1. Volume 20, Issue 1, Pages 112-123 (July 2017)
Somatostatin Neurons in the Basal Forebrain Promote High-Calorie Food Intake 
Chen Zhu, Yun Yao, Yan Xiong, Mingxiu Cheng, Jing Chen, Rui Zhao, Fangzhou Liao, Runsheng Shi, Sen Song 
Cell Reports 
Volume 20, Issue 1, Pages (July 2017)
DOI: /j.celrep
Copyright © 2017 The Authors Terms and Conditions

2. Cell Reports 2017 20, 112-123DOI: (10.1016/j.celrep.2017.06.007)
Copyright © 2017 The Authors Terms and Conditions

3. Figure 1 Stimulation of BF SOM Neurons Selectively Induces Fat Intake
(A–C) Images showing ChR2-eYFP expression in VP VGAT neurons (A), vBF SOM neurons (B), and VP PV neurons (C).
(D) Photo-stimulation of VP VGAT neurons significantly increases standard chow intake (p < , experimental group, ± g, n = 5; control group, ± g, n = 7).
(E) Photo-stimulation of vBF SOM neurons has no effect on standard chow intake (p = , experimental group, ± , n = 9; control group, ± , n = 7).
(F) Photo-stimulation of VP PV neurons has no effect on standard chow intake (p > , experimental group, ± g, n = 6; control group, ± g, n = 6).
(G) Photo-stimulation of VP VGAT neurons significantly increases high-fat chow intake (p < , experimental group, ± g, n = 5; control group, ± g, n = 7).
(H) Photo-stimulation of vBF SOM neurons significantly increases high-fat chow intake (p = , experimental group, ± g, n = 9; control group, ± g, n = 7).
(I) Photo-stimulation of VP PV neurons has no effect on high-fat chow intake (p > , experimental group, ± g, n = 6; control group, ± g, n = 6).
VP, ventral pallidum; HDB, nucleus of the horizontal limb of the diagonal band; MCPO, magnocellular preoptic nucleus; SI, substantia innominata; vBF, including HDB, MCPO, and SI. Upper panel: brain outlines adapted from Figure 30 of The Mouse Brain in Stereotaxic Coordinates, Compact 3rd Edition, Keith Franklin and George Paxinos (© copyright Elsevier, All Rights Reserved). Scale bars, 200 μm. Data represented as mean ± SEM. All significance values were tested by one-way ANOVA and post hoc Sidak’s test.
See also Figures S2–S4.
Cell Reports  , DOI: ( /j.celrep )
Copyright © 2017 The Authors Terms and Conditions

4. Figure 2 Genetically Defined Neuron Types in the Basal Forebrain
(A) Fluorescence images showing eYFP virus-labeled VGAT neurons (green) and FISH for SOM markers (red) in the VP area of a VGAT-Cre mouse.
(B) Quantification of percentage of SOM-positive and negative cells in VP VGAT-eYFP cells, 9.1% of eYFP-labeled VGAT neurons show co-expression with SOM FISH markers.
(C) Fluorescence images showing eYFP virus-labeled VGAT neurons (green) and FISH for SOM markers (red) in the vBF area of a VGAT-Cre mouse.
(D) Quantification of percentage of SOM-positive and negative cells in vBF VGAT-eYFP cells, 28.7% of eYFP-labeled VGAT neurons show co-expression with SOM FISH markers.
(E) Fluorescence images showing eYFP virus-labeled SOM neurons (green) and FISH for VGAT markers (red) in the vBF area of a SOM-Cre mouse.
(F) Quantification of percentage of VGAT-positive and negative cells in vBF SOM-eYFP cells, 68.3% of eYFP-labeled SOM neurons show co-localization with VGAT FISH markers.
Green, red, and yellow arrows indicate cells that are eYFP-positive, in situ hybridization-positive, and both, respectively. Scale bar, 20 μm.
See also Tables S1 and S2.
Cell Reports  , DOI: ( /j.celrep )
Copyright © 2017 The Authors Terms and Conditions

