Imaging light responses of retinal ganglion cells in the living mouse eye

Abstract

This study reports development of a novel method for high-resolution in vivo imaging of the function of individual mouse retinal ganglion cells (RGCs) that overcomes many limitations of available methods for recording RGC physiology. The technique combines insertion of a genetically encoded calcium indicator into RGCs with imaging of calcium responses over many days with FACILE (functional adaptive optics cellular imaging in the living eye). FACILE extends the most common method for RGC physiology, in vitro physiology, by allowing repeated imaging of the function of each cell over many sessions and by avoiding damage to the retina during removal from the eye. This makes it possible to track changes in the response of individual cells during morphological development or degeneration. FACILE also overcomes limitations of existing in vivo imaging methods, providing fine spatial and temporal detail, structure-function comparison, and simultaneous analysis of multiple cells.retinal ganglion cells; in vivo adaptive optics imaging; calcium imaging AVAILABLE METHODS FOR EXAMINING the physiology of retinal ganglion cells (RGCs) have marked limitations, which are resolved by the novel in vivo imaging method described here.

In vivo two-photon imaging of the mouse retina

“…Images were acquired at a frame rate of 22 Hz, slower than previous experiments with mouse imaging [4,15]. Due to the breathing motion of the anesthetized animal, there was noticeable motion in the image collected from one frame to the next.…”

“…A separate group of mice received retrograde injections of rabies viral vector, carrying GCaMP3 and DsRed genes (produced in the laboratory of Dr. Edward M. Callaway, Salk Institute for Biological Study, San Diego, CA, USA). Injections were made into the superior colliculus, a retino-recipient nucleus [15,19] labeling ganglion cells through retrograde transport. For in vivo imaging, mice were anesthetized with Ketamine/Xylazine cocktail injections.…”

“…Functional activation of ganglion cells labeled with a genetically encoded calcium indicator, G-CaMP3, has previously been recorded with an AOSLO in mouse eyes by Yin et al in vivo using conventional single-photon excitation methods [15]. We have applied two-photon imaging to visualize retinal ganglion cells labeled with green fluorescent protein (GFP) and G-CaMP3 and ascertained the feasibility and prospective accessibility of cell activation studies in the neural circuitry in the living mouse eye.…”

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

“…However, reports of in vivo imaging of the mouse retina with AO-enhanced SLO and OCT have only recently been published [2][3][4][5][6][7][8][9]. AO imaging of the mouse retina has been delayed by the challenge of designing a system for an eye ten-fold smaller than that of the human, and by the availability of highly developed ex vivo histochemical retinal imaging methods.…”

Imaging Light Responses of Foveal Ganglion Cells in the Living Macaque Eye

“…Measures of the luminance increment response of individual RGCs showed a graded light response of many cells over a 2 log 10 unit range of full-field intensity, and many cells showed saturated responses at the highest measured luminance. The study presented here extended the FACILE method first developed in the mouse eye by Yin et al (2013), to macaque fovea.…”