Fig. 4: Adrenoreceptor upregulation after fear conditioning enables FR-BAE induction.

a, t-Distributed stochastic neighbour embedding (tSNE) clustering of scRNA-seq data from amygdalar astrocytes across NoFC, FC, NoFR and FR conditions (n = 4 mice per group). b, Dot plots showing Fos expression. Dot colour indicates z-score (grey, depletion; green, enrichment). c, Left, dot plots showing adrenoreceptor gene expression. Undetected receptors (Adra1d, Adra2c and Adrb3) are not shown. Right, Adra1a and Adrb1 upregulation in NoFR versus FC. Horizontal lines represent median. Mann–Whitney test. d, RNAscope timeline. e, RNAscope images and violin plots of Adra1a and Adrb1 in Slc1a3⁺ astrocytes (18–28 cells from 5 mice per group). One-way ANOVA, Dunnett’s test versus NoFC. f, Co-expression index for Adra1a and Adrb1 (Methods) peaks at 1 day post-FC. One-way ANOVA, Dunnett’s test versus NoFC. n = 18–28 cells, 5 mice per group. g, BAE tagging timeline. h, Representative images and quantification of tagged FR-BAE, peaking at 1 day post-FC. n = 4–5 mice per group. One-way ANOVA, Dunnett’s test versus NoFC. i, RNAscope of Adrb1 and Fos in Slc1a3⁺ astrocytes (FR). Arrows indicate a Fos and Adrb1-high cell (representative images from 18 sections, 6 mice). Scale bar, 10 μm. j, Pearson’s correlation coefficient (r) for Fos and Adrb1 expression (z-scores). Fos-high: z > 2 (3.1% cells); Adrb1-high: z > 0.5 (26.3%). Rationale in Methods. n = 814 cells from 6 mice. k, Adrb1 expression is increased in Fos-high astrocytes (25 Fos-high cells, 789 Fos-low cells) and Fos is enriched in Adrb1-high astrocytes (215 Adrb1-high cells, 600 Adrb1-low cells). Mann–Whitney test. l, Schematic of Adra1a or Adrb1 conditional knockout (cKO). m, RNAscope of Adra1a and Adrb1 in Slc1a3⁺ astrocytes. Ctrl, control. n, Violin plots showing reduced targeted receptor after conditional knockout (77–117 cells, 3 mice per group). Horizontal lines represent the median. o,p, Images (o) and quantification (p) of FR-BAEs, showing decreased density following Adra1a or Adrb1 conditional knockout. Scale bar, 200 μm. n = 5–9 mice, One-way ANOVA, Dunnett’s test versus FR controls. Data are mean ± s.e.m. Sample sizes and replicates in Supplementary Table 1.