5. Figure 3
Optogenetic Stimulation of vBF SOM Neurons Also Increases Sucrose Intake
(A and B) Schematic diagrams showing experimental setup (A) and time line of the sucrose-preference test (B).
(C) Photo-stimulation of SOM neurons significantly increases probability of sucrose licks in vBF-SOM-ChR2 mice (p = , experimental group, 92.74% ± 1.67%, n = 7, control group, 80.31% ± 1.8%, n = 7).
(D) Photo-stimulation of SOM neurons significantly increases probability of sucrose lick duration (p = , experimental group, 92.46% ± 1.14%, n = 7, control group, 77.55% ± 3.28%, n = 7).
(E) Photo-stimulation of SOM neurons significantly increases probability of sucrose intake (p = , experimental group, 84.71% ± 1.26%, n = 7, control group, 71.74% ± 1.66%, n = 7).
(F) Schematic diagram showing time line of feeding tests after high-fat diet.
(G and H) Photo-stimulation of SOM neurons has no significant effect on high-fat chow intake (G) and standard chow intake (H) after HFD (high-fat chow: p = 0.9847, experimental group, ± g, n = 6, control group, ± g, n = 5; standard chow: p = , experimental group, ± g, control group, ± g).
Data represented as mean ± SEM. All significance values were tested by one-way ANOVA and post hoc Sidak’s test.
Cell Reports  , DOI: ( /j.celrep )
Copyright © 2017 The Authors Terms and Conditions

6. Figure 4
Optogenetic Stimulation of SOM Neuron Induces Anxiety-like Behaviors
(A) VP-VGAT-ChR2 mice spend significantly more time in the photo-stimulation-paired side (p < experimental group, 86% ± 2.24%, n = 5; control group, 46% ± 5.81%, n = 5).
(B) vBF-SOM-ChR2 mice spend significantly less time in the photo-stimulation-paired chamber (p = experimental group, 39% ± 4.55%, n = 9; control group, 51% ± 1.89%, n = 7).
(C) Photo-stimulation of VP PV neurons has no effect on real-time place preference test (p = experimental group, 46% ± 7.6%, n = 5; control group, 53% ± 4.02%, n = 5).
(D) Representative animal tracks of a vBF-SOM-ChR2 mouse for the first two epochs of the elevated-plus maze test. Left: schematic diagram showing the time line of elevated plus maze test.
(E) Activation of vBF SOM neurons significantly decreases the time spent in the open arm (p = , experimental group, ± s, n = 9, control group, ± s, n = 7).
(F) vBF SOM-ChR2 mice show a significantly lower percentage of open arm entry from the center (p = , experimental group, % ± %, n = 9, control group, % ± %, n = 7).
(G) Representative animal tracks of a vBF-SOM-ChR2 mouse in the open field test. Left: schematic diagram showing the time line of open field test.
(H) Photo-stimulation of SOM neurons significantly decreases the time spent in center (p = experimental group, ± s, n = 9; Control group, ± s, n = 7).
(I) Photo-stimulation of SOM neurons has no effects on velocity (p = experimental group, ±  cm/s, n = 9; control group, ±  cm/s, n = 7).
Data represented as mean ± SEM. All significance values were tested by one-way ANOVA and post hoc Sidak’s test.
See also Figure S5.
Cell Reports  , DOI: ( /j.celrep )
Copyright © 2017 The Authors Terms and Conditions

