Fig. 2: Expression of constitutively active GCGRs in liver results in high blood glucose levels in non-mammals.

a–d, Intravenous injection in chicks (a). Ns, normal saline; Ad, adomeglivant; Des, des-His¹Pro⁴-glucagon amide. b–d, Blood glucose and glycogen content (b), GCGR and G6pc2 expression (c), and GPa enzyme activity (d) four hours after injection. n = 3 per group. e–h, Intravenous injection of AAV-shGgGCGR in chicken embryo blood vessels (e). f–h, Blood glucose (f), GCGR expression (g), and glycogen content and GPa enzyme activity (h) in chicks 19 days after hatching. n = 3 per group. i–l, Intravenous injection of AAV-shGgGCGR in chicks (i). j–l, Blood glucose, glycogen content (j), GCGR, G6pc2 expression (k), GPa enzyme activity (l) ten days after injection. n = 3 per group. m–p, Intravenous injection of AAV-shLsGCGR in L. striata (m). n–p, Blood glucose and glycogen content (n), GCGR and G6pc1 expression (o), and GPa enzyme activity (p) four days after injection. n = 3 per group. q–t, Intravenous injection of AAV-shPvGCGR in P. vitticeps (q). r–t, Blood glucose and glycogen content (r), GCGR expression (s), and G6pc2 and Pck1 expression (t) five days after injection. n = 3 per group. u–x, Intrahepatic injection of AAV-HsGCGR and AAV-ScGCGR in E. macularius (u). v–x, Blood glucose and glycogen content (v), quantification of hepatic HsGCGR and ScGCGR mRNA (w), and G6pc3 and Pck2 expression (x) three days after injection. n = 3 per group. y–ab, Doxycyclxine-induced hepatic GgGCGR expression in D. rerio (y). z–ab, Blood glucose (z), HsGCGR and GgGCGR expression (aa), and Pck1 and G6pc1 expression (ab) 24 h after Dox induction. n = 4 per group. Data are mean ± s.e.m. from at least three independent experiments, performed in technical triplicate and were analysed by one-way ANOVA and Dunnett’s post hoc test (b–d,v,x,ab), two-way ANOVA followed by Šídák’s post hoc test (f,g), two-tailed unpaired Student’s t-test (h,j–l,n–p,r–t), two-tailed unpaired Welch’s t-test (w) or two-way ANOVA followed Tukey’s post hoc test (z). Cartoons were created in BioRender; Quanwei, Y. (2025) https://BioRender.com/j11v652. Sample sizes, statistical methods and P values are presented in Supplementary Table 15.