Journal: Nature Communications

Article Title: Stress induces oxytocin-Gαi-dependent remodeling of astrocytes to shape neuronal response in the amygdala

doi: 10.1038/s41467-025-68114-4

Figure Lengend Snippet: a Imaging of CeL/C astrocytes using targeted expression of eGFP using rAAV-GFAP-eGFP. b Representative micrograph showing synapses and surrounding astrocytic processes following IMARIS-based 3D reconstruction. Arrowheads show a putative synapse contacted by an astrocyte. eGFP: astrocyte, vGlut1/2: post-synaptic, Homer1: pre-synaptic c Quantification of the contacts between synapses and astrocyte processes per reconstructed astrocyte. n = 5 animals in each group. d Analysis of BLA-to-CeL/C synaptic transmission using optogenetic stimulation of the BLA axons and recordings of the EPSCs in CeL/C neurons in naïve and stressed mice. Injection of AAV-CaMKII-ChR2 was performed in the BLA of WT mice, and a second pair of injection containing rAAV-GFAP-Cre was done in the CeL/C of OTR-lox mice. 3 weeks after, we measured the currents evoked by BLA neurons activation in CeL/C neurons of presynaptic BLA neurons. Lower right panel: representative recording of EPSCs in CeL/C neurons evoked during photostimulation. e , f Time course and quantification of photostimulation-evoked EPSCs amplitude. n (neurons, mice): WT naive=14, 5, WT stress=19, 5, GFAP OTR cKO naïve=17, 9, GFAP OTR cKO stress=13, 5. g Experimental approach to record K + currents in CeL/C astrocytes following optogenetic stimulation of BLA neurons in naïve and stressed mice. AAV-CaMKII-ChR2 was injected in the BLA of mice, and 3 weeks after, the currents evoked by BLA neuron activation were measured in CeL/C astrocytes. Lower right panel: representative recording of photstimulation-evoked currents in control condition, after the application of Ba 2+ , and the subtraction of both (I K ). h , i Time course and quantification of photostimulation-evoked astrocytic current induced by BLA neurons stimulation before (black) and after a 15-min incubation in Ba 2+ (200µM, dark grey). The subtracted K + current is shown in light grey. n (astrocyte, mice): WT naive=13,6, WT stress=11, 8, GFAP OTR cKO naïve=11, 6, GFAP OTR cKO stress=8, 6. j Quantification of K ir 4.1 levels in naïve and stressed mice using 1mm punches of mice CeA was done on 400µm slices from naïve or stressed mice. k Image of representative Western blot reacted with antibody against K ir 4.1 in six animals from the indicated experimental groups. l Quantification of K ir 4.1 levels in amygdala punches. n naïve =6, n stress =8 mice. Data are expressed as mean across animals ± SEM for ( c ) and ( l ), and as mean across cells ± SEM for ( e ), ( f ), ( h ), and ( i ). Detailed statistics can be found in Source Data Table . * p < 0.05, ** p < 0.01. Drawings created in BioRender: Charlet, A. (2025) https://BioRender.com/bkv8qa0 .

Article Snippet: After 3x10min rinses in 9 mM NaCl Tris-HCl Buffer + 0.1% Tween 20 (TBST), membranes were blocked in TBST + 5% milk for 1 h at room temperature (RT) and incubated overnight at 4 °C sequentially with the following primary antibodies: anti-K ir 4.1 (1:1000, Rabbit; Alomone, #APC-035), anti-actin or anti-Tom20 in TBST + 5% milk.

Techniques: Imaging, Expressing, Transmission Assay, Injection, Activation Assay, Control, Incubation, Western Blot