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Ying Geng, Robin Sharma, Alfredo Dubra, Kamran Ahmad, Ted Twietmeyer, Benjamin Masella, Jennifer J. Hunter, Richard T. Libby, David R. Williams; High Resolution In Vivo Imaging Of The Mouse Retina Using An Adaptive Optics Scanning Laser Ophthalmoscope. Invest. Ophthalmol. Vis. Sci. 2011;52(14):5871. doi: https://doi.org/.
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Non-invasive microscopic imaging of the mouse retina would allow tracking of retinal development, disease progression, and the efficacy of therapy in single animals. Correction of the eye’s aberrations using adaptive optics (AO) could improve the resolution of in vivo mouse retinal images, but previous attempts have been limited by the small size of the mouse eye and the difficulty in measuring its aberrations due to a relatively optically thick retina which results in poor Shack-Hartmann (SH) wavefront spot quality. To improve retinal imaging in the mouse, we have developed a SH wavefront sensor tailored for the mouse eye that uses back-scattered light from the retina, incorporated it in a fluorescence adaptive optics scanning laser ophthalmoscope (FAOSLO), and tested its performance on the living mouse eye.
Adult transgenic mice that express YFP in a small subset of retinal ganglion cells (B6. Thy1-YFPH) were imaged. Mice were anesthetized and a contact lens was used to maintain corneal hydration. A SH wavefront sensor, operating at 843 nm, with a large diameter annular entrance pupil beacon was used to decrease the depth of focus and block the corneal reflection. Wave aberration was corrected with a 97-actuator large-stroke deformable mirror operating at 15 Hz. Simultaneous reflectance images and fluorescence images were acquired in vivo through a 2 mm dilated pupil.
Our wavefront sensor design allowed us to use the back-scattered light for closing the adaptive optics control loop in the mouse eye. Cellular structure can be resolved in reflectance imaging. Ganglion cell bodies, dendrites and axons were clearly resolved in registered in vivo fluorescent images. Both reflectance and fluorescence retinal images obtained with adaptive optics have improved resolution and contrast over images obtained without adaptive optics.
An AO instrument constructed with a wavefront sensor based on back-scattered light provides a fast and effective correction of mouse eye aberrations. The large numerical aperture of the mouse eye coupled with AO imaging potentially offers in vivo resolution at least two times larger than that of a diffraction-limited human eye, while collecting four times more light than that of the human eye. Thus, the FAOSLO could be a valuable tool for nonlinear imaging techniques and other light-starved imaging modalities.
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