To understand visual function and cellular metabolism in the living eye, we need a means to visualize individual cells and assess molecular dynamics non-invasively. Some of the key molecules involved in the visual cycle and cellular energy production are intrinsically fluorescent, but are inaccessible in the living eye through single-photon excitation because the needed wavelength range is not transmitted by the optics of the eye. Two-photon excited fluorescence ophthalmoscopy (TPEFO) can excite these otherwise inaccessible fluorophores with near-infrared light. In our previous experiments in the living macaque eye, we used an adaptive optics scanning light ophthalmoscope (AOSLO) to non-invasively image many different cell classes throughout the retina, including ganglion cells (Sharma, Williams et al., 2016). Additionally, we measured visual cycle function by characterizing the time course of two-photon excited fluorescence from photoreceptors (Sharma, Schwarz et al., 2016). We now propose to expand the capabilities of TPEFO in the living rodent and macaque eye to measure several important fluorescence properties, such as emission spectrum, fluorescence lifetime, and redox ratio, which may be indicators of cell health and function. This research has the potential to provide insight into normal and altered biochemical processes and improve our understanding of diseases that impact retinal metabolism and function such as glaucoma, macular degeneration and Leber hereditary optic neuropathy.
We will develop two-photon excited fluorescence ophthalmoscopy to non-invasively assess cellular metabolism and function in the living eye. We aim to investigate important properties of the excited fluorophores in response to visual stimulation, pharmacological manipulation or injury at a single cell level. This research may lead to advances in the diagnosis of retinal diseases, monitoring of disease progression, and assist in evaluating treatment efficacy.
| Schwarz, Christina; Sharma, Robin; Cheong, Soon Keen et al. (2018) Selective S Cone Damage and Retinal Remodeling Following Intense Ultrashort Pulse Laser Exposures in the Near-Infrared. Invest Ophthalmol Vis Sci 59:5973-5984 |
| Sharma, Robin; Schwarz, Christina; Hunter, Jennifer J et al. (2017) Formation and Clearance of All-Trans-Retinol in Rods Investigated in the Living Primate Eye With Two-Photon Ophthalmoscopy. Invest Ophthalmol Vis Sci 58:604-613 |
| Marcos, Susana; Werner, John S; Burns, Stephen A et al. (2017) Vision science and adaptive optics, the state of the field. Vision Res 132:3-33 |
| Rossi, Ethan A; Granger, Charles E; Sharma, Robin et al. (2017) Imaging individual neurons in the retinal ganglion cell layer of the living eye. Proc Natl Acad Sci U S A 114:586-591 |
| Sharma, Robin; Schwarz, Christina; Williams, David R et al. (2016) In Vivo Two-Photon Fluorescence Kinetics of Primate Rods and Cones. Invest Ophthalmol Vis Sci 57:647-57 |
| Schwarz, Christina; Sharma, Robin; Fischer, William S et al. (2016) Safety assessment in macaques of light exposures for functional two-photon ophthalmoscopy in humans. Biomed Opt Express 7:5148-5169 |
| Sharma, Robin; Williams, David R; Palczewska, Grazyna et al. (2016) Two-Photon Autofluorescence Imaging Reveals Cellular Structures Throughout the Retina of the Living Primate Eye. Invest Ophthalmol Vis Sci 57:632-46 |
| Masella, Benjamin D; Hunter, Jennifer J; Williams, David R (2014) New wrinkles in retinal densitometry. Invest Ophthalmol Vis Sci 55:7525-34 |
| Palczewska, Grazyna; Golczak, Marcin; Williams, David R et al. (2014) Endogenous fluorophores enable two-photon imaging of the primate eye. Invest Ophthalmol Vis Sci 55:4438-47 |
| Masella, Benjamin D; Hunter, Jennifer J; Williams, David R (2014) Rod photopigment kinetics after photodisruption of the retinal pigment epithelium. Invest Ophthalmol Vis Sci 55:7535-44 |
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