Cones in ageing and harsh environments: the neural economy hypothesis.

PURPOSE: Cones are at great risk in a wide variety of retinal diseases, especially when there is a harsh microenvironment and retinal pigment epithelium is damaged. We provide established and new methods for assessing cones and retinal pigment epithelium, together with new results. We investigated conditions under which cones can be imaged and could guide light, despite the proximity of less than ideal retinal pigment epithelium. RECENT FINDINGS: We used a variety of imaging methods to detect and localise damage to the retinal pigment epithelium. As age-related macular degeneration is a particularly widespread disease, we imaged clinical hallmarks: drusen and hyperpigmentation. Using near infrared light provided improved imaging of the deeper fundus layers. We compared confocal and multiply scattered light images, using both the variation of detection apertures and polarisation analysis. We used optical coherence tomography to examine distances between structures and thickness of retinal layers, as well as identifying damage to the retinal pigment epithelium. We counted cones using adaptive optics scanning laser ophthalmoscopy. We compared the results of five subjects with geographic atrophy to data from a previous normative ageing study. Using near infrared imaging and layer analysis of optical coherence tomography, the widespread aspect of drusen became evident. Both multiply scattered light imaging and analysis of the volume in the retinal pigment epithelial layer from the optical coherence tomography were effective in localising drusen and hyperpigmentation beneath the photoreceptors. Cone photoreceptors in normal older eyes were shorter than in younger eyes. Cone photoreceptors survived in regions of atrophy, but with greatly reduced and highly variable density. Regular arrays of cones were found in some locations, despite abnormal retinal pigment epithelium. For some subjects, the cone density was significantly greater than normative values in some retinal locations outside the atrophy. SUMMARY: The survival of cones within atrophy is remarkable. The unusually dense packing of cones at some retinal locations outside the atrophy indicates more fluidity in cone distribution than typically thought. Together these findings suggest strategies for therapy that includes preserving cones.

Central Macular Thickness in Diabetic Patients: A Sex-based Analysis.

SIGNIFICANCE: The pathological changes in clinically significant diabetic macular edema lead to greater retinal thickening in men than in women. Therefore, male sex should be considered a potential risk factor for identifying individuals with the most severe pathological changes. Understanding this excessive retinal thickening in men may help preserve vision. PURPOSE: The purpose of this study was to investigate the sex differences in retinal thickness in diabetic patients. We tested whether men with clinically significant macular edema had even greater central macular thickness than expected from sex differences without significant pathological changes. This study also aimed to determine which retinal layers contribute to abnormal retinal thickness. METHODS: From 2047 underserved adult diabetic patients from Alameda County, CA, 142 patients with clinically significant macular edema were identified by EyePACS-certified graders using color fundus images (Canon CR6-45NM). First, central macular thickness from spectral domain optical coherence tomography (iVue; Optovue Inc.) was compared in 21 men versus 21 women without clinically significant macular edema. Then, a planned comparison contrasted the greater values of central macular thickness in men versus women with clinically significant macular edema as compared with those without. Mean retinal thickness and variability of central macular layers were compared in men versus women. RESULTS: Men without clinically significant macular edema had a 12-µm greater central macular thickness than did women (245 ± 21.3 and 233 ± 13.4 µm, respectively; t40 = -2.18, P = .04). Men with clinically significant macular edema had a 67-µm greater central macular thickness than did women (383 ± 48.7 and 316 ± 60.4 µm, P < .001); that is, men had 55 µm or more than five times more (t20 = 2.35, P = .02). In men, the outer-nuclear-layer thickness was more variable, F10,10 = 9.34. CONCLUSIONS: Underserved diabetic men had thicker retinas than did women, exacerbated by clinically significant macular edema.

Near-infrared polarimetric imaging and changes associated with normative aging.

With aging, the human retina undergoes cell death and additional structural changes that can increase scattered light. We quantified the effect of normative aging on multiply scattered light returning from the human fundus. As expected, there was an increase of multiply scattered light associated with aging, and this is consistent with the histological changes that occur in the fundus of individuals before developing age-related macular degeneration. This increase in scattered light with aging cannot be attributed to retinal reflectivity, anterior segment scatter, or pupil diameter.

Subtle changes in diabetic retinas localised in 3D using OCT.

