Abstract
Two-photon ophthalmoscopy has potential for in vivo assessment of function of normal and diseased retina. However, light safety of the sub-100 fs laser typically used is a major concern and safety standards are not well established. To test the feasibility of safe in vivo two-photon excitation fluorescence (TPEF) imaging of photoreceptors in humans, we examined the effects of ultrashort pulsed light and the required light levels with a variety of clinical and high resolution imaging methods in macaques. The only measure that revealed a significant effect due to exposure to pulsed light within existing safety standards was infrared autofluorescence (IRAF) intensity. No other structural or functional alterations were detected by other imaging techniques for any of the exposures. Photoreceptors and retinal pigment epithelium appeared normal in adaptive optics images. No effect of repeated exposures on TPEF time course was detected, suggesting that visual cycle function was maintained. If IRAF reduction is hazardous, it is the only hurdle to applying two-photon retinal imaging in humans. To date, no harmful effects of IRAF reduction have been detected.
Highlights
In many blinding retinal diseases, severe function loss precedes any detectable structural changes in the retina, impeding a timely diagnosis
All exposures caused a significant reduction in infrared autofluorescence (IRAF) (p
Areas of reduced IRAF were larger than the marked location
Summary
In many blinding retinal diseases, severe function loss precedes any detectable structural changes in the retina, impeding a timely diagnosis. Photoreceptor dystrophies [1] initially compromise either rod or cone function [2,3], whereas glaucoma manifests in diminished ganglion cell responses [4]. Objective methods currently used to assess outer retinal function include electroretinography [5,6] and rhodopsin densitometry [7]. In both techniques, several retinal layers contribute to the signal, generating artifacts and complicating data interpretation [8,9,10]. To facilitate the diagnosis and prognosis of retinal diseases as well as the assessment of therapeutic interventions, more reliable, noninvasive methods are needed to evaluate retinal function
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