Optogenetic modulation of excitability using ChR2-NaV1.5 and automated patch clamp platforms for translational safety screening

Abstract

Channelrhodopsins (ChRs) are light-gated proteins that control phototaxis in protists and are commonly used in optogenetics to manipulate neuron activity. ChR2-Na_V1.5, a fusion of ChR2 with cardiac-specific Na⁺ channel, modulates sodium channel kinetics and produces photosensitive inward currents.

Using automated low and high throughput patch clamp (APC), we found that blue-light intensity directly correlates with ChR2 channel activity, with higher intensities resulting in larger peak amplitudes under constant light exposure durations. We also found that light could be used to activate ChR2, depolarizing the membrane sufficiently in current clamp mode to elicit action potentials.

This is demonstrated on both the Port-a-Patch, a semi-automated patch clamp device recording from a single cell at a time, and the high throughput fully automated device, the SyncroPatch 384. The SyncroPatch 384 could also be used to investigate different ChR2 variants. Several different constructs could be activated using blue light on a single NPC-384 chip. In this study we used HEK cells stably expressed ChR2-Na_V1.5 but the technology could be extended to hiPSC-neurons or other stem cells or primary cells which express light activated ion channels.

Our findings underscore the utility of APC combined with optogenetics in enhancing assay throughput, consistency, and mechanistic resolution. These advanced platforms could support early-stage safety screening and contribute to more predictive, human-relevant models for drug-induced excitability disorders and off-target toxicities.