Extended Data Fig. 8: Chemogenetic manipulation of astrocyte Ca²⁺ via hM4Di-actuation.

a, Optical fibre tip location for cyto-GCaMP6f in vivo astrocyte Ca²⁺ fibre photometry following pre-extinction CNO in animals expressing hM4Di or a mCherry control virus. b, Population CS-related Ca²⁺ activity during early extinction (E-Ext, first 5 trials of extinction session 1), late extinction (L-Ext, last 5 trials of extinction session 2), extinction retrieval (E-Ret), and fear renewal (F-Ren) (Control n = 8, hM4Di n = 6 mice). c, Quantification of CS-related Ca²⁺ activity shown in panel b (2-way repeated measures ANOVA and Šídák’s post hoc tests, during E-Ext *P = 0.0421, L-Ext *P < 0.0001, E-Ret *P = 0.0368, F-Ren *P = 0.0115, Control n = 8, hM4Di n = 6 mice). Extended timeline measuring effects of 3 mg/kg CNO on Ca²⁺ activity in animals placed in a neutral-cage, with corresponding quantification of d, area under the curve (AUC) e (2-way repeated measures ANOVA and Šídák’s post hoc tests, Control vs hM3Dq at all intervals *P < 0.0001, Control vs hM4Di 30 min #P < 0.0001, 60 min #P = 0.0005, 90 min #P = 0.0093, 120 min #P < 0.0001) and Ca²⁺ events f (unpaired t-test for transients at minute 120, *P = 0.0025, #P < 0.0001, Control n = 9, hM3Dq n = 5, hM4Di n = 5 mice). g, Effects of pre-retrieval 3 mg/kg CNO on freezing levels during fear conditioning (F-Con), Ext, E-Ret, and F-Ren in hM3Dq or mCherry control virus-expressing animals (2-way repeated measures ANOVA, Šídák’s or Fisher’s LSD post hoc tests, P > 0.05, Control n = 4, hM3Dq n = 8 mice). h, Effects of pre-retrieval 3 mg/kg CNO on freezing levels during F-Con, Ext, E-Ret, and F-Ren in animals expressing hM4Di or mCherry control virus (2-way repeated measures ANOVA, Šídák’s or Fisher’s LSD post hoc tests, P > 0.05, Control n = 4, hM4Di n = 8 mice). i, Effects of 3 mg/kg CNO on novel open field total distance and centre time (2-tailed unpaired t-tests, P > 0.05, Control n = 9, hM4Di n = 12 mice). j, Effects of 3 mg/kg CNO on elevated plus-maze total distance and open arm time (2-tailed unpaired t-tests, P > 0.05, Control n = 9, hM4Di n = 12 mice). k, Effects of 3 mg/kg CNO on light-dark exploration test total distance and dark compartment time (2-tailed unpaired t-tests, P > 0.05, Control n = 9, hM4Di n = 13 mice). Horizontal lines above traces in b denote permutation test-determined significant difference from chance for hM4Di (blue) and Control (black) or significant difference between groups (orange). Fluorescence values z-scored to the 30 min pre-CNO or 5-s pre-CS periods. Data mean ± SEM.

Source data