Extended Data Fig. 3: CANE-captured CeA_GA neurons are mostly inhibited by stress.

a, Schematic of CANE captured isoflurane-activated CeA_GA neurons followed by a second exposure to restraint stress (left, for Fos expression, right, for calcium imaging). b, Left, representative images of CANE_ISO-tdTomato neurons (red) and stress-activated Fos⁺ neurons (green) and their overlap. (n = 3 animals). Right, representative images of isoflurane-only activated Fos⁺ neurons (green, repeated experiments n = 5 biologically independent samples.) and compared to stress-only activated Fos⁺ neurons (green, repeated experiments n = 3 biologically independent samples.) in the CeA. c, Heatmap, activity patterns of CANE_ISO-GCaMP6m captured CeA_GA neurons in stress experiment sorted by the average activity during stress period (-8–0 min, pre-stress; 0–8 min, restraint stress; 8 -16 min, post-stress. 282 neurons from 5 mice × 2 trials). A small number of neurons (in the bottom of the heatmap) were activated by stress. d, Example traces of CANE_ISO-GCaMP6m captured CeA_GA neurons in stress experiment showing both stress-inhibited and stress-activated neurons. Norm. intensity: normalized calcium signals rescaled to 0–1. e, Scatter plots of the tracked same neurons based on isoflurane and stress related activity patterns. Each dot is calculated based on effectiveness corrected activity ratio between the post- and pre-stimulus periods for isoflurane-stimulus and stress-stimulus, separately. Corrected ratio, mean activity of (post-) effective time / mean activity of (pre-) effective time. Single dots represent individual neurons in the logarithmic scale coordinates, and the circled dots represent robustly firing neurons, with the maximum intensity of each neuron in the whole duration exceeding a threshold. f, Same plots with isoflurane responses calculated by actively firing time exceeding a threshold (1 min). Active duration, total (post-) effective time of effective moment >1 min. “Effective time” refers to the time points of a neural trace whose intensity is two median absolute deviation above its mean. Only these time points were considered to compute the mean. g, Neuron count summary of e. Left, Neuron count distribution of activity of isoflurane-suppressed neurons during stress. Right, Neural count distribution of activity of isoflurane-activated neurons during stress. h, Neural count distribution of f. Left, marginal count distribution of activity of CANE_ISO-GCaMP6m captured CeA_GA neurons during stress. Right, marginal count distribution of activity of CANE_ISO-GCaMP6m captured CeA_GA neurons during isoflurane GA.