1. Volume 23, Issue 1, Pages 103-112 (January 2016)
Striatal Dopamine Links Gastrointestinal Rerouting to Altered Sweet Appetite 
Wenfei Han, Luis A. Tellez, Jingjing Niu, Sara Medina, Tatiana L. Ferreira, Xiaobing Zhang, Jiansheng Su, Jenny Tong, Gary J. Schwartz, Anthony van den Pol, Ivan E. de Araujo 
Cell Metabolism 
Volume 23, Issue 1, Pages (January 2016)
DOI: /j.cmet
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2. Cell Metabolism 2016 23, 103-112DOI: (10.1016/j.cmet.2015.10.009)
Copyright © 2016 Elsevier Inc. Terms and Conditions

3. Figure 1
Chronic Exposure to Sugar Adjusts the Sensitivity to Satiating Intra-gastric Preloads, an Effect Mediated by the Duodenal-Jejunal Route
(A) Behavioral preparation. Mice are placed in a behavioral box where licks for the artificial sweetener sucralose trigger an infusion pump that delivers either glucose (blue) or more sucralose (red) directly into the gut. After 13 days of daily 1-hr exposure sessions, mice are tested for intake levels after administration of satiating intra-gastric preloads of either glucose or sucralose that correspond to average daily intake levels.
(B) The behavioral protocol was initially used in mice with preserved gastrointestinal routing.
(C) Glucose-exposed mice (n = 12) self-administer significantly more sucralose than sucralose-exposed mice (n = 12) after an intra-gastric preload of glucose (t[22] = 4.0, ∗p = 0.001). y axis indicates final volume self-administered into the gut, so that the bottom half of the y axis (yellow area) shows the volume of the preload, and the upper half the amounts consumed after the preload. Note the preload volumes correspond to their baseline intake of each solution.
(D) Glucose-exposed mice (n = 8) self-administer significantly more glucose than sucralose-exposed mice (n = 8) after an intra-gastric preload of glucose (t[14] = 2.6, ∗p < 0.02).
(E) Glucose-exposed mice (n = 6) self-administer significantly more glucose than sucralose-exposed mice (n = 6) after an intra-gastric preload of sucralose (t[10] = 6.3, ∗p = 0.001).
(F) Glucose- and sucralose-exposed mice (n = 6 each group) self-administer similar levels of sucralose after an intra-gastric preload of sucralose (t[10] = 0.58, p = 0.57).
(G) The behavioral protocol was also used in mice sustaining a duodenal-jejunal bypass (DJB) intervention. Scheme representation of the DJB, which involves pylorus ligation (black line drawn under stomach) and a gastric-jejunal anastomose.
(H) DJB glucose-exposed mice (n = 5) self-administer significantly less sucralose than glucose-exposed sham controls (n = 5) after an intra-gastric preload of glucose (with respect to preload: SHAM = 202.4% ± 47.4%; DJB = 11.1% ± 8.5%, Paired t test t[4] = 4.2, ∗p = 0.013).
(I) DJB glucose-exposed mice self-administer significantly less glucose than glucose-exposed sham controls after an intra-gastric preload of glucose (SHAM = 93.7% ± 8.6%; DJB = 50.3% ± 12.2%, t[4] = 8.4, ∗p = 0.001).
(J) DJB glucose-exposed mice showed a tendency to self-administer significantly less glucose than glucose-exposed sham controls after an intra-gastric preload of sucralose (SHAM = 151.2% ± 15.8%; DJB = 90.19% ± 8.7%, t[4] = 6.8, ∗p = 0.002).
(K) DJB glucose-exposed mice self-administered similar amounts of sucralose to glucose-exposed sham controls after an intra-gastric preload of sucralose (SHAM = 237.3% ± 26.5%; DJB = 204.4% ± 33.7%, t[4] = 1.0, p = 0.37).
I, intake after preload; p, preload; n.s., statistically non-significant. Data are represented as mean ± SEM.
Cell Metabolism  , DOI: ( /j.cmet )
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4. Figure 2
The Duodenal-Jejunal Route Regulates Gut-Induced Striatal Dopamine
(A) Assay for assessing gut-induced striatal dopamine in awake mice. Injections of sucralose or glucose into stomach are performed concomitantly to brain microdialysis in dorsal or ventral striatum.
