The brain contralaterally projects to the liver.

(A) Representative 3D images of TH-immunostained mouse liver (left), with quantification of sympathetic axon density and lobe volume (right). Scale bar, 3,000 μm. (B) Representative immunofluorescence images of PRV-labeled neurons (EGFP) in the left (top) and right (bottom) LPGi following PRV injections into the right anterior (left) and median (right) lobes. Scale bars, 100 μm. (C) Quantification of PRV-labeled neurons in left and right LPGi across different hepatic lobes: left lateral, median, right posterior, right anterior, caudate, and porta hepatis (n = 3). (D) Sankey diagram showing projection patterns from left and right LPGi to individual hepatic lobes. (E and F) Representative slices of EGFP+ and mRFP+ neurons in left (top) and right (bottom) LPGi following PRV-EGFP (right anterior lobe) and PRV-mRFP (median lobe) injections. Proportions of EGFP+, mRFP+, and co-labeled neurons in left and right LPG (F, n = 3). Scale bars, 100 μm. Data are expressed as means ± SEM, with individual values shown for all experiments. *p < 0.05, **p < 0.01. Unpaired Student’s t-test for (C).

LPGi stimulation raises blood glucose by driving contralateral, lobe-specific hepatic glycogenolysis.

(A) Scheme for injecting AAV-DIO-hM3Dq or AAV-DIO-mCherry (top). Blood glucose levels following chemogenetic activation of the left, right, or bilateral LPGi, measured at −10, 30, 60, 90, and 120 min (bottom, n = 6). (B and C) Representative immunofluorescence images of pGS (B) and pPYGL (C) expression in the median (top) and right anterior (bottom) lobes after 1 h chemogenetic activation of left, right, or bilateral LPGi, with quantification of relative fluorescence intensity (n = 5). Scale bars, 100 μm. (D) Western blot analysis of pPYGL, PYGL, pGS, and GS in median (left) and right anterior (right) lobes after 1 h activation, with densitometric quantification (bottom, n = 3). (E) Plasma concentrations of norepinephrine (NE) (left, n = 5) and epinephrine (EPI) (right, n = 5) following 1 h activation. (F) NE contents in median (left, n = 5) and right anterior (right, n = 5) lobes after 1 h activation. Data are expressed as means ± SEM, with individual values shown for all experiments. *p < 0.05, **p < 0.01, ***p < 0.001; #p < 0.05, ##p < 0.01, ###p < 0.001; $$$p < 0.001. One-way ANOVA with the Tukey test for (A -F).

LPGi maintains glucose balance through compensatory use of ipsilateral hepatic lobes under contralateral lobe impairment.

(A and B) Blood glucose levels in the left- (A, n = 7) or right-sided (B, n = 7) lobes denervated mice. Western blot of TH protein in median and right anterior lobes after denervating left- (top) or right-sided (bottom) lobes. (D and E) Representative immunofluorescence images of pGS (top) and pPYGL (bottom) expression in right anterior and median lobes in left- (D) or right-sided (E) lobes denervated mice, with quantification of relative fluorescence intensity (right, n = 5). Scale bars, 100 μm. (F and G) Western blot analysis of pPYGL, PYGL, pGS, and GS proteins in median and right anterior lobes (top) in left- or right-sided lobes denervated mice, with densitometric quantification (bottom, n = 3). (H) NE contents of median and right anterior lobes in left- or right-sided lobes denervated mice (n = 6). (I) Representative immunofluorescence images of c-FOS expression in left and right LPGi in left- or right-sided lobes denervated mice. Scale bars, 100 μm. Data are presented as means ± SEM, with individual values shown for all experiments. *p < 0.05, **p < 0.01, ***p < 0.001. Unpaired Student’s t-test for (A-B, D-H).

Sympathetic projections to the liver decussate at the porta hepatis, progressively formed during development.

(A) Representative 3D images of mice following PRV injection into right- (left) or left-sided (right) lobes. Scale bars, 3000 µm. (B) Representative 3D images of CG-SMG projections to peripheral organs. Scale bar, 3,000 μm. (C) Representative images of sympathetic axons after unilateral CG injection, with quantification of EGFP-labeled axon distribution across hepatic lobes. Scale bars, 100 μm. (D) Representative images of WGA-labeled neurons in left and right CG, with percentages of WGA555+, WGA647+, and co-labeled neurons. Scale bars, 100 μm. (E) Representative images showing the developmental progression of CG-SMG projections to the liver from postnatal week 0 to week 2. Scale bars, 3,000 µm.

Supplementary evidence for contralateral projections from the brain to the liver, related to Figure 1.

