Putti, E. | Search bioRxiv

bioRxiv2025

Lrrns define a visual circuit underlying brightness and contrast perception

Brightness and contrast are fundamental features of vision, crucial for object detection, environmental navigation, and feeding. Here, we identify a brightness-and contrast-processing circuit in the zebrafish visual system and uncover the role of Leucine-rich repeat neuronal (Lrrn) cell adhesion molecules (CAMs) in regulating its assembly. Deep-projecting retinal ganglion cells (RGCs) serve as the first synaptic relay to the brain requiring Lrrn2 and Lrrn3a for precise axonal targeting and connectivity within the optic tectum. Genetic targeting of these CAMs leads to circuit disorganization and impairments in contrast sensitivity, leading to deficits in visually guided behaviors. Additionally, ultrastructural circuit reconstruction and functional imaging analysis revealed their critical role in luminance processing. These studies define a fundamental visual processing pathway and establish Lrrn CAMs as essential molecular drivers of its assembly.

neuroscience↗

bioRxiv2024

Two-photon voltage imaging with rhodopsin-based sensors

The recent advances in sophisticated optical techniques, coupled with two-photon sensitive genetic voltage indicators (GEVIs), have enabled in-depth voltage imaging in vivo at single spike and single-cell resolution. To date, these results have been only achieved using ASAP-type sensors, as the complex photocycle of rhodopsin-based voltage indicators posed challenges for their two-photon use, restricting their application to one-photon approaches. In this work, we demonstrate that rhodopsin-based GEVIs (FRET-opsin) can be used under two-photon illumination when their peculiar light intensity dependence of kinetics and sensitivity are considered. We rationally engineer a fully genetically-encoded, rhodopsin-based voltage indicator with the brightest known fluorophore AaFP1, Jarvis, and demonstrate its utility under both one- and two-photon illumination. We also showed two-photon usability of the similar FRET-opsin sensor pAce. Our comparison of 2P scanless with fast 2P scanning illumination revealed that the latter approach is less suitable for this class of indicators and, on the contrary, both sensors responded well when scanless approaches were used. Furthermore, utilising Jarvis, we demonstrated high-fidelity, high-SNR action potential detection at kilohertz-imaging rates both in mouse hippocampal slices and in zebrafish larvae. To the best of our knowledge, this study represents the first report of a fully genetically-encoded rhodopsin-based voltage indicator for high contrast action potential detection under two-photon illumination in vitro and in vivo.

neuroscience↗