7. Figure 5
SOM and VGAT Neurons Receive Different Inputs but Target Similar Downstream Regions
(A) Schematic diagram showing the design of the recombinant AAV-DIO-ChR2 (H134R)-eYFP virus and injection site to anterogradely trace the outputs of VP VGAT neurons and vBF SOM neurons.
(B) VP VGAT neurons and vBF SOM neurons both send projections to LHA, LHb, and VTA. Scale bar, 200 μm.
(C) Virus used in the cell-type-specific retrograde trans-synaptic tracing.
(D) Schematic diagrams showing the experimental design for cell-type-specific retrograde trans-synaptic tracing. AAV-DIO-EGFP-TVA and AAV-DIO-RG virus were injected on day 1 to mark the starter neurons in VGAT-Cre or SOM-Cre mice and SADΔG-dsRed (Enva) virus were injected on day 14 to retrogradely trace the inputs.
(E and F) Images from a VGAT-ires-Cre mouse showing AAV-DIO-EGFP-TVA expression in VP VGAT neurons (E) and SADΔG-dsRed (Enva) labeling of input neurons in nucleus accumbens and lateral septum (F). Green, EGFP-TVA; orange, SADΔG-dsRed; blue, DAPI. Scale bars, 200 μm (E), 500 μm (D left), 200 μm (D right).
(G and H) Images from a SOM-Cre mouse showing AAV-DIO-EGFP-TVA expression in vBF SOM neuron (G) and SADΔG-dsRed (Enva) labeling of input neurons in nucleus accumbens and lateral septum (H). Green, EGFP-TVA; orange, SADΔG-dsRed; blue, DAPI. Scale bar, 200 μm (G), 500 μm (H left), 200 μm (H right).
(I) Numbers of starter neuron in each SOM-Cre and VGAT-Cre mouse.
(J) The ratio of input neurons in NAc and total starter neurons in each mouse.
(K) The ratio of input neurons in LS and total starter neurons in each mouse.
(L) Significantly more NAc neurons innervate VP VGAT neurons and more LS neurons innervate vBF SOM neurons. NAc input ratio: p = (VP-VGAT,  ± , n = 4; vBF-SOM, ± , n = 4). LS input ratio: p = (VP-VGAT, ± , n = 4; vBF-SOM, ± , n = 4).
LHA, lateral hypothalamus area; LHb, lateral habenula; VTA, ventral tegmental area; AcbC, nucleus accumbens core; AcbSh, nucleus accumbens shell; LAcbSh, lateral nucleus accumbens shell; VDB, nucleus of the vertical limb of the diagonal band; LS, lateral septum; Tu, olfactory tubercle. MS, medial septal nucleus; SFI, septofimbrial nucleus. Data represented as mean ± SEM. All significance values were tested by one-way ANOVA and post hoc Sidak’s test. Panels (E) and (G): brain outlines adapted from Figure 30 of The Mouse Brain in Stereotaxic Coordinates, Compact 3rd Edition, Keith Franklin and George Paxinos (© copyright Elsevier, All Rights Reserved).
See also Table S3.
Cell Reports  , DOI: ( /j.celrep )
Copyright © 2017 The Authors Terms and Conditions

8. Figure 6
The Projection from vBF SOM Neurons to LHA Is Responsible for Fat Intake Only, but Not Sucrose Preference
(A and B) ChR2-eYFP expression in vBF SOM neurons (A) and their axonal projections in the LHA (B). Scale bars, 200 μm (up), 20 μm (bottom).
(C) Photo-stimulation of SOM axons in LHA significantly increases high-fat chow intake (p = , experimental group, ± g, n = 7; control group, ± g, n = 7).
(D) Photo-stimulation of SOM axons in LHA has no effects on standard chow intake (p = , experimental group, ± g, n = 7; control group, ± g, n = 7).
(E–G) Photo-stimulation of SOM axons in LHA has no effects on sucrose lick number (E), sucrose lick duration (F), and sucrose preference (G) (sucrose lick number: p > , experimental group, 85.37% ± 3.16%, n = 6; control group, 85.17% ± 5.36%, n = 6; sucrose lick duration: p = , experimental group, 84.25% ± 3.87%, n = 6; control group, 84.75% ± 4.08%, n = 6; sucrose preference: p = , experimental group, 77.82% ± 3.47%, n = 6; control group, 79.15% ± 7.21%, n = 6).
Upper panel: brain outlines adapted from Figure 30 (left) and Figure 45 (right) of The Mouse Brain in Stereotaxic Coordinates, Compact 3rd Edition, Keith Franklin and George Paxinos (© copyright Elsevier, All Rights Reserved). Data represented as mean ± SEM. All significance values were tested by one-way ANOVA and post hoc Sidak’s test.
Cell Reports  , DOI: ( /j.celrep )
Copyright © 2017 The Authors Terms and Conditions