Fig. 2: Silencing of CeAGABA neurons reduces ARS-induced respiratory and behavioral changes. | Nature Communications

Fig. 2: Silencing of CeAGABA neurons reduces ARS-induced respiratory and behavioral changes.

From: GABAergic neurons in central amygdala contribute to orchestrating anxiety-like behaviors and breathing patterns

Fig. 2

a Schematic of the chemogenetic strategy by delivering a virus encoding hM4Di into the CeA. b Representative image showing immunohistochemical detection of hM4Di expression (green) in the CeA. Scale bar, 100 µm. c RNAscope-FISH images demonstrating colocalization of hM4Di (green) and Slc32a1 (Vgat, pink) mRNA. Scale bars, 20 μm. d Quantitative analysis confirming specific expression of hM4Di-eYFP in CeAGABA neurons. n = 4 mice. e Representative traces showing that bath application of CNO (5 μM) induced membrane hyperpolarization in a hM4Di-expressing CeAGABA neuron, as measured by whole-cell patch clamp slice recordings. f Schematic of the experimental procedure. g Distribution of RF (bin size 0.5 Hz) over a 40-min assay of ARS mice injected with saline and CNO. n = 12 mice per group. p < 0.0001 for 3 and 3.5 Hz. h Accumulative duration spent in active and quiescent states (n = 10 mice for each group). p = 0.0008 for active; p = 0.0008 for quiescent. Percent of time (i, n = 10 mice for each group) and accumulative duration (j, n = 10 mice for each group) of each behavior defined in Fig. 1c. CNO injection significantly reduced ARS-induced prolonged grooming time and increased eupnea duration (groom, p = 0.0059; eupnea, p = 0.0078). k Percent of time spent on each behavior in every 5-min interval (n = 10 mice for each group). Time-series changes in the duration of moving (l), grooming (m), sniff (n), eupnea (o) and sleep (p) in every 5 min. (m, 31-35 min, p = 0.0395). q CNO injection significantly decreased RF at 20- and 30-min mark post-ARS compared to saline treatment. p = 0.0028 for 20 min and <0.0001 for 30 min. *p < 0.05, **p < 0.01, ****p < 0.0001 by two-way ANOVA with Bonferroni’s multiple comparisons tests (g, l–q) and two-tailed unpaired t-test (h, j). All data are presented as the mean ± SEM. Source data is provided as a Source Data file. The materials depicted in panels a and f are created in BioRender. Xiaoyi, W. (2025) https://BioRender.com/i81e326.

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