Abstract
Purpose :
The perceptual roles of most retinal ganglion cell (RGC) classes are controversial in part because we lack a paradigm for measuring the perceptual consequences of activating cells of only one class in the living primate eye. Recent success in classifying single RGCs with in vivo calcium imaging (McGregor 2018, PLoS One, 13), in vivo optogenetic activation of RGCs en masse (McGregor 2020, Nat. Commun. 11, 1703; Sahel 2021, Nat. Med. 27., 1223), and in vivo targeted excitation of single cones (Harmening 2014, J. Neurosci. 34, 5667) now bring such a paradigm within reach. As a first step, we demonstrate the optogenetic stimulation of single foveal RGCs in the living macaque.
Methods :
RGCs in an anesthetized female macaque were stimulated and imaged using adaptive optics scanning light ophthalmoscopy. Co-expression of the optogenetic actuator and calcium indicator was achieved via intravitreal injection of AAV2-CAG-ChrimsonR-tdTomato and AAV2-CAG-GCaMP6s. We selected single RGC somas from those closest to the foveal center because of their large separation from their nearest neighbors (>20 µm). On each trial, the targeted soma was exposed to 12.5 µm diameter flashes (640 nm, 900 µW) of 0.8 second duration. Four consecutive flashes were delivered, each separated by 15s. The GCaMP responses to the flash of both targeted and nearby cells were imaged with a 1° field (488 nm, 25 µW) and the ΔF/F for each soma’s response was averaged over the four flashes.
Results :
In two cells serving cones at the foveal center, it was possible to record calcium responses in the targeted cell on single trials, a result that was repeatable in the same cells in two sessions one week apart. In the first session, the targeted cell responses showed mean ΔF/F values of 0.314 and 0.177. Nearby cells showed little or no response with a mean of 0.003 (se = 0.008, n = 52, 95% CI [-0.013, 0.018]). In the second session, the same two targeted cells gave values of 0.291 and 0.189. Nearby cells gave a mean value of 0.015 (se = 0.007, n = 66, 95% CI [0.002, 0.028]).
Conclusions :
We can directly activate single RGCs in the living eye and measure the resulting calcium signal, the first step toward a method to measure the sensations produced by individual RGCs in the awake behaving monkey. Such a paradigm could also provide valuable information for RGC-based optogenetic or optoelectronic vision restoration therapies.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.