Conceived and designed the experiments: CE. Performed the experiments: CE. Analyzed the data: CE AS. Contributed reagents/materials/analysis tools: CE AS. Wrote the paper: CE AS.
The authors have declared that no competing interests exist.
Chronic sleep disruption in laboratory rats leads to increased energy expenditure, connective tissue abnormalities, and increased weights of major organs relative to body weight. Here we report on expanded findings and the extent to which abnormalities become long-lasting, potentially permanent changes to health status after apparent recuperation from chronic sleep disruption. Rats were exposed 6 times to long periods of disrupted sleep or control conditions during 10 weeks to produce adaptations and then were permitted nearly 4 months of undisturbed sleep. Measurements were made in tissues from these groups and in preserved tissue from the experimental and control groups of an antecedent study that lacked a lengthy recuperation period. Cycles of sleep restriction resulted in energy deficiency marked by a progressive course of hyperphagia and major (15%) weight loss. Analyses of tissue composition in chronically sleep-restricted rats indicated that protein and lipid amounts in internal organs were largely spared, while adipose tissue depots appeared depleted. This suggests high metabolic demands may have preserved the size of the vital organs relative to expectations of severe energy deficiency alone. Low plasma corticosterone and leptin concentrations appear to reflect low substrate availability and diminished adiposity. After nearly 4 months of recuperation, sleep-restricted rats were consuming 20% more food and 35% more water than did comparison control rats, despite normalized weight, normalized adipocytes, and elevated plasma leptin concentrations. Plasma cholesterol levels in recuperated sleep-restricted rats were diminished relative to those of controls. The chronically increased intake of nutriments and water, along with altered negative feedback regulation and substrate use, indicate that internal processes are modified long after a severe period of prolonged and insufficient sleep has ended.
Repeated exposure to a deficiency of a basic need, whether food, water, oxygen, or warmth, results in physiological adaptations and phenotypic changes—modifications that are not apparent after acute deficiencies (e.g.,
Inadequate sleep, in nature and in society, may be expected to be severe and recurrent during a given lifespan. We recently studied the effects of repeated exposure to 10-day periods of restricted sleep in rodents to study physiological adaptations that result from chronic limitations of sleep
Latent or long-lasting effects of insufficient sleep may exert injurious effects and lower resistance to disease, or otherwise alter proper development and healthy aging, in the same way that chronic restrictions of other requirements would be expected to impact health. The purpose of the present study, therefore, was to broaden understanding of the physiological characteristics that arise from chronic sleep deficiency and to determine if residual consequences remain after recuperation. To accomplish this, we analyzed the composition of preserved tissues from rats in the antecedent study of 6 cycles of sleep restriction or control conditions without extended recovery
Procedures were carried out in accordance with protocols for animal care and use approved by institutional animal care and use committees at The Medical College of Wisconsin and the Zablocki Veterans Administration Medical Center, project numbers 2560-02N and 2560-04. Subjects were adult male Sprague-Dawley rats obtained from Harlan (Madison, WI). Live animal experiments in the present study were composed of 16 rats that weighed 491 (SD 29) g and were 24 to 26 wk old at the start of the study. Preserved tissues from the antecedent study of sleep restriction without an extended recovery period were obtained from 20 rats that were 452 (SD 32) g and were 28 (SD 1) wk old at the time of study
Control and experimental animals were housed under constant light conditions in rooms with ordinary ceiling lights to minimize the effects sleep deprivation may have on the amplitude and phase of the circadian rhythm (see
Surgery was conducted to ensure equal treatment of rats under conditions of chronic sleep restriction or ambulation control conditions both with an extended recovery period and those of the antecedent study without an extended recovery period. Electrodes were implanted into the cranium and temporalis muscles using the procedures described for the antecedent investigation, during which electroencephalographic and electromyographic signals were obtained to determine sleep stages and wakefulness
Schematics of the experimental apparatus designed by Rechtschaffen, Bergmann, and colleagues are shown elsewhere
During a 7-day baseline period, the platform was rotated once per hour to acquaint the rats with the movement of the housing platform.
