Time-dependent behavioral, neurochemical, and metabolic dysregulation in female C57BL/6 mice caused by chronic high-fat diet intake

Abstract

High-fat diet (HFD) induced obesity is associated not only with metabolic dysregulation, e.g., impaired glucose homeostasis and insulin sensitivity, but also with neurological dysfunction manifested with aberrant behavior and/or neurotransmitter imbalance. Most studies have examined HFD's effects predominantly in male subjects, either in the periphery or on the brain, in isolation and after a finite feeding period. In this study, we evaluated the time-course of selected metabolic, behavioral, and neurochemical effects of HFD intake in parallel and at multiple time points in female (C57BL/6) mice. Peripheral effects were evaluated at three feeding intervals (short: 5-6 weeks, long: 20-22 weeks, and prolonged: 33-36 weeks). Central effects were evaluated only after long and prolonged feeding durations; we have previously reported those effects after the short (5-6 weeks) feeding duration. Ongoing HFD feeding resulted in an obese phenotype characterized by increased visceral adiposity and, after prolonged HFD intake, an increase in liver and kidney weights. Peripherally, 5 weeks of HFD intake was sufficient to impair glucose tolerance significantly, with the deleterious effects of HFD being greater with prolonged intake. Similarly, 5 weeks of HFD consumption was sufficient to impair insulin sensitivity. However, sensitivity to insulin after prolonged HFD intake was not different between control, low-fat diet (LFD) and HFD-fed mice, most likely due to age-dependent decrease in insulin sensitivity in the LFD-fed mice. HFD intake also induced bi-phasic hepatic inflammation and it increased gut permeability. Behaviorally, prolonged intake of HFD caused mice to be hypoactive and bury fewer marbles in a marble burying task; the latter was associated with significantly impaired hippocampal serotonin homeostasis. Cognitive (short-term recognition memory) function of mice was unaffected by chronic HFD feeding. Considering our prior findings of short-term (5-6 weeks) HFD-induced central (hyperactivity/anxiety and altered ventral hippocampal neurochemistry) effects and our current results, it seems that in female mice some metabolic/inflammatory dysregulations caused by HFD, such as gut permeability, appear early and persist, whereas others, such as glucose intolerance, are exaggerated with continuous HFD feeding; behaviorally, prolonged HFD consumption mainly affects locomotor activity and anxiety-like responses, likely due to the advanced obesity phenotype; neurochemically, the serotonergic system appears to be most sensitive to continued HFD feeding.

Keywords: Anxiety; Glucose intolerance; Hepatic inflammation; High-fat diet; Hypoactivity; Insulin sensitivity.

Figure 1

Detailed outline of the study's experimental design. For Abbreviations: see the text.

Figure 2

Effect of low-fat diet (LFD) or high-fat diet (HFD) consumption on mean body weights of female C57BL/6 mice after 6, 22, or 36 weeks on respective diets. Graphical representations are mean ± SEM (n = 8/group/time point). *** P ≤ 0.001 indicates significant difference from LFD within a time point. “a, b, c” indicate significant difference between time points within a dietary treatment (P ≤ 0.001).

Figure 3

Absolute (g) organ weights of female C57BL/6 mice fed either a high fat diet (HFD) or a control, low fat diet (LFD) for 6 (A), 22 (B), or 36 (C) weeks. Graphical representations are mean ± SEM (n = 8/group/time point). ** P ≤ 0.01 represent effects within a time point.

Figure 4

Glucose and test (GTT; 2 g/kg BW, oral; A, B, and C) and insulin sensitivity test (IST; 0.5 IU, i.p.: A', B', and C') of female C57BL/6 mice fed either a high fat diet (HFD) or a control (low fat diet; LFD) for 6 (A, A'), 22 (B, B'), or 36 (C, C') weeks. Integrated areas under the curve (AUC) of GTT and IST responses are presented, respectively, in Fig. 4D and Fig. 4D'. Graphical representations are mean ± SEM (n = 8/group/time point). * P ≤ 0.05, ** P ≤ 0.01 and *** P ≤ 0.001 represent effects within a time point (min, A, A', B, B', C and C'; weeks, D and D'). Note: for simplicity, only the differences between LFD and HFD groups within a time point are represented in A, A', B, B', C, and C' panels. “a” indicates significant difference between time points within treatment (P ≤ 0.001).

Figure 5

Effect of low-fat diet (LFD) or high-fat diet (HFD) consumption on gastrointestinal permeability (plasma FITC-dextran levels) of female C57BL/6 mice after 6, 22 or 36 weeks on respective diets. Graphical representations are mean ± SEM (n = 8/group/time point). * P ≤ 0.05 represents treatment effect within feeding duration. “a” indicates significant difference between time points within a dietary treatment.

Figure 6

Effect of 6, 22 or 36 weeks of high-fat diet (HFD) consumption by female C57BL/6 mice on liver mRNA levels of tumor-necrosis factor alpha (TNFα), interleukin 6 (IL-6) and haptoglobin (Hp). The house keeping gene (HKG) 18S was used to normalize the mRNA data, which are presented as fold change relative to respective low-fat diet (LFD) group at each time point. Graphical representations are means ± SEM (n = 6–8 per group). * P < 0.05, ** P < 0.01 indicate treatment effect within a feeding duration.

Figure 7

Effect of 21 or 32 weeks of high-fat diet (HFD) consumption on: (A) distance traveled (per 5 min interval); (B) time spent per 5 min interval in the center of the open field arena during open field testing; (C) average time to turn during the pole test; and (D) mean forelimb grip strength. Graphical representations are mean ± SEM (n = 8/group/time point). * P ≤ 0.05, ** P ≤ 0.01 and *** P ≤ 0.001 indicates significant difference from low-fat diet (LFD) within a time point. “a” indicates significant difference between time points within a dietary treatment (P ≤ 0.05).

Figure 8

Effect of 21 or 32 weeks of high-fat diet (HFD) consumption on: (A) time spent with a familiar vs. a novel object (%) in a novel object recognition test (NOR); (B) total time spent swimming in a forced swim test (FST); and (C) number of marbles buried (≥ 70%) during a marble burying test (MBT). Graphical representations are mean ± SEM (n = 8/group/time point). “#” indicates significant (P ≤ 0.05) difference between the novel vs. familiar object within a dietary treatment group. ** P ≤ 0.01 indicates significant difference from low-fat diet (LFD) within a time point. “a” indicates significant difference between time points within a dietary treatment (P ≤ 0.001).

Figure 9

Effect of 22 or 36 weeks of high-fat diet (HFD) consumption on concentrations of serotonin (5-HT; A, C and E) and its metabolite, 5-hydroxyindole acetic acid (5-HIAA; B, D and F) in the prefrontal cortex (PFC; A and B), dorsal hippocampus (dHIP, C and D) and ventral hippocampus (vHIP, E and F) of female C57BL/6 mice. 5-HT and 5-HIAA concentrations are normalized on a per mg protein basis and are presented as mean ± SEM (n = 8/group/time point). *P ≤ 0.05 indicates significant difference from low-fat diet (LFD) within a time point. (^) indicates trend towards significance (P = 0.06).