(A) Schematic illustration of hepatic lobes with anatomical labeling. (B) Represent images of hepatic lobes after PRV-EGFP injections. Scale bars, 150 μm. (C-F) Representative immunofluorescence images of PRV-labeled neurons (EGFP) in the left (top) and right (bottom) LPGi following PRV injections into left lateral lobe (C), right posterior lobe (D), caudate lobe (E), and porta hepatis (F). Scale bars, 100 μm.

Chemogenetic activation of LPGi induces c-Fos expression, promotes hepatic glycogenolysis, and activates glycogenolytic enzymes, related to Figure 2.

(A) Representative immunofluorescence images of c-FOS+ LPGi neurons in hM3Dq mice following 1 h stimulation (Left), with quantification of c-FOS+ neuron percentage (right, n = 3). Scale bars, 100 μm. (B) Representative PAS staining images of median (top) and right anterior (bottom) lobes in hM3Dq mice following 1 h stimulation, with semi-quantitative analysis of glycogen levels (right, n = 5). Scale bars, 100 µm. (C and D) Western blot analysis of PEPCK and G6PC proteins in the median (C, top) and right anterior (D, top) lobes, with densitometric quantified (bottom, n = 3). (E and F) Enzyme activity of PEPCK and G6PC in the median (E) and right anterior (F) lobes (n = 3). Data are expressed as means ± SEM, with individual values shown for all experiments. *p < 0.05, ***p < 0.001; ##p < 0.01, ###p < 0.001; $$$p < 0.001. A one-way ANOVA with the Tukey test was used for (A-F).

Optogenetic activation of LPGi confirms contralateral lobe-specific mobilization observed in Figure 2.

(A) Scheme for injecting AAV-DIO-ChR2 or AAV-DIO-mCherry (top). Blood glucose levels following optogenetic activation of left, right, or bilateral LPGi, measured at −10, 30, 60, 90, and 120 min (bottom, n = 6) (B and C) Representative immunofluorescence images of pGS (B) and pPYGL (C) expression in median (top) and right anterior (bottom) lobes after 1 h optogenetic activation of left, right, or bilateral LPGi, with quantification of relative fluorescence intensity (n = 5). Scale bars, 100 μm. (C) Western blot analysis of pPYGL, PYGL, pGS, and GS in median (left) and right anterior (right) lobes after 1 h activation, with densitometric quantification (bottom, n = 3). (D) Plasma concentrations of norepinephrine (NE) (left, n = 5) and epinephrine (EPI) (right, n = 5) following 1 h optogenetic activation. (E) NE contents in median (left, n = 5) and right anterior (right, n = 5) lobes after 1 h optogenetic activation. Data are expressed as means ± SEM, with individual values shown for all experiments. *p < 0.05, **p < 0.01, ***p < 0.001; #p < 0.05, ##p < 0.01, ###p < 0.001; $p < 0.05, $$$p < 0.001. A one-way ANOVA with the Tukey test was used for (A -F).

Optogenetic activation of LPGi recapitulates c-Fos expression, hepatic glycogenolysis, and activity of glycogenolysis-related enzymes observed in Figure S2.

(A) Representative immunofluorescence images of c-FOS+ neurons in LPGi after 1 h optogenetic stimulation, with quantification of ratio of c-FOS+ neurons (right, n = 3). Scale bars, 100 μm. (B) Representative PAS staining images of the median (top) and right anterior lobes (bottom) lobes following 1 h optogenetic stimulation, with semi-quantitative analysis of glycogen levels (n = 5). Scale bars, 100 µm. (C and D) Western blot analysis of PEPCK and G6PC proteins in median (C) and right anterior( D) lobes, with densitometric quantification (n = 3). (E and F) Enzyme activity of PEPCK and G6PC in median (E) and right anterior lobes (F) (n = 3). Data are expressed as means ± SEM, with individual values shown for all experiments. *p < 0.05, **p < 0.01, ***p < 0.001; #p < 0.05, ##p < 0.01, ###p < 0.001; $$$p < 0.001. A one-way ANOVA with the Tukey test was used for (A-F).

Validation of sympathetic nerve ablation, enhanced glycogenolysis in non-denervated hepatic lobes, and accurate targeting of AAV and WGA injections, related to Figure 3 and 4.

(A) Densitometric quantification of TH protein in median and right anterior lobes after denervating left- (left) or right-sided (right) lobes. (B-E) Representative PAS staining images of median (top) and right anterior (bottom) lobes in left- (B) and right-sided (D) lobes denervated mice, with semi-quantitative analysis of glycogen levels (C and E, n = 5). Scale bars, 100 µm. (A) Represent slices of the CG in unilateral AAV-TH-axon-EGFP injected mice. Scale bars, 20 μm. (B) Represent slices of hepatic lobes after injecting WGA 647 and WGA 555 into left- and right-sided lobes. Scale bars, 50 μm. Data are expressed as means ± SEM, with individual values shown for all experiments. ***p < 0.001; Unpaired Student’s t-test for (A, C, and E).