As in the antecedent study, sleep-restricted rats experienced a cycle consisting of a 10-day sleep restriction period and a 2-day
Besides greatly fragmented sleep in sleep-restricted rats, we previously showed that the total accumulation of sleep is reduced by this technique
The experimental conditions were matched in control animals and included surgery, daily measurement procedures, and the same total duration of ambulation requirements, except that the rotations were consolidated to permit lengthy opportunities to obtain uninterrupted sleep. The platform rotation schedule consisted of a 90-min period, during which the platform was rotated for 150 s and then was stationary for 30 s, followed by 198 min without platform rotations. This schedule was repeated 5 times per day. By these means, we have shown that NREM sleep is reduced from a baseline value of 55% of the time to between 46 and 48% of the time during a given 10-day ambulation period, while the percentage of time in paradoxical sleep does not differ from baseline. During the 2-day
Food and water intake and body weights were recorded daily. During the middle of both the 5th and 6th cycles, food and fecal waste were collected to measure caloric values, which were previously reported to rule out malabsorption and feeder waste as explanations for weight loss in rats examined in the antecedent study
Rats in the antecedent study did not experience an extended recovery period; for those rats, tissue harvesting and necropsy occurred, as described below, at the conclusion of the 6 cycles of sleep restriction or ambulation control conditions. Rats in the new live animal experiments in the present study were provided a 17- to 18-wk extended recovery period after the same 6 cycles (72 days) of sleep restriction or ambulation control conditions, at which time the rats were removed from the apparatuses and placed individually in cylindrical cages (12″ diam., 12″ high). These cages were equipped with the same food tubes, water bottles, and counterbalanced boom assemblies used in the experimental apparatuses and were placed within environmental chambers under conditions of constant light and 25°C ambient temperature. Food and water intake and body weights were measured each day during the first month of extended recovery, and then every 48 hrs until Day 115 in the sleep-restricted rats, when it appeared clear that the range of body weights of this group did not differ from those of the ambulation control group. The surgically implanted head plug assemblies became detached at various points during the study, as expected; the affected individuals remained under study because the effects were transient and not serious (e.g., temporary appearance of malaise and decreased food intake).
Procedures of necropsy evaluations and tissue harvests began on Days 125 and 122 for sleep-restricted and ambulation control rats, respectively, and continued over a 7-day period, between 0900 and 1500 hr. Each rat was injected with bromodeoxyuridine (0.5 mg/kg ip) under very brief isoflurane anesthesia 90 min prior to necropsy examination to allow for future studies of cell proliferation. After this 90-min period, each rat was anesthetized by isoflurane inhalation and injected with ketamine·HCl (50 mg/kg ip), xylazine·HCl (8 mg/kg im), and atropine sulfate (0.003 mg/kg im). Once deep anesthesia was attained, a cardiac puncture was performed for blood collection and exsanguination. Blood was injected into chilled Vacutainer® tubes containing EDTA K3, rotated, and transferred to microcentrifuge tubes for centrifugation at 10 000 g for 10 min at 4°C.
Tissues were rapidly dissected and preserved for the present analyses and for follow-up investigations in 1 or more of the following ways: 1) enclosed in aluminum foil, snap frozen in liquid nitrogen, and sealed in small plastic bags (Whirl Pak); 2) grossed, placed in cassettes, and fixed in 10% neutral buffered formalin or 4% paraformaldehyde (adipose tissue); and 3) embedded in freezing media and fixed frozen in chilled methylbutane. The site chosen for a specimen of non-weight-bearing skeletal muscle was the ventral abdominal wall, sampled just off midline, between the rib cage and the bladder. The small intestine was dissected at the pylorus and at the junction with the cecum, laid out straight for a length measurement, and washed thrice through with saline to remove contents before preservation. Plasma and snap frozen tissues were stored at −80°C.
