Purpose.: To study the variation of cone photoreceptor packing density across the retina in healthy subjects of different ages. Methods.: High-resolution adaptive optics scanning laser ophthalmoscope (AOSLO) systems were used to systematically image the retinas of two groups of subjects of different ages. Ten younger subjects (age range, 22–35 years) and 10 older subjects (age range, 50–65 years) were tested. Strips of cone photoreceptors, approximately 12° × 1.8° long were imaged for each of the four primary retinal meridians: superior, inferior, nasal, and temporal. Cone photoreceptors within the strips were counted, and cone photoreceptor packing density was calculated. Statistical analysis (three-way ANOVA) was used to calculate the interaction for cone photoreceptor packing density between age, meridian, and eccentricity. Results.: As expected, cone photoreceptor packing density was higher close to the fovea and decreased with increasing retinal eccentricity from 0.18 to 3.5 mm (∼0.6–12°). Older subjects had approximately 75% of the cone density at 0.18 mm (∼0.6°), and this difference decreased rapidly with eccentricity, with the two groups having similar cone photoreceptor packing densities beyond 0.5 mm retinal eccentricity on average. Conclusions.: Cone packing density in the living human retina decreases as a function of age within the foveal center with the largest difference being found at our most central measurement site. At all ages, the retina showed meridional difference in cone densities, with cone photoreceptor packing density decreasing faster with increasing eccentricity in the vertical dimensions than in the horizontal dimensions.

Adaptive Optics (AO) retinal imaging has provided revolutionary tools to scientists and clinicians for studying retinal structure and function in the living eye. From animal models to clinical patients, AO imaging is changing the way scientists are approaching the study of the retina. By providing cellular and subcellular details without the need for histology, it is now possible to perform large scale studies as well as to understand how an individual retina changes over time. Because AO retinal imaging is non-invasive and when performed with near-IR wavelengths both safe and easily tolerated by patients, it holds promise for being incorporated into clinical trials providing cell specific approaches to monitoring diseases and therapeutic interventions. AO is being used to enhance the ability of OCT, fluorescence imaging, and reflectance imaging. By incorporating imaging that is sensitive to differences in the scattering properties of retinal tissue, it is especially sensitive to disease, which can drastically impact retinal tissue properties. This review examines human AO retinal imaging with a concentration on the use of the Adaptive Optics Scanning Laser Ophthalmoscope (AOSLO). It first covers the background and the overall approaches to human AO retinal imaging, and the technology involved, and then concentrates on using AO retinal imaging to study the structure and function of the retina.

Retinal vascular diseases are a leading cause of blindness and visual disability. The advent of adaptive optics retinal imaging has enabled us to image the retinal vascular at cellular resolutions, but imaging of the vasculature can be difficult due to the complex nature of the images, including features of many other retinal structures, such as the nerve fiber layer, glial and other cells. In this paper we show that varying the size and centration of the confocal aperture of an adaptive optics scanning laser ophthalmoscope (AOSLO) can increase sensitivity to multiply scattered light, especially light forward scattered from the vasculature and erythrocytes. The resulting technique was tested by imaging regions with different retinal tissue reflectivities as well as within the optic nerve head.

We present a technique for estimating the density of the human macular pigment noninvasively that takes advantage of the autofluorescence of lipofuscin, which is normally present in the human retinal pigment epithelium. By measuring the intensity of fluorescence at 710 nm, where macular pigment has essentially zero absorption, and stimulating the fluorescence with two wavelengths, one well absorbed by macular pigment and the other minimally absorbed by macular pigment, we can make accurate single-pass measurements of the macular pigment density. We used the technique to measure macular pigment density in a group of 159 subjects with normal retinal status ranging in age between 15 and 80 years. Average macular pigment density was 0.48 ± 0.16 density unit (D.U.) for a 2°-diameter test field. We show that these estimates are highly correlated with reflectometric (mean: $0.23\pm 0.07 \mathrm{D}.\mathrm{U}.)$ and psychophysical (mean: $0.37\pm 0.26 \mathrm{D}.\mathrm{U}.;$ obtained by heterochromatic flicker photometry) estimates of macular pigment in the same subjects, despite the fact that systematic differences in the estimated density exist between techniques. Repeat measurements over both short- and long-time intervals indicate that the autofluorescence technique is reproducible: The mean absolute difference between estimates was less than 0.05 D.U., superior to the reproducibility obtained by reflectometry and flicker photometry. To understand the systematic differences between density estimates obtained from the different methods, we analyzed the underlying assumptions of each technique. Specifically, we looked at the effect of self-screening by visual pigment, the effect of changes in optical property of the deeper retinal layers, including the role of retinal pigmented epithelium melanin, and the role of secondary fluorophores and reflectors in the anterior layers of the retina.