PURPOSE: To detect and localise subtle changes in retinas of diabetic patients who clinically have no diabetic retinopathy (DR) or non-proliferative DR (NPDR) as compared to age- and sex- matched controls. Spectral Domain Optical Coherence Tomography (SD-OCT) and software to examine all retinal layers, including deeper layers, were used to quantify foveal avascular zone size and inner and outer retinal layer thicknesses, as well as to detect axial location of prominent lesions. METHODS: Diabetic subjects, 19 total with 16 having no DR and three having non-proliferative retinopathy, were matched with 19 controls with respect to age and sex. Macular-centred SD-OCT grids of 20 × 15° were taken with the Spectralis. En face or transverse images were generated from the SD-OCT data by automatically segmenting all retinal layers. The transverse images were investigated for foveal avascular zone (FAZ) size, retinal vessel calibre, and structural changes. The size of the FAZ was compared for diabetics vs controls using vendor software and manual marking in Photoshop. Inner retinal layer (IRLFAZ ) and outer nuclear layer (ONLFAZ ) thicknesses at the margins of the FAZ were measured using vendor software. RESULTS: The FAZ area was larger for diabetics (mean ± S.D. = 0.388 ± 0.074 mm2 ) than controls (0.243 ± 0.113 mm2 ), t18 = 5.27, p < 0.0001, using vendor software. The mean IRLFAZ was thicker for the diabetics (86.8 ± 14.5 µm) than controls (65.2 ± 16.3 µm), t18 = 4.59, p = 0.00023, despite lack of exudation by clinical exam. There was no significant association between FAZ area and mean IRLFAZ for the diabetics, r = 0.099, p = 0.69. Vessels not clinically detected were visible in the NFL transverse image of most diabetics, especially for a mild NPDR patient. A prominent lesion found in the en face infra-red image of a mild NPDR subject was localised in the photoreceptor layer by SD-OCT, as well as additional outer retinal changes in other subjects. CONCLUSIONS: Our results demonstrate changes in inner and outer diabetic retinas not readily detectable by clinical exam. IRLFAZ had not thinned at the margins of the large FAZs, indicating neural mass did not yet decrease despite potential ischemia.

SD-OCT and Adaptive Optics Imaging of Outer Retinal Tubulation.

PURPOSE: To investigate outer retinal tubulation (ORT) using spectral domain optical coherence tomography (SD-OCT) and an adaptive optics scanning laser ophthalmoscope (AOSLO). To document the frequency of ORT in atrophic retinal conditions and quantify ORT dimensions versus adjacent retinal layers. METHODS: SD-OCT images were reviewed for the presence of retinal atrophy, scarring, and/or exudation. The greatest width of each ORT was quantified. Inner and outer retinal thicknesses adjacent to and within the area of ORT were measured for 18 patients. AOSLO imaged ORTs in five subjects with direct and scattered light imaging. RESULTS: ORT was identified in 47 of 76 subjects (61.8%) and in 65 eyes via SD-OCT in a wide range of conditions and ages, and in peripapillary atrophy. ORTs appeared as finger-like projections in atrophy, seen in the en face images. AOSLO showed some ORTs with bright cones that guide light within atrophic areas. Multiply scattered light mode AOSLO visualized variegated lines (18-35 µm) radiating from ORTs. The ORTs' width on OCT b-scan images varied from 70 to 509 µm. The inner retina at the ORT was significantly thinner than the adjacent retina, 135 vs.170 µm (P = .004), whereas the outer retina was significantly thicker, 115 vs. 80 µm (P = .03). CONCLUSIONS: ORTs are quite common in eyes with retinal atrophy in various disorders. ORTs demonstrate surviving photoreceptors in tubular structures found within otherwise nonsupportive atrophic areas that lack retinal pigment epithelium and choriocapillaris.

Fixation stability and scotoma mapping for patients with low vision.

PURPOSE: To develop a simplified device that performs fundus perimetry techniques such as fixation mapping and kinetic perimetry. METHODS: We added visual stimulation to a near-infrared retinal imager, the laser scanning digital camera (LSDC). This device uses slit scanning illumination combined with a two-dimensional CMOS (complementary metal oxide semiconductor) detector, with continuous viewing of the retina. The CMOS readout was synchronized with the slit scanning, thereby serving as a confocal aperture to reduce stray light in retinal images. A series of retinal images of 36 degrees was automatically aligned to provide data for fixation maps and quantification of fixation stability. The LSDC and alignment techniques also provided fundus viewing with retinal location correction for scotoma mapping. RESULTS: First, fixation mapping was readily performed in patients with central scotoma or amblyopia. The automatic alignment algorithm allowed quantification of fixation stability in patients with macular pathologies that did not cause scotoma. Second, fixation stability was rapidly and quantitatively assessed by the automatic registration of the series of retina images. There was no significant difference in the fixation stability with automatic versus manual alignment. Kinetic perimetry demonstrated that fundus imaging helped reduce the variability of perimetric data by identifying and preventing false-positives caused by eye motion. We found that the size of the blind spot was significantly larger for dark targets on brighter backgrounds than when the contrast was reversed (p < 0.045). This is consistent with incremental targets being detected partially or wholly because of scattered light falling on more sensitive retinal locations. CONCLUSIONS: Fundus perimetry with the LSDC allows for a wide range of fixation and perimetry tasks.

