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. 2022 Jun 8;8(23):eabn5345. doi: 10.1126/sciadv.abn5345

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Fig. 6. — (A) Experimental paradigm for synaptophysin-mGFP–mediated anterograde viral tracing in GLP-1R-Cre mice. (B) Cre-dependent mGFP expression in the brainstem including NTS and DMV originating from GLP-1R afferents. (C) Schematic showing tracing of the input-output relationships between DMH^GLP-1R+ neurons. Rabies EnVA-ΔG-GFP virus was injected into the DMV. AAV-DIO-RVG and AAV-DIO-TVA-dsRed were coinjected into the DMH of GLP-1R-Cre mice. (D) Representative images showing a substantial rabies-GFP signal from DMV-projecting DMH^GLP-1R+ neurons in the NTS. (E) Schematic showing tracing of DMH^GLP-1R+ neuron–pancreas projection: PRV-GFP and AAV-DIO-mCherry were injected into pancreas and DMH, respectively, in GLP-1R-Cre mice. (F) Representative image showing GLP-1R⁺ neurons (red) and pancreas-projecting neurons (green). Representative pancreas-projecting GLP-1R⁺ neurons indicated by white arrow (left). PRV-positive cell proportion (relative to total GLP-1R cells) is about 20% (right). (G) Experimental paradigm for optogenetic stimulation of GLP-1R terminals in the DMV in GLP-1R-Cre mice. t = 0, prestimulation, t = 30, during stimulation, t = 60, post-stimulation. (H) Fasting blood glucose level is significantly decreased after exposure to blue light in AAV-DIO-ChR2-YFP–infected GLP-1R-Cre mice when compared to the control group (AAV-DIO-YFP–infected GLP-1R-Cre mice). ***P < 0.001 (ANOVA test). (I) Experimental paradigm for Exn4 injection in DMH of WT mice. (J) Fasting plasma insulin level increased 0.5 hours after Exn4 injection. ***P < 0.001 (t test). (K) Experimental paradigm for viral injection in DMH of GLP-1R-Cre mice. (L) Activation of GLP-1R neurons in the DMH increased fasting plasma insulin levels 1 hour after intraperitoneal injection of hM₃Dq-specific agonist, CNO. *P < 0.05 (t test). (M) Experimental paradigm for viral injection of AAV-Cre in GLP1R^f/f mice. (N) Plasma insulin levels are significantly lower in GLP-1R knockdown group than in control group. *P < 0.05 (t test). n numbers are indicated in each panel.