OPTOGENETIC SILENT SUBSTITUTION: 50 SHADES OF COLORS TO DECOUPLE OPTOGENETIC AND PHOTORECEPTOR ACTIVATION IN THE RETINA

Optogenetics enables precise control of neural activity, but its application in the retina is limited by the intrinsic photosensitivity of photoreceptors. Light used to activate optogenetic tools overlaps with the absorption spectra of endogenous opsins, leading to unavoidable photoreceptor activation. Here, we introduce Optogenetic Silent Substitution (OSS), inspired by the silent substitution used in visual psychophysics. We switched between two spectral configurations, one including a red stimulation capable of activating a red-shifted transfected opsin (such as ReachR), and another without red illumination.

We aim to maintain a constant photoreceptor isomerization rate between configurations, while only the red configuration activates the optogenetic protein. We achieved this by adjusting green and yellow LED intensities to compensate for photoreceptor activation induced by the red LED. To find the proper light intensity levels, we used multi-electrode arrays to measure responses of wild-type retinas, that did not express optogenetic protein, to various spectral configurations. Our goal was to identify pairs of configurations, one including red illumination and one without, that evoked equivalent activation of photoreceptors, such that alternating between them did not change the ganglion cell response.

Preliminary results suggest that there is a range of LED intensity values that achieves an effective “silent substitution”: they are indistinguishable by photoreceptors, but one configuration has a strong red light to activate optogenetic proteins. Our findings demonstrate that OSS is a viable technique to decouple optogenetic and photoreceptor activation. This new tool should allow to use optogenetics in the retina without photoreceptor crosstalk nor blocking photoreceptors.