Extended Data Fig. 5: Dual-color imaging of 5-HT and DA dynamics in acute mouse brain slices with high spatial-temporal resolution.
From: Improved green and red GRAB sensors for monitoring spatiotemporal serotonin release in vivo
a, Schematic illustrates the mouse brain slice experiments. b–h, Electrical stimulation evoked 5-HT and DA release. b, Representative fluorescence and pseudocolor images of g5-HT3.0 (top) and rDA3m (bottom) at baseline and in response to the indicated electrical stimuli, in the presence of artificial cerebrospinal fluid (ACSF) or 100 μM RS. Similar results were observed for 4 slices. The white dashed circle (50 μm in diameter) indicates the ROI used for further analysis; the white line indicates the stimulating electrode location. Scale bar, 100 μm. c–d, Representative traces and summary data for changes in g5-HT3.0 (c) and rDA3m (d) fluorescence in response to the indicated stimuli in ACSF or RS. e, Example time-lapse pseudocolor images of g5-HT3.0 (top) and rDA3m (bottom) in response to indicated electrical stimuli. Similar results were observed for 4 slices. The dashed lines were used to analyze spatial and temporal dynamics; image averaged from three trials conducted in one slice. Scale bar, 100 μm. f, Example spatial dynamics of the fluorescence changes shown in (e). g, Summary of the full width at half maximum (FWHM) of activity-dependent 5-HT and DA signals measured in f at the indicated time points. Two-tailed paired t-tests, P = 0.7559, 0.1318, 0.741 and 0.9301 for 1 s, 2 s, 5 s and 10 s, respectively. h, Group summary of on and off kinetics for the 100-pulse evoked response of g5-HT3.0 and rDA3m. Two-tailed paired t-tests, P = 0.4308 and 0.1415 for on and off kinetics, respectively. i–n, Spontaneous 5-HT and DA release. i, Representative pseudocolor images of the cumulative spontaneous transients during a 10-min recording. Similar results were observed for 7 slices. Scale bar, 100 μm. j, Representative time-lapse pseudocolor images, and ΔF/F0 traces of ROIs (10 μm in diameter) from the area indicated by the gray dashed rectangle in i. Scale bar, 20 μm. k, Number of transients in g5-HT3.0 and rDA3m fluorescence. Two-tailed paired t-tests, P = 0.0226. l, Distribution of the peak response of individual events. m, Example traces showing the rise and decay kinetics (t50) of g5-HT3.0 and rDA3m (left), and the distribution of individual events (right). n, Distribution of the area of individual events. n = 4 slices from 3 mice in c,d,g,h; n = 7 slices of 3 mice in k; n = 1060 and 47 events for g5-HT3.0 and rDA3m, respectively, from 7 slices of 3 mice in l–n. Data are shown as mean ± s.e.m. in c,d,g,h,k, with the error bars or shaded regions indicating the s.e.m. *P < 0.05, n.s., not significant.