Potential vision tester using adaptive optics, Maxwellian view, and small pupil.

We demonstrate a free-space, trolley-mounted potential vision tester (PVT), designed to study and improve the accuracy of visual acuity (VA) measurements in the aging eye. Key features include a high-resolution visual display presented in Maxwellian view, a 3 mm pupil to limit wavefront (WF) aberrations, and a moderate cost deformable mirror to induce or correct higher order optical aberrations. The visual display supported accurate measurement of visual acuities down to 20/5. The moderate cost, piezo deformable mirror induced seven nominal aberrations, calibrated as 0, -0.32, -0.23, + 0.27, and +0.39 microns spherical aberration; + 0.49 microns Y coma; and -0.51 microns X coma. A custom Hartmann Shack (HS) calibration (HSc) system demonstrated that induced aberrations were repeatable and stable. A Badal optometer provided the coarse focus. WF aberrations were measured for five normal subjects with a commercially available HS device (HSP) (OCULUS Pentacam AXL Wave), providing estimates of WF errors for 3 mm and other pupil sizes. VA was measured using four alternative forced-choice for a single black on white E stimulus in each trial. Using the method of constant stimuli yielded robust standard deviation measurements. The 50% fit for VA plotted against induced aberration resulted in linear functions for each subject for the range of our positive and negative spherical aberration data. Subjects differed, but higher order terms were unnecessary to describe data across spherical aberrations.

Cone Photoreceptors in Diabetic Patients.

Purpose: Cones in diabetic patients are at risk due to metabolic and vascular changes. By imaging retinal vessel modeling at high magnification, we reduced its impact on cone distribution measurements. The retinal vessel images and retinal thickness measurements provided information about cone microenvironment. Methods: We compared cone data in 10 diabetic subjects (28-78 yr) to our published norms from 36 younger and 10 older controls. All subjects were consented and tested in a manner approved by the Indiana University Institutional Review Board, which adhered to the Declaration of Helsinki. Custom adaptive optics scanning laser ophthalmoscopy (AOSLO) was used to image cones and retinal microcirculation. We counted cones in a montage of foveal and temporal retina, using four non-contiguous samples within 0.9-7 deg that were selected for best visibility of cones and least pathology. The data were fit with a two parameter exponential model: ln(cone density) = a * microns eccentricity + b. These results were compared to retinal thickness measurements from SDOCT. Results: Diabetic cone maps were more variable than in controls and included patches, or unusually bright and dark cones, centrally and more peripherally. Model parameters and total cones within the central 14 deg of the macula differed across diabetic patients. Total cones fell into two groups: similar to normal for 5 vs. less than normal for 2 of 2 younger diabetic subjects and 3 older subjects, low but not outside the confidence limits. Diabetic subjects had all retinal vascular remodeling to varying degrees: microaneurysms; capillary thickening, thinning, or bends; and vessel elongation including capillary loops, tangles, and collaterals. Yet SD-OCT showed that no diabetic subject had a Total Retinal Thickness in any quadrant that fell outside the confidence limits for controls. Conclusions: AOSLO images pinpointed widespread retinal vascular remodeling in all diabetic eyes, but the SDOCT showed no increased retinal thickness. Cone reflectivity changes were found in all diabetic patients, but significantly low cone density in only some. These results are consistent with early changes to neural, glial, or vascular components of the retinal without significant retinal thickening due to exudation.

Quantifying frequency content in cross-sectional retinal scans of diabetics vs. controls.

PURPOSE: To examine subtle differences in the structure of diabetic vs. control retinas. METHODS: Spectral-domain optical coherence tomography (SD-OCT) images were compared for the retinas of 33 diabetic subjects who did not have clinical evidence of diabetic macular edema and age-matched controls, with central macular thicknesses of 275 and 276 microns, respectively. Cross-sectional retinal images through the fovea, called B-scans, were analyzed for spatial frequency content. The B-scans were processed to remove and smooth the portions of the retinal image not within regions of interest in the retina. The remaining retinal images were then quantified using a Fast Fourier Transform (FFT) approach that provided amplitude as a function of spatial frequency. RESULTS: The FFT analysis showed that diabetic retinas had spatial frequency content with significantly higher power compared to control retinas particularly for a deeper fundus layer at mid-range spatial frequencies, ranging from p = 0.0030 to 0.0497 at 16.8 to 18.2 microns/cycle. There was lower power at higher spatial frequencies, ranging from p = 0.0296 and 0.0482 at 27.4 and 29.0 microns/cycle. The range of mid-range frequencies corresponds to the sizes of small blood vessel abnormalities and hard exudates. Retinal thickness did not differ between the two groups. CONCLUSIONS: Diabetic retinas, although not thicker than controls, had subtle but quantifiable pattern changes in SD-OCT images particularly in deeper fundus layers. The size range and distribution of this pattern in diabetic eyes were consistent with small blood vessel abnormalities and leakage of lipid and fluid. Feature-based biomarkers may augment retinal thickness criteria for management of diabetic eye complications, and may detect early changes.