(B) Representation of dialysis probe locations into dorsal (blue) and ventral (red) striatal sectors.
(C) Intra-gastric infusions of glucose induced significantly greater dopamine release in dorsal striatum of sham mice (n = 5) compared to sucralose infusions (two-way repeated-measures ANOVA, sampling time × intra-gastric stimulus interaction F[14,56] = 4.39, ∗p < 0.001). The y axis represents percent change with respect to baseline levels. Mean absolute dopamine concentrations for baseline samples were 0.24 ± 0.04 pg/μL and 0.24 ± 0.03 pg/μL for glucose and sucralose sessions, respectively.
(D) This effect was completely abolished in DJB mice (n = 5, F[14,70] = 0.7, p = 0.7). Mean absolute dopamine concentrations for baseline samples were 0.22 ± 0.01 pg/μL and 0.20 ± 0.01 pg/μL for glucose and sucralose sessions, respectively.
(E) Intra-gastric infusions of glucose induced significantly greater dopamine release in ventral striatum of sham mice (n = 5) compared to sucralose infusions (F[14,56] = 3.9, ∗p < 0.001). Mean absolute dopamine concentrations for baseline samples were 0.09 ± 0.03 pg/μL and 0.09 ± 0.02 pg/μL for glucose and sucralose sessions, respectively.
(F) Similar effects were observed in DJB mice (n = 5, F[14,56] = 7.3, ∗p < 0.001). Mean absolute dopamine concentrations for baseline samples were 0.10 ± 0.01 pg/μL and 0.09 ± pg/μL for glucose and sucralose sessions, respectively.
(G) Injections of sucralose or glucose into different parts of intestine of awake mice are performed concomitantly to brain microdialysis in dorsal striatum of sham mice.
(H) Small 100mg/kg glucose infusions into duodenum induced significantly greater dopamine release than equivalent jejunal infusions (F[14,56] = 4.1, ∗p < 0.001). Mean absolute dopamine concentrations for baseline samples were 0.21 ± 0.01 pg/μL and 0.23 ± 0.02 pg/μL for duodenal and jejunal sessions, respectively.
(I) Similar experiments were performed in DJB mice.
(J) The effects were not preserved when infusions were made into the bypassed duodenum and compared to jejunum (between-subjects two-way ANOVA F[14,112] = 1.390, p = 0.17). Mean absolute dopamine concentrations for baseline samples were 0.19 ± 0.01 pg/μL and 0.22 ± 0.03 pg/μL for duodenal and jejunal sessions, respectively.
(K) Mice were implanted with portal (n = 5) or jugular (n = 5) intravenous catheters, through which the same dose of glucose as in intestine was infused.
(L) Portal infusions of glucose produced significant increases over baseline, and were significantly greater than equivalent infusions into the jugular vein (two-way mixed-model ANOVA, sampling time × vein interaction F[14,112] = 9.2, ∗p < 0.001).
Mean absolute dopamine concentrations for baseline samples were 0.23 ± 0.02 pg/μL and 0.25 ± 0.01 pg/μL for portal and jugular sessions, respectively. Saline infusions did not produce vein-specific effects (F[14,112] = 0.4 p = 0.9). n.s., statistically non-significant. Data are represented as mean ± SEM.
Cell Metabolism  , DOI: ( /j.cmet )
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5. Figure 3
Optogenetic Stimulation of D1r-Neurons in Dorsal Striatum Diminishes Sensitivity to Satiating Intra-Gastric Preloads and Counters the Appetite-Suppressing Effects of Duodenal-Jejunal Rerouting
(A) Detected licks trigger 3 s intra-gastric infusions concomitantly to 3 s blue light laser pulse to either dorsal or ventral striatum. Animals were tested after administration of glucose or sucralose intra-gastric preloads.
(B) EYFP visualization confirms that ChR2 transfection was contained to DS. Note the dense bundle of axon terminals in entopenducular nucleus and substantia nigra, pars reticulata, and absence of labeled fibers in globus pallidus, confirming exclusive labeling of D1r-neurons. Panel is a composite of sequential 10× images of the entire sagital section.