Sections of paraffin-embedded adipose tissue from perirenal, omental, and epididymal sites in sleep-restricted and ambulation control rats with extended recovery were cut into 4-µm thick slices and stained with hematoxylin and eosin by the Children's Research Institute Histology Core, an affiliate of the Medical College of Wisconsin. Stained sections were coded to ensure that investigators were blind to each animal's experimental conditions. Morphometrics were performed under brightfield microscopy and digital image analysis (Olympus BX51 microscope and DP71 camera, ImagePro Plus image analysis software by Media Cybernetics, Inc., Bethesda, MD). The areas of unilocular adipocytes in each of 10 representative fields were measured. The total region sampled was approximately 0.4 mm2 and contained 161 to 408 adipocytes. The areas of multilocular adipocytes, identified as small, uniformly-sized lipid droplets encased within a single membrane (illustrated in
Rat plasma insulin and rat plasma corticosterone were measured by enzyme immunoassays (SPI-Bio, France, and Immunodiagnostic Systems Ltd, Fountain Hills, AZ, respectively). Rat plasma leptin was measured by radioimmunoassay (Millipore, St. Charles, MO). Determinations were made in duplicate. The intra-assay coefficients of variation in our hands were <4% for insulin and leptin and <10% for corticosterone. No values fell beyond the range of assay detection. The following clinical chemistry measurements were considered viable determinants in EDTA-treated plasma and were completed at the Research Animal Diagnostic Laboratory (University of Missouri, Columbia, MO): cholesterol, triglycerides, high- and low-density lipoproteins (HDL and LDL), glucose, albumin, total protein, blood urea nitrogen, creatinine, and phosphorus (Olympus AU680, Beckman-Coulter, Center Valley, PA). In addition, plasma osmolality, an important consideration in causes of thirst, was measured by the freeze-point method (Osmette, Precision Systems, Inc., Natick, MA), which is the most direct method and one that does not rely on electrolyte measurements.
Fresh frozen aliquots of liver, heart, intestine, skeletal muscle, kidney, and spleen were prepared for tissue composition analysis of fat, protein, moisture, and ash by conventional procedures (e.g.,
Values for food intake, water intake, and body weight in ambulation control and sleep-restricted rats during the six cycles and the subsequent 114 to 115 days of extended recovery, first were expressed relative to individual baseline values, then log transformed. Analyses were completed by mixed-effects models to account for different treatment conditions across time and individual variability, and to elucidate common trends. Selection of time-trend structures was guided by the Bayesian information criterion, which balances model complexity and goodness of fit. For body weight, the trend during the 6 sleep restriction cycles was modeled linearly with 1 change point, and the trend during the extended recovery period was modeled by an exponential return to an underlying linear growth rate. Both food and water intake were modeled by linear models for the separate components of sleep restriction, 2-day
Mixed-effects modeling was performed using the nlme 3.1–96 package of R 2.10.1. Other computations were performed in SAS 9.2 using PROC MULTTEST for the multiple comparison adjustments. Values are means (SD) if individual values composed an average and means (SE) if data first were averaged within animals and then averaged for the group.
Weight loss in sleep-restricted rats was progressive across cycles of sleep restriction and averaged 15 (SD 6)% below baseline levels by the last 2 days before the extended recovery period for recuperation. This was highly significantly different from the growth of ambulation control rats during the same period (
Data are expressed as a percentage change from baseline 2-day averages in sleep-restricted (▪) and ambulation control (○) rats. The first 72 days were arranged in 6 cycles, each composed of a 10-day period of sleep restriction or ambulation control (unshaded), followed by a 2-day period of
Food and water consumption during the periods of sleep restriction or ambulation control conditions before extended recovery replicated those of the antecedent study without extended recovery in overall dynamics. Peak daily food consumption based on 48-hr averages ranged from 179 to 495% of basal consumption in sleep-restricted rats of the present study. We detected initial increases in food intake at the start of both the sleep restriction and ambulation control conditions of 1.18- and 1.24-fold basal amounts, respectively, presumably due to scheduled ambulation. The positive slope of food intake during the sleep restriction periods was statistically significantly larger than that in the ambulation control group (
Water intake initially dropped in sleep-restricted rats by 0.58-fold (
One rat was removed from the experiment at the beginning of the 5th sleep restriction period because of a change in health status marked by lethargy, apparent weakness, and an inability to contend with platform rotations. This was the same clinical picture presented by each of 2 sleep-restricted rats that did not survive the 6th period during the antecedent study. Measurements on this animal just 24 hr prior to its removal from the formal study indicated robust food intake of 131% of baseline, reduced water intake to 47% of baseline, and loss of body weight from baseline of 12%; which should not have been lethal. Observations indicated that this rat was awake much of the time during recovery periods, when sleep was allowed
Dermatoses began to develop on the paws of all sleep-restricted rats of this study at about the same time and at the same sites as those on sleep-restricted rats of the antecedent study
Adipocytes in the omentum, epididymus, and the surrounds of the mesenteric lymph nodes in sleep-restricted rats with extended recovery did not differ in size from those of corresponding ambulation control rats. Only 1 multilocular region was found in the mesentery of 1 sleep-restricted rat with extended recovery.
