Sleep/wake fragmentation disrupts metabolism in a mouse model of narcolepsy

Abstract

Recent population studies have identified important interrelationships between sleep duration and body weight regulation. The hypothalamic hypocretin/orexin neuropeptide system is able to influence each of these. Disruption of the hypocretin system, such as occurs in narcolepsy, leads to a disruption of sleep and is often associated with increased body mass index. We examined the potential interrelationship between the hypocretin system, metabolism and sleep by measuring locomotion, feeding, drinking, body temperature, sleep/wake and energy metabolism in a mouse model of narcolepsy (ataxin-ablation of hypocretin-expressing neurons). We found that locomotion, feeding, drinking and energy expenditure were significantly reduced in the narcoleptic mice. These mice also exhibited severe sleep/wake fragmentation. Upon awakening, transgenic and control mice displayed a similar rate of increase in locomotion and food/water intake with time. A lack of long wake episodes partially or entirely explains observed differences in overall locomotion, feeding and drinking in these transgenic mice. Like other parameters, energy expenditure also rose and fell depending on the sleep/wake status. Unlike other parameters, however, energy expenditure in control mice increased upon awakening at a greater rate than in the narcoleptic mice. We conclude that the profound sleep/wake fragmentation is a leading cause of the reduced locomotion, feeding, drinking and energy expenditure in the narcoleptic mice under unperturbed conditions. We also identify an intrinsic role of the hypocretin system in energy expenditure that may not be dependent on sleep/wake regulation, locomotion, or food intake. This investigation illustrates the need for coordinated study of multiple phenotypes in mouse models with altered sleep/wake patterns.

Figure 1. Diurnal fluctuation of locomotor activity (LMA, A), core body temperature (Tb, B), and wakefulness (C) in wild-type (WT) and transgenic (TG) mice

Data shown are from different groups of mice. LMA was measured using infrared monitors. Two-way ANOVA indicates a significant difference in LMA between the WT and TG mice (n = 8 in each genotype), and no difference in mean Tb (n = 12 in each genotype) or wake time (n = 5 WT, 6 TG). The P-values from two-way ANOVA are presented above the curves. The dashed lines with open circles represent group averages of the WT mice and the continuous lines with filled circles represent group averages of the TG mice. Bars underneath indicate light and dark phases. *Significant difference (P < 0.05, unpaired, two-tailed t test) between the WT and TG mice at a specific time point. Data of 24 h are double plotted side by side to better illustrate the diurnal rhythms.

Figure 3. Diurnal fluctuation of energy expenditure (EE, A) and respiratory quotient (RQ, B) in wild-type (WT) and transgenic (TG) mice

Two-way ANOVA indicates a significant difference in EE between the WT and TG mice (n = 8 in each genotype), and no difference in RQ (n = 8 in each genotype). The P-values from two-way ANOVA are presented above the curves. The dashed lines with open circles represent group averages of the WT and the continuous lines with filled circles represent group averages of the TG. Bars underneath indicate light and dark phases. *Significant difference (P < 0.05, unpaired, two-tailed t test) between the WT and TG mice at a specific time point. Data are double plotted.

Figure 2. Histograms of wake (A) and NREM sleep (B) duration in wild-type (WT) and transgenic (TG) mice

For equal bin histograms (left), the wake data are binned in 4 min intervals, and the NREM sleep data are binned in 1 min intervals. The dashed lines with open circles represent group averages of the WT mice and the continuous lines with filled circles represent group averages of the TG mice. The distributions of wake and NREM sleep durations were significantly different between the two genotypes (P < 0.001, Kolmogorov–Smirnov test). The bar graphs (right) show the differences between the WT and TG mice in three arbitrary bins of state duration. The open bars represent the WT mice and the filled bars represent the TG mice. *Significant difference (P < 0.05, unpaired, two-tailed t test) between the WT and TG mice.

Figure 4. Energy expenditure during different states and transitions in wild-type (WT) and transgenic (TG) mice

A, hypnograms coupled with energy expenditure (EE) and core body temperature (Tb) measurements. One representative animal from each genotype is shown. EE and Tb are expressed in four colours with each colour representing the values at each of the four states: active wake (AW), quiet wake (QW), NREM and REM. B, average EE during AW, QW, NREM and REM. Note the differences among the states are underrepresented due to frequent state transition and measurement carry-over. Two-way ANOVA showed significant difference in overall EE between the WT and TG mice (P < 0.05, n = 5 for WT, n = 6 for TG). The open bars represent the WT mice and the filled bars represent the TG mice. Data are expressed as mean ±s.e.m.*Significant difference between AW or QW and NREM or REM in the WT mice; #significant difference between AW and NREM in the TG mice (P < 0.05, unpaired, two-tailed t test). C, z-score transform of EE at state transitions. The dashed lines with open circles represent the WT mice and the continuous lines with filled circles represent the TG mice. Data are selected to avoid using those from transient periods (e.g. < 60 s of REM). Numbers in parentheses represent the numbers of data points used in analyses: (WT, TG). Data are expressed as means ±s.d.

Figure 5. Progression of locomotor activity (LMA, A), feeding (B), and drinking (C) upon awakening in wild-type (WT) and transgenic (TG) mice

LMA was measured using telemetry. Two-way ANOVA shows no significant difference in LMA (P = 0.32), feeding (P = 0.96) or drinking (P = 0.97) between the WT and TG mice (n = 5 for WT, n = 6 for TG). The open bars represent the WT mice and the filled bars represent the TG mice. *Significant difference (P < 0.05, unpaired, two-tailed t test) between the WT and TG mice at a specific period.

Figure 6. Progression of core body temperature (Tb, A) and energy expenditure (EE, B) in wake and NREM sleep in wild-type (WT) and transgenic (TG) mice

Two-way ANOVA indicates no significant difference in Tb during wake (P = 0.78) and NREM sleep (P = 0.10) between the WT and TG mice (n = 5 for WT, n = 6 for TG). The EE values during both wake and NREM sleep were significantly lower in the TG than those in the WT mice. The rate change during wake was significantly lower in the TG mice (P < 0.05, ANCOVA for the first 3 min and beyond), and was similar during NREM sleep in both genotypes (P > 0.11, ANCOVA for data within the first 8 min). The open bars represent the WT mice and the filled bars represent the TG mice. *Significant difference (P < 0.05, unpaired, two-tailed t test) between the WT and TG mice at a specific period.