Fig. 2: OXT alters the astrocytic morphological phenotype in a PKC, MEK and OXTR-dependent manner and astrocyte-neuron spatial relationships in vitro (in primary cortical rat astrocytes) and in vivo.
From: OXTR-mediated signaling in astrocytes contributes to anxiolysis
a Representative images of primary astrocytes stained for GFAP (green) and DAPI (blue) showing OXT-induced morphological alterations. Scale bar = 20 µm. b Dose-dependent effect of 10-min exposure to OXT on the length of the longest primary process in vitro (F4,539 = 2.760, p = 0.027; *p = 0.016 for 500 nM OXT vs. Veh). c Effect of 10-min OXT or AVP exposure on process length of primary astrocytes following pre-treatment with either Veh, U0126, Gö6983 or L368,889 (for details see legend to Fig.1; interaction: F3,646 = 4.480, p = 0.004; **p = 0.008 Veh/Veh vs. Veh/OXT. d, e in vitro effects of 180-min OXT exposure (d) on astrocytic F-actin levels reflected by phalloidin immunofluorescence (t13 = 3.225, **p = 0.007 vs. Veh), and (e) on phospho(Ser19)-myosin-light-chain-kinase (pMLC/Ser19) immunofluorescence (t12 = 3.136, **p = 0.009 vs. Veh). f Representative images of PVN astrocytes (GFAP; green) co-stained with neurophysin (OXT; red) and DAPI (blue) in rats 10 min after icv Veh or OXT (1 nmol/5 µl). White arrows mark points of GFAP/neurophysin (OXT) colocalization indicating astrocyte-neuron contacts. Scale bar = 10 µm. g–i Effects of icv infusion of OXT on primary process length after 10 min (g; t10 = 3.48, **p = 0.006), primary process number (H; t10 = 2.47, *p = 0.033) and GFAP/OXT-colocalization (I; t10 = 3.09, * p = 0.011) within the rat PVN. j Representative confocal microscopy image of a GFP-expressing mouse hippocampal astrocyte. Scale bar = 10 µm. k Representative 3D reconstructions of astrocytes created from Veh or OXT (500 nM, 10 min) -treated acute mouse hippocampal slices. Scale bar = 10 µm. l Assessment of cellular surface area from 3D reconstructed astrocytes (t20 = 3.302, **p = 0.004). m Assessment of cellular volume from 3D-reconstructed astrocytes (t20 = 2.155, *p = 0.046). n Representative confocal microscopy image of a GFP-expressing mouse hippocampal astrocyte. Scale bar = 10 µm. White dotted box indicates inlay for middle panel. Inlay includes deconvolved confocal image of astrocytic element (GFP; red), as well as deconvolved STED-images of pre-synaptic (VGlut1; Magenta) and post-synaptic (Homer1; green) markers. Scale bar for inlays = 1 µm. Right panel displays representative astrocyte processes in their spatial relationship to pre- and post-synaptic elements from slices treated with either Veh or OXT. o Average synapse/astrocyte distance per analyzed inlay (t15 = 3.003, **p = 0.009). Data represent mean absolute or relative values +/−SEM.
