Fig. 4: Pupil-induced RGC activity is relayed to the visual cortex.

A Experimental setup. 2-photon imaging of GCaMP8f expressing LGN axon terminals in V1 (green). B Example field of view (representative of N = 3 animals). C Responses of LGN axons to full field visual stimulation of contralateral (purple) or ipsilateral (green) eye (n = 82 boutons, N = 3 animals). D Response quality index of responses during stimulation. Contralateral stimulation (left, purple): mean = 0.24, above response threshold (0.3): 18.3%. Ipsilateral stimulation (right, green): mean=0.31, above response threshold (0.3): 56.1%. (n = 82 boutons, N = 3 mice). Boxplot values, from left to right: Mean (0.24, 0.31), lower box (0.19, 0.23), higher box (0.29, 0.39), lower whisker (0.12, 0.13), higher whisker (0.42, 0.57), max (0.5, 0.57), min (0.12, 0.13). E Response dynamics of LGN responses. Left: Population response of boutons to contralateral (top, purple) and ipsilateral (bottom, green) stimulation. Right: Time of peak response. Right top, Purple: the difference between stimulus offset and maximal response to contralateral stimulation. Right bottom, Green: the difference between stimulus onset and maximal response to ipsilateral stimulation. Red: time shown in histogram. F Experimental setup of layer 2/3 neurons in V1 (green). G Example field of view (representative of N = 3 animals). H K-means clustering in PCA space of the average responses of V1 cells (n = 152 cells, N = 2 animals), 2 clusters: OFF (green) and ON (yellow)-responding. I, J top: examples of one OFF (green) and one ON (yellow) cell response. Black, vertical lines = beginning ipsilaterally-presented LED flash. Bottom, gray: trial-averaged pupil area traces (z-score). Solid trace = mean, Shaded areas = SEM. K Histogram of the delay onset, for all the cells, color-coded according to the functional cluster (OFF green, ON yellow). L Average responses (n = 152 cells, N = 2 animals). Black, vertical lines = flash; the black dashed line = mean pupil response ± SEM. Blue is an activity increase, teal decrease. M Population response (OFF in green, ON in yellow), plotted against pupil response (gray, same as in (L)), all mean ± SEM. N Histogram of the percentage of the trial correlated with the pupil, calculated for all the cells recorded (n = 250 cells, N = 2 animals).