3D imaging of cone photoreceptors over extended time periods using optical coherence tomography with adaptive optics

Optical coherence tomography with adaptive optics (AO-OCT) is a highly sensitive, noninvasive method for 3D imaging of the microscopic retina. The purpose of this study is to advance AO-OCT technology by enabling repeated imaging of cone photoreceptors over extended periods of time (days). This sort of longitudinal imaging permits monitoring of 3D cone dynamics in both normal and diseased eyes, in particular the physiological processes of disc renewal and phagocytosis, which are disrupted by retinal diseases such as age related macular degeneration and retinitis pigmentosa. For this study, the existing AO-OCT system at Indiana underwent several major hardware and software improvements to optimize system performance for 4D cone imaging. First, ultrahigh speed imaging was realized using a Basler Sprint camera. Second, a light source with adjustable spectrum was realized by integration of an Integral laser (Femto Lasers, lambda(c)=800nm, Delta lambda=160nm) and spectral filters in the source arm. For cone imaging, we used a bandpass filter with lambda(c)=809nm and Delta lambda=81nm (2.6 mu m nominal axial resolution in tissue, and 167 KHz A-line rate using 1,408 px), which reduced the impact of eye motion compared to previous AO-OCT implementations. Third, eye motion artifacts were further reduced by custom ImageJ plugins that registered (axially and laterally) the volume videos. In two subjects, cone photoreceptors were imaged and tracked over a ten day period and their reflectance and outer segment (OS) lengths measured. High-speed imaging and image registration/dewarping were found to reduce eye motion to a fraction of a cone width (1 mu m root mean square). The pattern of reflections in the cones was found to change dramatically and occurred on a spatial scale well below the resolution of clinical instruments. Normalized reflectance of connecting cilia (CC) and OS posterior tip (PT) of an exemplary cone was 54 +/- 4, 47 +/- 4, 48 +/- 6, 50 +/- 5, 56 +/- 1% and 46 +/- 4, 53 +/- 4, 52 +/- 6, 50 +/- 5, 44 +/- 1% for days #1,3,6,8,10 respectively. OS length of the same cone was 28.9, 26.4, 26.4, 30.6, and 28.1 mu m for days # 1,3,6,8,10 respectively. It is plausible these changes are an optical correlate of the natural process of OS renewal and shedding.