Among rats that did not experience the extended recovery period, plasma corticosterone was significantly lower in sleep-restricted rats than in comparison ambulation control rats (
Horizontal bars indicate group means for each treatment condition. Significance of *
As previously reported for sleep-restricted rats without extended recovery, the liver, heart, kidney, and small intestine each accounted for a significantly greater proportion of body weight in sleep-restricted rats compared with ambulation control rats, and spleen weight was maintained
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Organ/Group | Organ Weight (g) |
Per Gram (%) | Weight in Milligrams | % Organ Mass | Per Gram (%) | Weight in Milligrams | % Lipid Free Mass | Per Gram (%) | Weight in Milligrams | % Lipid Free Mass | Per Gram (%) | Weight in Milligrams | % Lipid Free Mass |
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AC | 20.05 (2.99) | 26.3 (3.3) | 5317 (1213) | 26.3 (3.3) | 25.2 (2.6) | 5056 (951) | 34.3 (4.2) | 47.3 (5.0) | 9436 (1323) | 64.1 (4.2) | 1.2 (0.2) | 240.3 (36.9) | 1.7 (0.3) |
SR | 18.65 (2.55) | 20.5 (1.9)* | 3850 (816) | 20.5 (1.9)* | 25.8 (2.1) | 4815 (768) | 32.5 (2.4) | 52.5 (2.2) | 9759 (1136) | 66.1 (2.4) | 1.2 (0.1) | 221.9 (22.9) | 1.5 (0.1) |
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AC | 1.33 (0.10) | 4.1 (0.6) | 54.3 (10.0) | 4.1 (0.6) | 9.9 (0.9) | 131.9 (16.5) | 10.3 (0.9) | 84.6 (1.1) | 1128 (85) | 88.2 (1.0) | 1.5 (0.2) | 19.5 (3.6) | 1.5 (0.2) |
SR | 1.44 (0.15) | 3.4 (0.3) | 48.9 (6.1) | 3.4 (0.3) | 10.0 (1.0) | 144.6 (16.9) | 10.4 (1.0) | 84.6 (1.0) | 1222 (135) | 87.6 (0.9) | 2.0 (0.2)** | 28.4 (4.5)** | 2.0 (0.2)** |
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AC | 2.68 (0.18) | 5.0 (0.9) | 135.0 (26.1) | 5.0 (0.9) | 15.0 (0.6) | 402.9 (33.1) | 15.8 (0.6) | 78.2 (1.0) | 2095 (137) | 82.4 (0.8) | 1.7 (0.2) | 45.5 (6.2) | 1.8 (0.2) |
SR | 2.68 (0.35) | 5.1 (0.6) | 135.1 (18.3) | 5.1 (0.6) | 13.2 (1.2)*** | 352.9 (40.1) | 13.9 (1.3)** | 79.7 (1.3) | 2191 (271) | 84.0 (1.1)* | 2.0 (0.5) | 55.2 (17.1) | 2.1 (0.5) |
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AC | 1.34 (0.29) | 3.1 (0.6) | 40.8 (12.4) | 3.1 (0.6) | 14.1 (2.1) | 191.4 (59.0) | 14.5 (2.2) | 80.7 (2.2) | 1077 (219) | 83.3 (2.1) | 2.2 (0.3) | 28.8 (6.4) | 2.2 (0.3) |
SR | 1.27 (0.29) | 3.2 (0.6) | 42.6 (9.6) | 3.2 (0.6) | 12.8 (1.0) | 163.2 (37.6) | 13.2 (1.0) | 81.5 (1.0) | 1032 (240) | 84.2 (0.9) | 2.5 (0.3)* | 32.2 (8.7) | 2.6 (0.3)* |
