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Biomed Opt Express. 2015 May 21;6(6):2191-210. doi: 10.1364/BOE.6.002191. eCollection 2015 Jun 1.

Adaptive-optics SLO imaging combined with widefield OCT and SLO enables precise 3D localization of fluorescent cells in the mouse retina.

Author information

1
UC Davis RISE Eye-Pod Laboratory, Dept. of Cell Biology and Human Anatomy, University of California Davis, 4320 Tupper Hall, Davis, California 95616, USA ; Vision Science and Advanced Retinal Imaging Laboratory (VSRI) and Department of Ophthalmology & Vision Science, UC Davis, 4860 Y Street, Ste. 2400, Sacramento, CA 95817, USA ; Depts. of Ophthalmology & Vision Science and of Cell Biology & Human Anatomy 4303 Tupper Hall, Davis California 95616, USA ; rjzawadzki@ucdavis.edu.
2
UC Davis RISE Eye-Pod Laboratory, Dept. of Cell Biology and Human Anatomy, University of California Davis, 4320 Tupper Hall, Davis, California 95616, USA.
3
Center for Neuroscience, University of California, Davis Sacramento, CA 95817, USA.
4
Vision Science and Advanced Retinal Imaging Laboratory (VSRI) and Department of Ophthalmology & Vision Science, UC Davis, 4860 Y Street, Ste. 2400, Sacramento, CA 95817, USA.
5
Beckman Laser Institute Korea & Biomed. Engineering, Dankook University, Cheonan, Chungnam 330-715, South Korea.
6
Dept. of Molecular and Cellular Biology, University of California, Berkeley 94720, USA.
7
Center for Neuroscience, University of California, Davis Sacramento, CA 95817, USA ; Depts. of Ophthalmology & Vision Science and of Cell Biology & Human Anatomy 4303 Tupper Hall, Davis California 95616, USA.
8
UC Davis RISE Eye-Pod Laboratory, Dept. of Cell Biology and Human Anatomy, University of California Davis, 4320 Tupper Hall, Davis, California 95616, USA ; Depts. of Cell Biology & Human Anatomy, and of Physiology & Membrane Biology 4303 Tupper Hall, Davis California 95616, USA ; enpugh@ucdavis.edu.

Abstract

Adaptive optics scanning laser ophthalmoscopy (AO-SLO) has recently been used to achieve exquisite subcellular resolution imaging of the mouse retina. Wavefront sensing-based AO typically restricts the field of view to a few degrees of visual angle. As a consequence the relationship between AO-SLO data and larger scale retinal structures and cellular patterns can be difficult to assess. The retinal vasculature affords a large-scale 3D map on which cells and structures can be located during in vivo imaging. Phase-variance OCT (pv-OCT) can efficiently image the vasculature with near-infrared light in a label-free manner, allowing 3D vascular reconstruction with high precision. We combined widefield pv-OCT and SLO imaging with AO-SLO reflection and fluorescence imaging to localize two types of fluorescent cells within the retinal layers: GFP-expressing microglia, the resident macrophages of the retina, and GFP-expressing cone photoreceptor cells. We describe in detail a reflective afocal AO-SLO retinal imaging system designed for high resolution retinal imaging in mice. The optical performance of this instrument is compared to other state-of-the-art AO-based mouse retinal imaging systems. The spatial and temporal resolution of the new AO instrumentation was characterized with angiography of retinal capillaries, including blood-flow velocity analysis. Depth-resolved AO-SLO fluorescent images of microglia and cone photoreceptors are visualized in parallel with 469 nm and 663 nm reflectance images of the microvasculature and other structures. Additional applications of the new instrumentation are discussed.

KEYWORDS:

(110.1080) Active or adaptive optics; (110.4500) Optical coherence tomography; (170.0110) Imaging systems; (170.4460) Ophthalmic optics and devices; (170.4470) Ophthalmology; (330.7324) Visual optics, comparative animal models

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