(C) Whole-cell slice recordings reveal robust optogenetic activation of a ChR2+ D1r-neuron by 1/5/10/20 Hz blue light pulses (10 ms) and by a continuous light train.
(D) The y axis indicates final volume self-administered into the gut, so that the bottom half of the y axis (yellow area) shows the volume of the preload, and the upper half the amounts consumed after the preload. Laser pulses overrode the satiating effects of intra-gastric glucose preload in a region-dependent manner (two-way mixed model ANOVA, brain region × laser state F[2,15] = 5.6, p = 0.01). In animals transfected with ChR2 in D1r-neurons of dorsal striatum, laser pulses induced a drastic increase in sucralose intake despite the glucose preload (laser ON versus OFF, n = 6, paired t test t[5] = 7.0, Bonferroni ∗p < 0.01). However, no effect was observed when ventral striatum was activated (n = 6, t[5] = 0.5, p = 0.5). Control mice (D1r-Cre mice implanted with optical fibers and transfected in dorsal striatum with a blue-light insensitive ion channel) showed no light-dependent effects as expected (n = 6, t[5] = 0.7, p = 0.48). Laser activation in dorsal striatum significantly increased intake compared to both control (t[10] = −4.211, ∗∗p = 0.002) and ventral striatum (t[10] = 4.5, ∗∗∗p = 0.001) mice.
(E) In D1r-Cre DJB mice transfected with ChR2 in dorsal striatum, laser pulses induced a marked increase in sucralose intake despite the glucose preload (laser ON versus OFF, n = 5, paired t test t[4] = 3.4, ∗p = 0.027).
(F) Similar effects were observed for glucose intake after sucralose preload (t[4] = 10.4, ∗p < 0.001).
I, intake after preload; p, preload. n.s., statistically non-significant. Data are represented as mean ± SEM.
Cell Metabolism  , DOI: ( /j.cmet )
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6. Figure 4
Cell-Specific Ablation of D1r-Neurons in Dorsal Striatum Enhances Sensitivity to Satiating Intra-gastric Preloads and Mimics the Appetite-Suppressing Effects of Duodenal-Jejunal Rerouting
(A) Strong labeling throughout dorsal striatum is observed when retrograde fluorescent beads were injected into globus pallidus, the exclusive target of D2r-expressing neurons of dorsal striatum.
(B) Injection site associated with globus pallidus injections.
(C) Weaker labeling throughout dorsal striatum is observed when retrograde fluorescent beads were injected into substantia nigra, pars reticulata, the exclusive target of D1r-expressing neurons of dorsal striatum.
(D) Injection site associated with substantia nigra, pars reticulata injections.
(E) Ablation of D1r-expressing cells in dorsal, but not in ventral, striatum resulted in glucose preloads producing strong inhibitory effects on sucralose intake (two-way ANOVA between-group effect F[3,26] = 30.6, p < 0.001). Post hoc, Bonferroni corrected tests: DS-casp versusVS-casp (t[13] = 9.5 ∗p < 0.004), DS-casp versus D1-CTL (t[12] = 10.9 ∗∗p < 0.004), DS-casp versus WT-casp (t[13] = 6.4, ∗∗∗p < 0.004). The lower half of the y axis (yellow area) shows the volume of the preload, and the upper half the amounts consumed after the preload. Note the preload volumes correspond to their baseline intake of each solution.
(F) Similar effects were observed during glucose intake after glucose preloads (F[3,26] = 35.3 p < 0.001). Post hoc, Bonferroni corrected tests: DS-casp versusVS-casp (t[13] = 8.4, ∗p < 0.004), DS-casp versus D1-CTL (t[12] = 11.6, ∗∗p < 0.004), DS-casp versus WT-casp (t[13] = 8.8, ∗∗∗p < 0.004).
(G) No effects were observed during glucose intake after sucralose preloads (F[3,26] = 0.01 p = 0.9) or (H) during sucralose intake after sucralose preloads (F[3,26] = 0.4 p = 0.7).
Data are represented as mean ± SEM.
Cell Metabolism  , DOI: ( /j.cmet )
Copyright © 2016 Elsevier Inc. Terms and Conditions