Distances From Capillaries to Arterioles or Venules Measured Using OCTA and AOSLO.

Purpose: To investigate distances from retinal capillaries to arterioles or venules noninvasively. Methods: An adaptive optics scanning laser ophthalmoscope (AOSLO) and optical coherence tomography angiography (OCTA) imager acquired detailed maps of retinal vasculature. Using OCTA, we quantified the distance from the edge of an arteriole or venule to the middle of the nearest capillaries (periarteriole or perivenule capillary-free zones, respectively) within the superficial vascular plexus of 20 young healthy subjects with normal axial lengths. These distances were compared to AOSLO images for three subjects. We tested the relation between the peripheral capillary-free zones and FAZ horizontal, vertical, effective diameters, and asymmetry indices in the deep vascular plexus. We examined enlargement with OCTA of capillary-free zones in a type 2 diabetic patient. Results: The periarteriole capillary-free zone (67.2 ± 25.3 µm) was readily visible and larger than the perivenule capillary-free zone (42.7 ± 14.4 µm), F(1, 998) = 771, P < 0.0001. The distance from foveal center (P = 0.003) and diameter (P = 0.048) were predictive of perivenule capillary-free zone values. OCTA and AOSLO corresponded for arterioles. FAZ effective diameter was positively associated with asymmetry indices, r = 0.49, P = 0.028, but not peripheral capillary-free zones, although focal enlargements were found in a diabetic patient. Conclusions: For normal retinas, periarteriole and perivenule capillary-free zones are readily visible with OCTA and AOSLO. Periarteriole capillary-free zones were larger, consistent with arterioles carrying oxygen rich blood that diffuses to support the retina.

Distribution differences of macular cones measured by AOSLO: Variation in slope from fovea to periphery more pronounced than differences in total cones.

Large individual differences in cone densities occur even in healthy, young adults with low refractive error. We investigated whether cone density follows a simple model that some individuals have more cones, or whether individuals differ in both number and distribution of cones. We quantified cones in the eyes of 36 healthy young adults with low refractive error using a custom adaptive optics scanning laser ophthalmoscope. The average cone density in the temporal meridian was, for the mean±SD, 43,216±6039, 27,466±3496, 14,996±1563, and 12,207±1278cones/mm2 for 270, 630, 1480, and 2070µm from the foveal center. Cone densities at 630µm retinal eccentricity were uncorrelated to those at 2070µm, ruling out models with a constant or proportional relation of cone density to eccentricity. Subjects with high central macula cone densities had low peripheral cone densities. The cone density ratio (2070:630µm) was negatively correlated with cone density at 630µm, consistent with variations in the proportion of peripheral cones migrating towards the center. We modelled the total cones within a central radius of 7deg, using the temporal data and our published cone densities for temporal, nasal, superior, and inferior meridians. We computed an average of 221,000 cones. The coefficient of variation was 0.0767 for total cones, but higher for samples near the fovea. Individual differences occur both in total cones and other developmental factors related to cone distribution.

Cone photoreceptor packing density and the outer nuclear layer thickness in healthy subjects.

PURPOSE: We evaluated the relationship between cone photoreceptor packing density and outer nuclear layer (ONL) thickness within the central 15 degrees. METHODS: Individual differences for healthy subjects in cone packing density and ONL thickness were examined in 8 younger and 8 older subjects, mean age 27.2 versus 56.2 years. Cone packing density was obtained using an adaptive optics scanning laser ophthalmoscope (AOSLO). The ONL thickness measurements included the ONL and the Henle fiber layer (ONL + HFL), and were obtained using spectral domain optical coherence tomography (SDOCT) and custom segmentation software. RESULTS: There were sizeable individual differences in cone packing density and ONL + HFL thickness. Older subjects had on average lower cone packing densities, but thicker ONL + HFL measurements. Cone packing density and ONL + HFL thickness decreased with increasing retinal eccentricity. The ratio of the cone packing density-to-ONL2 was larger for the younger subjects group, and decreased with retinal eccentricity. CONCLUSIONS: The individual differences in cone packing density and ONL + HFL thickness are consistent with aging changes, indicating that normative aging data are necessary for fine comparisons in the early stages of disease or response to treatment. Our finding of ONL + HFL thickness increasing with aging is inconsistent with the hypothesis that ONL measurements with SDOCT depend only on the number of functioning cones, since in our older group cones were fewer, but thickness was greater.