Fig. 3

Optically enhanced formation of protrusions on regenerating axons after axotomy. a Schematic diagram of the axotomy experimental scheme. Neurons were loaded in the left compartment (cell body side) and axons grew to the right compartment (axonal side) through the microgroove barrier. On DIV4, neurons were transduced with AAVs expressing mScarlet and optoRET. On DIV11, neurons underwent axotomy and axons were allowed to regenerate for two consecutive days, during which the axonal side of the chip was illuminated with a blue LED plate. On DIV13, the axonal side of the microfluidics chip was imaged. b Representative confocal images of the axonal sides of microfluidics chips on DIV13. Scale bars = 200 µm. c Comparison of regenerated axon length (mm) between Dark (n = 64) and Light (n = 70) groups. Data are presented as means ± SEM (*p < 0.01; Kolmogorov–Smirnov test). d Representative skeletal images of regenerated axons. e Heatmap of the frequency distribution of low, intermediate, and high classes of axon morphological complexity. f Comparison of protrusion densities (protrusion count/µm) between Dark (n = 61) and Light (n = 69) groups. Data are presented as means ± SEM (****p < 0.0001; Kolmogorov–Smirnov test)