Fig. 4: Binge drinking rapidly alters the intracellular calcium levels to release glutamates and activate mGluR5 of Kupffer cells. | Nature Communications

Fig. 4: Binge drinking rapidly alters the intracellular calcium levels to release glutamates and activate mGluR5 of Kupffer cells.

From: Binge drinking triggers VGLUT3-mediated glutamate secretion and subsequent hepatic inflammation by activating mGluR5/NOX2 in Kupffer cells

Fig. 4

a A schematic figure of Ca2+ channels and transporters in HEPs. Heatmaps of genes in Pair and EtOH group (n = 3/group). b Representative intravital liver imaging stained with Fluo-4 AM and lectin. Statistical analysis of Fluo-4 AM-positive cells per image field (n = 4/group). Scale bar, 100 μm. c Heatmaps showing relative mRNA expression of SNARE complex-related genes analyzed by RNA-sequencing before and after binge drinking in EtOH-fed mice (n = 3/group). d Glutamate concentration in the supernatant after in vitro additional EtOH treatment of HEPs isolated from 2-week Pair-fed and EtOH-fed mice (n = 3/group). e Flow cytometry analyses of mGluR5 expression and ROS generation in isolated KCs. f ROS generation and gene expression of Grm5, Cybb, and Cxcl1 in KCs treated with mGluR5 selective agonist (CHPG) or monosodium glutamate (MSG) compared to vehicle (n = 4/group). g Size distribution analysis of HEPs isolated from Pair- or EtOH-fed mice. Bar graph shows the average diameter of HEPs in each group (n = 6/group). h Transmission electron microscopy in liver section fed with Pair or EtOH diet for 2 weeks. Bar graph indicates the average distance between adjacent hepatocytes (HEPs) and Kupffer cells (KCs), measured across multiple cell pairs per mouse (n = 12 HEP–KC pairs/group). Scale bar, 5 μm. i Co-culturing of KCs directly or indirectly with HEPs isolated from EtOH-fed mice, followed by treatment with 50 mM EtOH for 6 h. Relative mRNA expression of Cybb, and Il1b in isolated KCs (n = 6/group). LDH was measured in the supernatant (n = 4/group). Statistical comparisons were made using one-way ANOVA with Tukey’s multiple comparisons test (b, i), and multiple comparisons by two-stage step-up method of Benjamini, Krieger and Yekutieli. F (d), or two-tailed unpaired t-test (f–h). Data are presented as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001. Source data are provided as a Source Data file.

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