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Robin Sharma, Lu Yin, Ying Geng, William H. Merigan, David R. Williams, Jennifer J. Hunter; In vivo two-photon imaging of the mouse retina. Journal of Vision 2012;12(14):51. doi: https://doi.org/10.1167/12.14.51.
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Two-photon imaging in the living eye has applications such as imaging compounds in structures that are otherwise transparent and measuring cellular function at wavelengths that minimally excite the visual response. Imaging of two-photon autofluorescence from photoreceptors has been demonstrated previously in the primate retina (Hunter et al. 2011). Our goal is to develop two-photon imaging of retinal cells in the living mouse eye because of (1) the substantially higher spatial resolution and light collecting efficiency of the mouse eye than that of humans (Geng et al. 2012), (2) the availability of mouse models of retinal disease and, (3) the facility with which mouse retinal cells can be fluorescently labeled for both structural and functional imaging. To utilize these advantages, we have recently built a two-photon adaptive optics scanning laser ophthalmoscope for imaging the mouse retina. Using this instrument, we were able to collect two-photon autofluorescence from the photoreceptor layer using 730 nm excitation light and also observed functional changes in fluorescence signal over time. Also, for the same incident power signal levels in the mouse were nearly 60 times greater than the primate. Extrinsic fluorophores can also be used to generate signal contrast in two photon imaging. We have successfully imaged ganglion cell somas and sometimes their dendrites labeled with green fluorescent protein (GFP) and a GFP based calcium indicator G-CaMP3.
Meeting abstract presented at OSA Fall Vision 2012
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