Fig. 1: OXT affects astrocytic proteins in vitro and in vivo, and impairs gap-junctional intercellular communication (GJIC) in primary rat cortical astrocytes in a MEK, PKC and OXTR-dependent manner.
From: OXTR-mediated signaling in astrocytes contributes to anxiolysis
a Heatmap of percentage changes in protein or protein phosphorylation levels in primary astrocytes following 10-min exposure to 500 nM OXT (upper panel), and in brain tissue punches taken from the PVN, hippocampus and amygdala 10 min or 20 min after icv administration of OXT (1 nmol/5 µl) (lower panel). Downregulations are colored in blue, while upregulations are colored in red. b Representative images of streptavidin staining visualizing the distance of biocytin diffusion and, thus, reflecting GJIC within the astrocytic network. Exposure of primary astrocytes to either OXT (500 nM) or the gap-junction blocker carbenoxolone (CBX, 1 µM) impaired relative GJIC compared to vehicle (Veh) treatment. Scale bar = 100 µm. c Dose-dependent effects of OXT (applied for 10 min) on relative GJIC in primary astrocytes (treatment F4,73 = 6.847, p < 0.001 versus Veh; **p = 0.001 for 10 nM; ***p = 0.0004 for 1000 nM). d Relative GJIC of astrocytes pre-treated for 1 h with either Veh, the MAP kinase inhibitor U0126 (1 µM), the PKC inhibitor Gö6983 (10 µM) or the OXTR antagonist L368,889 (1 µM) prior to OXT exposure (500 nM, 15 min) or AVP exposure (1 µM, 15 min) (interaction: F3,62 = 5.574, p = 0.002; *p = 0.01 Veh/OXT vs. Veh/Veh treatment). e Relative GJIC in astrocytic cultures after treatment with CBX (t15 = 14.75, ***p < 0.0001 versus Veh). f Representative immunocytochemical images of astrocytes stained for Cx43 (green), the tight-junction protein ZO1 (red) and DAPI (blue) showing lower levels of Cx43/ZO1 colocalization in OXT- compared with Veh-treated cells. White arrows indicate points of Cx43/ZO1 colocalization. g Dose-dependent effects of OXT (15 min) on Cx43 localization at cell-cell contacts (treatment F5,36 = 8.810, p < 0.001 versus Veh; *100 nM: p = 0.050, ***500 nM: p = 0.0002, **1 µM: p = 0.003). h Effect of differing exposure times to OXT (500 nM) on Cx43 localization at cell-cell contacts (treatment F5,36 = 6.077, p < 0.001 vs. basal (t = 0); **p = 0.005 for 10 min; *p = 0.024 for 15 min; **p = 0.003 for 180 min). i Impact of OXT (500 nM, 15 min) or AVP (1 µM, 15 min) on Cx43/Zo1 colocalization following pre-treatment with either Veh, U0126, Gö6983 or L368,889 (see legend in D) (interaction: F3,46 = 5.267, p = 0.003; **p = 0.008 for Veh/OXT vs. Veh/Veh; ** p = 0.001 U0126/OXT vs Veh/Veh). Data represent mean absolute or relative values +/−SEM.
