High fat diet blunts stress-induced hypophagia and activation of Glp1r dorsal lateral septum neurons in male but not in female mice

Figures

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  Figure 1. Restraint Stress Hypophagia in Lean Male Mice but Not in Obese Male or Lean or Obese Female Mice. A) Body weight of chow-fed and HFD-fed male and female mice on the day of restraint. B-D; I–K) Food intake following no restraint (Control; black lines/bars) or restraint stress (Restraint; red lines/bars) which occurred for 1 h prior to dark onset in lean male (B–D) and female (I–K) wild-type mice. E-H) Meal patterns (meal number, intermeal interval, meal duration, and meal size) in lean (solid bars) and obese (hatched bars) male mice. ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001, ∗∗∗∗P < 0.0001 in a t-test (A) or Two-way ANOVA (B–K). Data are presented as Mean ± SEM. N = 9–12/group.

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  Figure 2. Metabolic Readings Following Restraint Stress or No Restraint Stress in Lean and Obese Male and Female Mice. EE in chow-fed (A) and HFD-fed (B) control (black line/bar) and restrained (red line/bar) male mice during 1 h control or restraint stress period (yellow highlight) and the subsequent 12 h. C) Average EE during the control and restraint period. RER in chow-fed (D) and HFD-fed (E) control (black line/bar) and restrained (red line/bar) male mice during 1 h control or restraint stress period (yellow highlight) and the subsequent 12 h. F) Average RER during the control and restraint period. Locomotor activity in chow-fed (G) or HFD-fed (H) male mice following no restraint (Control; black) or restraint stress (Restraint; red) 1 h prior to dark onset. EE in chow-fed (I) and HFD-fed (J) control (black line/bar) and restrained (red line/bar) female mice during 1 h control or restraint stress period (yellow highlight) and the subsequent 12 h. K) Average EE during the control and restraint period. RER in chow-fed (L) and HFD-fed (M) control (black line/bar) and restrained (red line/bar) female during 1 h control or restraint stress period (yellow highlight) and the subsequent 12 h.…

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  Figure 3. Restraint Stress Increases dLS Glp1r Neuronal Activity in Lean Male Mice but Not in Obese Male or Lean or Obese Female Mice. A) Representative fluorescent image of unamplified GCaMP expression (green), native tdTomato in Glp1r+ neurons (red), and co-localization (yellow, highlighted with white arrows) in the dLS. Of 159 red cells (Glp1r+) counted, 146 were yellow, yielding an efficiency of GCaMP expression in Glp1r+ neurons of 91.82%. Of 162 green cells (GCaMP+), 16 were not yellow (Glp1r−), yielding non-selective expression of 9.88%. Hashed lines outline the fiber-optic cannula placement. LV = lateral ventricle. Scale bar = 200 μm. Fluorescent signal, a proxy for neuronal activity, in a lean (B, C) and an obese (D, E) male mouse during no restraint (B, D) and restraint (C, E). Orange line and initial spike in fluorescent signal occurred when mouse was lifted by the tail to simulate putting the mouse in a restrainer (No Restraint [B, D]) or to put the mouse in a restrainer (Restraint C, E). The mLspike algorithm, which is a validated way to impute spike frequency based on Ca2+ transients, was used to measure spike (red hatch marks) frequency in MATLAB and generated the ‘best…

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  Figure 4. Struggle Behaviors Show Lean and Obese mice Struggled Similarly and dLS Glp1r Neurons are Activated during Struggle in Lean Male Mice compared to Obese Male Mice, whereas Neuronal Responses in Lean and Obese Female Mice do Not Differ during Struggle. Struggle analysis showed no significant differences between dietary conditions within sex suggesting that chow-fed and HFD-fed mice experienced the restraint stress similarly. Average length of each bout (A), number of bouts per animals (B), and total time in bout (C) were analyzed. Male (D) and female (E) fiber photometry recordings time-locked to the struggle analysis. Maximum peaks were calculated for the 5s after bout onset to normalize across bouts and time-locked to the fiber photometry recordings via MATLAB. Movement bout begins at zero seconds. Shaded area is SEM. (F) Average AUC by group. ∗∗P < 0.01; ∗∗∗P < 0.001. (N = 6–14/group).

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  Figure 5. Inhibiting dLS Glp1r Neurons Blunts Stress-Induced Hypophagia. Food intake in male Glp1r-Cre; tdTomato mice expressing inhibitory DREADD in the dLS following saline (A) or CNO injection (B; 2 mg/kg, i. p.) 30 min before either restraint or no restraint 1 h before dark onset. ∗∗∗P < 0.0001 in a Two-way ANOVA. Data are presented as Mean ± SEM. N = 12–13/group. C–H) Fluorescent images of dLS, outlined in hatched lines, in saline-injected (C–E) and CNO-injected (F–H) mice showing cFos in control (C, F) and restraint stressed (D, G) mice and HA tag confirming inhibitory DREADD expression (E, H). I-R) Higher magnification representative fluorescent images of dLS showing native tdTomato fluorescence (red), cFos (cyan), and merge (magenta) in saline-treated control (I–K), saline-treated restraint stressed (L–N), and CNO-treated restraint stressed (O–Q) Glp1r-Cre; tdTomato mice as well as cFos, native tdTomato fluorescence, and HA tag (i.e., DREADD; green) in a restraint stressed, CNO-treated mouse (R). S) cFos quantification of control and restrained mice given either saline or CNO (2 mg/kg, i. p.) 90 min before perfusion. ∗P < 0.05, ∗∗∗P < 0.001. Data are presented as Mean ± SEM. N =…