Fig. 3: Ion selectivity and physiological features of VirChR1.

a Schematic comparison of VirChR1 and CrChR2 ion channeling activity under different calcium concentrations, membrane boundaries are shown schematically as black horizontal lines. b Voltage-clamp records from n = 1 representative SH-SY5Y cell, expressing VirChR1 with (left) 10 mM HEPES pH 7.4, 110 mM NaCl, 2 mM MgCl₂ and (right) 110 mM L-arginine hydrochloride replacing NaCl in bath. Pipette solution during experiments was: 10 mM HEPES pH 7.4, 110 mM NaCl, 2 mM MgCl₂, 10 mM EGTA, illumination by LED (470 nm) lamp is indicated with light blue color. c Current–voltage dependences for n = 1 representative SH-SY5Y cell in 110 mM NaCl (red) and 110 mM L-arginine hydrochloride (blue). Currents are reproducible and typical to those in n = 9 experiments with other cells (and n = 21 experiments under slightly different NaCl concentrations varied from 110 mM to 140 mM). d Action spectrum of VirChR1 measured using equal photon fluxes (Sample size, n = 18–20). e Voltage-clamp records from n = 1 representative SH-SY5Y cell expressing VirChR1 in bath solution (left) 10 mM HEPES pH 7.4, 110 mM NaCl, 2 mM MgCl₂ and (right) in 80 mM CaCl₂ replacing NaCl in bath solution. f Current–voltage dependences for n = 1 representative SH-SY5Y cell in 110 mM NaCl (red) and 80 mM CaCl₂ (indigo) solutions. g Excitation recovery of photocurrent after a short pulse of nanosecond laser (500 nm) activation. Tau-off was measured in n = 5 independent experiments. Current–voltage dependences for n = 1 cell for different bath/pipette solution. Traces are shown for h bath solutions: 110 mM NaCl (red) and 110 mM KCl (green) (pipette solution is standard) and i pipette solution 110 mM L-arginine hydrochloride salt solution of pH 5.0 (bath solution is standard). Estimation of relative conductivities for different ions was done by fitting traces with Goldman-equation. l Current dependence on calcium concentration in bath solution measured at +80 mV (inflection point is at ~2.2 mM of calcium). For all electrophysiological recordings at n = 1 cell currents were reproducible in n = 3–10 independent experiments with other cells. No current averaging between cells was done, since different cells have significantly different protein expression levels. Data are presented as mean values ± SEM of current value under illumination in the cell measured.