Figure 1.
12-hr IAM produced concentration-dependent fructose bingeing.
A: Schematic diagram showing the timeline of the daily light cycle and water, fructose and chow access during the IAM. Bingeing was defined as a statistically significant increase in the 1st hr (dashed red box) fructose or chow intake between day 1 and subsequent diet days. B: Time course of 1st hr additional bottle intake throughout IAM for the four treatment groups: control (a second novel water bottle), 4% fructose, 8% fructose and 12% fructose. ∧ Denotes the day at which stable fructose bingeing was achieved in the 8 and 12% fructose groups. C: 1st hr total liquid intake of both the ad libitum water bottle (white columns) and the additional bottle that contained either water (diagonal striped column), or 4%, 8%, or 12% fructose solution (progressively darker blue columns) on the first and last day of the IAM. D: 1st hr chow intake for the four groups throughout IAM. ∧, # Denotes the day at which stable chow bingeing was achieved in control and 4% fructose groups, respectively. E: 1st hr caloric intake represented as chow (horizontal striped column) and fructose (blue column) calories on the first and last day of IAM. Mean values ± SEM (n = 8/group). *p<0.05 within group 1-way RMANOVA compared to day 1 with Tukey’s post hoc analysis.
Table 1.
Long-term fructose bingeing did not alter D1R or D2R number.
Figure 2.
Both long- and short-term IAM produced similar levels of bingeing behavior.
A: 1st hr water (black symbols; control) or fructose (blue symbols; 8% fructose) intake on days 1 and 2 for the short-term IAM experiment (closed symbols) and on days 1, 2, 8 and 21 for the long-term IAM experiment (open symbols). B: 1st hr chow intake on days 1 and 2 of short-term IAM and days 1, 2, 6, and 21 of long-term IAM. ∧ denotes the day at which stable fructose or chow bingeing was achieved in the long-term IAM experiment. *p<0.05 paired t-test short-term IAM only. Mean values ± SEM (n = 8/group).
Figure 3.
Long-term fructose bingeing reduced NAc shell neuron and enhanced LH/PeF Orx neuron activation.
Immunohistochemical (IHC) staining results following long-term IAM (see Fig. 2). A: Anatomic outlines of NAc shell and core with enlarged representative images of IHC staining in NAc shell from control (Ctrl.) and 8% fructose (Fruc.) group animals with Fos-IR in green and Nissl stain in blue. B: Group results for NAc shell Fos-IR. C: Anatomic outlines of LH/PeF and DMH with enlarged representative images of IHC staining in Ctrl. and Fruc. group animals. The Fos-IR nuclei are in green, Orx-IR cells are in fuchsia and Nissl stain is in blue. D: Group results for the ratio of Orx neuron activation (Fos-IR nuclei in Orx-IR cells) in the LH/PeF. E: Significant negative correlation between LH/PeF Orx neuron activation and NAc shell neuron activation in the fructose IAM group. F: No correlation in the control IAM group. Dotted bands represent 95% confidence intervals of the linear regression. Mean values ± SEM (n = 7–8 per group) *p<0.05 unpaired t-test. Abbreviations: anterior commissure (ac), lateral ventricle (LV), fornix (f).
Table 2.
Fos-IR in hypothalamic and reward-related regions.
Table 3.
Fos-IR in Orx-IR neurons after long- or short-term IAM.
Figure 4.
Ox1R antagonist pretreatment reduced caloric intake in both IAM feeding groups.
A: 1st hr total liquid intake for ad libitum water (white column) and additional bottle, containing water (diagonal striped) or 8% fructose (blue column), for control and fructose groups on day 21 of IAM (no injection) and on day 22 of IAM after a 30-min pretreatment with vehicle (2% DMSO and 10% β-cyclodextrin in sterile water; Veh.) or SB-334867 (30 mg/kg, i.p; SB) B: The 1st hr caloric intake, expressed as chow (horizontal striped columns) and 8% fructose (in blue columns) calories. Red lines indicate day 1 intake of water or fructose in panel A and calories in panel B. Mean values ± SEM (n = 6–9/group) *p<0.05 within group 1-way RMANOVA compared to day 21.
Figure 5.
Ox1R antagonist pretreatment reduced feeding-induced neuronal activation only in the control IAM group.
A: Orx neuron Fos-IR in LH/PeF, B: Fos-IR in the NAc shell, and C: Fos-IR in VMH following vehicle (Veh.) or SB-334867 (SB) pretreatment and 1st hr bingeing on day 22 of IAM (see Fig. 4 for treatment details). Mean values ± SEM (n = 6–9 animals/group) *p<0.05 within feeding group 1-way ANOVA.