Adaptive optics imaging of microscopic structures in rat retina in vivo

Abstract

Purpose.

Rodent transgenic and knockout models enhance our understanding of normal and diseased retina. However, in vivo microscopic resolution of retinal cells is compromised by ocular aberrations. We have characterized the performance of the Rochester fluorescence adaptive optics scanning laser ophthalmoscope (fAOSLO) for imaging cells in the living rat retina.

Methods

GFP was expressed in ganglion cells of normal SD rats via intravitreal injections of AAV vectors. An fAOSLO acquired simultaneous reflectance and fluorescence retinal images. For comparison, histological images were obtained in the same eyes using a confocal SLO. In vivo resolution with the fAOSLO was characterized by comparing the transverse cross-section across individual dendrites from in vivo images to cross-sections from histology.

Results

Capillaries and fluorescently-labeled ganglion cell bodies, axons, and dendrites were clearly resolved after AO correction. The full width at half maximum (FWHM) of the in vivo line spread function was estimated by using deconvolution to correct the in vivo light distribution corresponding to a single dendrite for the dendrite diameter measured from histology. The line spread function FWHM was about 2 µm, which is twice as wide as the FWHM of the diffraction-limited line spread function.

Conclusions

If the aberrations of the rat eye were completely corrected, the resolution could be ∼2X that of the human eye. While our instrument corrects a substantial fraction of the aberrations as indicated by the RMS wavefront error after AO correction, direct measurements of retinal image quality reveal some blur beyond that expected from diffraction. Nonetheless, subcellular features of ganglion cells can be resolved, which offers promise for using adaptive optics to investigate the rodent eye in vivo.

Footnotes

 Supported by grants from NIH (EY014375 and EY01319) and Research to Prevent Blindness. This work has also been supported in part by the NSF Science and Technology Center for Adaptive Optics, managed by the University of California at Santa Cruz under cooperative agreement No. AST-9876783. We thank Mina Chung, Lana Nagy, Terry Schaefer, & Joe Stamm for their assistance with this work.