Cationic Channelrhodopsin from the Alga Platymonas subcordiformis as a Promising Optogenetic Tool

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Olga S. Idzhilova^1,2, Gulnur R. Smirnova^1,2, Lada E. Petrovskaya³, Darya A. Kolotova^1,2, Mikhail A. Ostrovsky², and Alexey Y. Malyshev^1,a*

¹Institute of Higher Nervous Activity and Neurophysiology, Russian Academy of Sciences, 117485 Moscow, Russia

²Emanuel Institute of Biochemical Physics, Russian Academy of Sciences, 119334 Moscow, Russia

³Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia

^* To whom correspondence should be addressed.

Received September 27, 2022; Revised October 10, 2022; Accepted October 10, 2022

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The progress in optogenetics largely depends on the development of light-activated proteins as new molecular tools. Using cultured hippocampal neurons, we compared the properties of two light-activated cation channels – classical channelrhodopsin-2 from Chlamydomonas reinhardtii (CrChR2) and recently described channelrhodopsin isolated from the alga Platymonas subcordiformis (PsChR2). PsChR2 ensured generation of action potentials by neurons when activated by the pulsed light stimulation with the frequencies up to 40-50 Hz, while the upper limit for CrChR2 was 20-30 Hz. An important advantage of PsChR2 compared to classical channelrhodopsin CrChR2 is the blue shift of its excitation spectrum, which opens the possibility for its application in all-optical electrophysiology experiments that require the separation of the maxima of the spectra of channelrhodopsins used for the stimulation of neurons and the maxima of the excitation spectra of various red fluorescent probes. We compared the response (generation of action potentials) of neurons expressing CrChR2 and PsChR2 to light stimuli at 530 and 550 nm commonly used for the excitation of red fluorescent probes. The 530-nm light was significantly (3.7 times) less efficient in the activation of neurons expressing PsChR2 vs. CrChR2-expressing neurons. The light at 550 nm, even at the maximal used intensity, failed to stimulate neurons expressing either of the studied opsins. This indicates that the PsChR2 channelrhodopsin from the alga P. subcordiformis is a promising optogenetic tool, both in terms of its frequency characteristics and possibility of its application for neuronal stimulation with a short-wavelength (blue, 470 nm) light accompanied by simultaneous recording of various physiological processes using fluorescent probes.

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KEY WORDS: optogenetics, neuron, bacterial opsin, channelrhodopsin, light-induced current, patch clamp, intracellular recording, Platymonas subcordiformis

DOI: 10.1134/S0006297922110116

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