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. 2023 Apr 12;12:e82649. doi: 10.7554/eLife.82649

Figure 5. The α3-GABAA receptor signaling within the MC4RDMH mediates feeding and glucose tolerance.

(A) Expression level of Gabra1- Gabra6 in the DMH neurons of control and Agrp-Lepr KO mice. (B) Immunostaining showing the expression of GABAA α3 in the DMH. Scale bar, 100 μm. (C–E) Colocalization of GABAA α3 and transsynaptically labeled neurons by WGA-ZsGreen in the DMH after injection of AAV9-DIO-WGA-ZsGreen into the ARC of AgrpCremice. Scale bar in E for C–E, 50 μm. (F–H) The food intake (F), body weight (G), and GTT (H) were performed in Mc4rCre::Rosa26fs-Cas9 mice with a bilateral injection of vehicle, AAV9-Gabra3sgRNA-tdTomato (GABAA-α3 KO), or AAV9-DIO-Gabra3cDNA-EYFP (GABAA-α3 OE) into the DMH. (I) Feeding efficiency in the mice described in F–H. (J) Weight of adipose tissues in the mice described in F–H. n=8 per group in F–J; *p<0.05 between Control and GABAA-α3 KO, #p<0.05 between Control and GABAA-α3 OE. (K–O) The food intake (K), body weight (L), GTT (M), feeding efficiency (N), and weight of adipose tissues (O) were performed in control mice and GABAA-α3 OE mice followed by the microinjection of leptin into ARC. (n=8 per group in K-O; *p<0.05 between Control +leptin and GABAA-α3 OE +leptin). (P–R) The 24-hr food intake (P), body weight change (Q), and refeeding test (R) were performed in HFD-treated with or without knockout of GABAA-α3. (n=8 per group; *p<0.05). Error bars represent mean ± SEM. unpaired two-tailed t test in A and N–R; one-way ANOVA and followed by Tukey comparisons test in I and J; two-way ANOVA and followed by Bonferroni comparisons test in F–H and K–M. (S) Diagram showing a leptin regulated GABAergic neural circuit reverses obesity. The GABAergic AgRPLepR→DMH circuit plays a critical role in control of leptin-mediated food intake, body weight, and glucose tolerance through the α3-GABAA signaling within the MC4RDMH neurons.

Figure 5—source data 1. The original data of body weight, food intake, and GTT for Figure 5.

Figure 5.

Figure 5—figure supplement 1. Quantification of gabra3 transcription levels in the DMH by qPCR 21 days after injection of AAV9-Gabra3sgRNA-tdTomato into DMH of Mc4rCre::Rosa26fs-Cas9 mice (A) or injection of AAV9-DIO-Gabra3cDNA-EYFP into DMH of Mc4rCre mice (B).

Figure 5—figure supplement 1.

(n=6 per group; *p<0.05; unpaired two-tailed t test).

Figure 5—figure supplement 2. The AgRP-DMH circuit regulates feeding and glucose tolerance depending on the α3-GABAA receptor signaling in the MC4RDMH neurons.

Figure 5—figure supplement 2.

(A–B) 1-hr food intake (A) and GTT (B) in mice with AAV9-fDIO-ChR2-EYFP into ARC, AAV9-DIO-Cas9-mCitrine and AAV9-Gabra3sgRNA-tdTomato injected into the DMH of NpyFlp::Mc4Cre mice followed with implantation of optic fiber in the DMH. n=8 per group; *p<0.05 between ChR2 +AAV-tdTomato and ChR2 +AAV-a3-sgRNA-tdTomato, #p<0.05 between ChR2 +AAV-tdTomato and AAV-tdTomato, ‡p>0.05 between ChR2 +AAV-a3-sgRNA-tdTomato and AAV-a3-sgRNA-tdTomato. Error bars represent mean ± SEM. One-way ANOVA and followed by Tukey comparisons test in A; two-way ANOVA and followed by Bonferroni comparisons test in B.