Journal: Nature Communications

Article Title: Triggering action potentials of a single neuron by multiphoton excitation elicits visually guided behavior

doi: 10.1038/s41467-026-69446-5

Figure Lengend Snippet: a The Ca 2+ amplitude of neuronal activities depends on the photostimulation powers in vitro ( n = 163 neurons). b The Ca 2+ level of photostimulated neurons in the presence of Tempo (400 μM; n = 20 neurons for control and n = 19 neurons for Tempo; Two-tailed Mann-Whitney test). c The Propidium Iodide (PI, 20 μM) and Fluo-4 fluorescence in neurons suffered photostimulation and Triton X-100 (0.1%; positive control), respectively. Right: Compared the fluorescence intensity of PI in neurons suffered photostimulation and Triton X-100 ( p = 4.0 × 10 −21 ; n = 18 neurons of photostimulation and n = 26 neurons of Triton X-100; Two-tailed Mann-Whitney test). Scale bar: 10 μm. d The Ca 2+ amplitude of neuronal activities depends on the photostimulation powers in vivo ( n = 25 neurons from 4 mice). e The activation efficiency of photostimulation in vivo ( n = 24 neurons from 4 mice). f The in vivo PI (Propidium Iodide) assay of photostimulation. The GCaMP6S (black line) and PI (50 μM, 800 nl, red line) fluorescence in neurons suffered photostimulation ( n = 11 neurons). Scale bar: 20 μm. g The neuronal responses to stimulation by a CW laser in vivo. Left: Ca 2+ response of the same neuron after CW laser stimulation and femtosecond laser stimulation. Both lasers were 900 nm. Right: Quantified Ca 2+ response of neurons after the CW and femtosecond laser stimulation at the same wavelength ( p = 0.0003; n = 6 neurons; Two-tailed Mann-Whitney test). Scale bar: 20 μm. h The neuronal responses to photostimulation at different locations inside or outside the neuron in vivo ( p = 0.0312; n = 6 neurons; Two-tailed Wilcoxon matched-pairs signed rank test). Scale bar: 10 μm. i The neuronal responses to the photostimulation by the same femtosecond laser at different repetition rates (1 MHz vs. 10 MHz) in vivo. The depth was 90 – 120 μm. n = 13 neurons for 1 MHz, 24 neurons for 10 MHz. Scale bar: 20 μm. j Photostimulation applied to a specific neuron (upper panel) and a single axon structure (lower panel) led to a Ca 2+ response in that neuron (right plots). n = 122 neurons from 11 mice. Scale bar: 20 μm. k The projection presentation (over time) of the fluorescence integral of neurons for 40 s since the photostimulation to the target (Neuron 1). Scale bar: 20 μm. Inserts: Ca 2+ responses from the neighbors (Neuron 2–5). Scale bar: 10 μm. Upper-right: heat map of ΔF/F 0 . Lower-right: ΔF/F 0 of neuron 1–5. n = 13 neurons. l The graph depicts how neighboring neurons respond relative to their distance from the photostimulation site. 0 μm: the neuron that was photostimulated. The red line represents the average background ΔF/F 0 for all non-stimulated neurons during periods of non-stimulation. p = 2.0 × 10 −9 ; n = 13 photostimulated neurons from 4 mice; Two-tailed Mann-Whitney test. m The responses of neurons in other planes at the z direction. The data acquired by performing multi-plane imaging. Right: light-blue line from individual neurons and dark-blue the average. Scale bar: 20 μm. n = 6 neurons from 3 mice. Error bars for the averaged plot indicate mean ± s.e.m. Source data are provided as a Source Data file.

Article Snippet: Triton X-100 (HY-Y1883A, MedChemExpress) was added as a positive control.

Techniques: In Vitro, Control, Two Tailed Test, MANN-WHITNEY, Fluorescence, Positive Control, In Vivo, Activation Assay, Imaging