Fig. 1
Spatial confinement influences migration decision-making, cellular ATP:ADP ratio, and glucose uptake. a Confocal reflectance image of a tapered collagen microtrack, and track width when cells contact two side walls of the microtrack or turn around and reverse migration direction (n = 20 cells). b Confocal reflectance image of a Y-shaped collagen microtrack and time-series images of decision-making during migration in Y-shaped microtracks (cell body outlined in yellow). c Final migration choice of cells based on cell-matrix contact before reaching the bifurcation (n = 34, 29, 41 cells for one wall (7 μm), one wall (12 μm), or two walls). d Time taken for cells touching two walls to pass through the bifurcation and enter a branch (n = 29 cells). e Confocal reflectance images of microtrack structure, and normalized PercevalHR ratio and 2-NBDG heatmaps of cells in each microtrack (yellow lines show microtrack walls; red arrowheads show areas of matrix displacement around the cell body). f, g Quantification of cell body aspect ratio (f) and track displacement (g) for cells in 15, 12, and 7 μm microtracks (n = 95, 96, and 91 cells, respectively). h, i Normalized PercevalHR ratio (n = 95, 96, and 91 cells, respectively) (h) and 2-NBDG uptake (n = 54, 40, and 56 cells, respectively) (i) for cells in 15, 12, and 7 μm microtracks. j Intracellular ATP:ADP ratio (normalized PercevalHR ratio) during decision-making in Y-shaped microtracks (yellow lines show microtrack walls). k Quantification of normalized PercevalHR ratio as a function of distance from the bifurcation and after final migration choice (n = 10 cells for 12 μm path, 5 cells for 7 μm path). l Normalized PercevalHR ratio in tapered collagen microtracks (n = 62 cells). Data shown as median ± interquartile range (box), 5th–95th percentiles (whiskers), and mean (+) (a, d, f, g, k), or mean ± s.e.m. (c, h, i); dashed lines show exponential growth; Clopper–Pearson confidence interval for observed proportion (c), two-tailed Mann–Whitney test (d, f, g, k), or extra sum-of-squares F-test (h, i, l); *P < 0.05, **P < 0.01, ***P < 0.001. Scale bar, 50 μm (a, b), 25 μm (b, j), and 15 μm (e)
