuning properties of the ensemble of cone photoreceptors is due to the tuning properties of individual cones convolved with the disarray in pointing direction between the cones. We used direct imaging with the Rochester adaptive optics ophthalmoscope to directly image these properties in individual cones in living human eyes. We found that cone disarray is very small, accounting for less than 1% of the breadth of the tuning function of an ensemble of cones. The implication is that the optical fiber properties of an ensemble of cones mimic the tuning properties of a single cone.

Noninvasive visualization and analysis of parafoveal capillaries in humans.

PURPOSE To demonstrate a noninvasive method to visualize and analyze the parafoveal capillary network in humans. METHODS An adaptive optics scanning laser ophthalmoscope was used to acquire high resolution retinal videos on human subjects. Video processing tools that enhance motion contrast were developed and applied to the videos to generate montages of parafoveal retinal capillaries. The capillary network and foveal avascular zone (FAZ) were extracted using video and image analysis algorithms. The capillary densities in the zone immediately outside the FAZ were calculated and the variation in density as a function of direction was investigated. Extracted FAZ geometries were used to calculate area and effective diameters. The authors also compared their method against fluorescein angiography (FA) for one subject. RESULTS The parafoveal capillaries were clearly visible when the motion contrast in noninvasive videos was enhanced. There was a marked improvement in the contrast of the parafoveal capillaries when compared to the unprocessed videos. The average FAZ area was 0.323 mm(2), with an average effective diameter of 633 microm. There was no variation in capillary density near the FAZ in different directions. CONCLUSIONS Using motion cues to enhance vessel contrast is a powerful tool for visualizing the capillary network, in the absence of contrast agents. The authors demonstrate a tool to study the microcirculation of healthy subjects noninvasively.

MEMS-based adaptive optics scanning laser ophthalmoscopy

We have developed a compact, robust adaptive optics (AO) scanning laser ophthalmoscope using a microelectromechanical (MEMS) deformable mirror (DM). Facilitated with a Shack-Hartmann wavefront sensor, the MEMS-DM-based AO operates a closed-loop modal wave aberration correction for the human eye and reduces wave aberrations in most eyes to below 0.1 μm rms. Lateral resolution is enhanced, and images reveal a clear cone mosaic near the foveal center. The significant increase in throughput allows for a confocal pinhole whose diameter is less than the Airy disc of the collection lens, thereby fully exploiting the axial resolution capabilities of the system.