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AC | 31.08 (4.45) | 26.4 (2.8) | 8229 (1528) | 26.4 (2.8) | 24.1 (1.8) | 7290 (728) | 32.1 (1.9) | 48.4 (3.2) | 15216 (2736) | 66.4 (1.8) | 1.1 (0.1) | 340.6 (55.2) | 1.5 (0.1) |
SR | 28.35 (4.91) | 25.3 (6.2) | 7600 (3046) | 25.3 (6.2) | 26.6 (2.7) | 7560 (839) | 36.5 (1.7) | 47.1 (4.9) | 12906 (1732) | 62.1 (1.7) | 1.0 (0.1) | 286.0 (33.4) | 1.4 (0.0) |
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AC | 1.90 (0.20) | 5.0 (2.4) | 80.0 (14.7) | 5.0 (2.4) | 10.0 (0.9) | 189.0 (22.7) | 10.4 (1.0) | 83.2 (2.4) | 1592 (158) | 87.5 (0.9) | 1.9 (0.4) | 35.9 (5.5) | 2.0 (0.3) |
SR | 2.04 (0.40) | 5.5 (2.5) | 106.6 (64.8) | 5.5 (2.5) | 9.2 (0.5) | 184.3 (39.8) | 9.5 (0.8) | 83.9 (2.8) | 1724 (311) | 88.7 (0.7) | 1.4 (0.3)* | 29.2 (7.3) | 1.5 (0.5)* |
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AC | 3.31 (0.33) | 4.9 (0.9) | 168.7 (20.7) | 4.9 (0.9) | 14.4 (0.9) | 488.3 (52.4) | 15.5 (0.8) | 79.1 (1.8) | 2599 (286) | 82.7 (0.7) | 1.6 (0.5) | 56.0 (13.3) | 1.7 (0.1) |
SR | 2.94 (0.20) | 5.4 (1.3) | 139.5 (17.0) | 5.4 (1.3) | 13.8 (0.8) | 421.8 (35.6) | 15.1 (0.3) | 79.3 (1.1) | 2333 (146) | 83.3 (0.2) | 1.5 (0.1) | 45.6 (2.2) | 1.6 (0.1) |
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AC | 1.46 (0.57) | 4.4 (1.1) | 61.4 (21.1) | 4.4 (1.1) | 13.6 (1.1) | 200.4 (80.8) | 14.3 (1.1) | 79.8 (1.1) | 1161 (456) | 83.3 (1.1) | 2.3 (0.1) | 34.1 (13.5) | 2.4 (0.1) |
SR | 1.53 (0.49) | 3.7 (1.4) | 57.0 (26.8) | 3.7 (1.4) | 13.0 (1.9) | 202.1 (78.6) | 13.6 (2.1) | 81.0 (2.3) | 1229 (387) | 83.9 (2.1) | 2.3 (0.2) | 36.6 (13.4) | 2.4 (0.2) |
Values are means (SD).
N = 7–10;
N = 6–8, except for N = 3–5 for AC and SR hearts and SR kidney weights and associated calculations of g/tissue and % organ mass. Statistical comparisons are provided for AC and SR rats without extended recovery, and AC and SR rats with extended recovery: *
Reported for SR and AC rats without extended recovery (10).
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Organ/Group | N | Per Gram (%) | Per Gram (%) | Lipid Free Mass (%) | Per Gram (%) | Lipid Free Mass (%) | Per Gram (%) | Lipid Free Mass (%) |
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AC | 9 | 11.3 (2.0) | 12.4 (5.3) | 14.0 (3.7) | 74.5 (4.0) | 84.2 (1.2) | 1.6 (0.2) | 1.8 (0.2) |
SR | 8 | 10.3 (3.2) | 9.2 (1.1) |
10.3 (1.6) |
78.9 (4.3) | 87.9 (1.9) |
1.6 (0.3) | 1.8 (0.3) |
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AC | 10 | 9.6 (3.6) | 19.4 (4.0) | 21.4 (4.0) | 69.6 (3.7) | 77.1 (4.0) | 1.4 (0.3) | 1.5 (0.4) |
SR | 7 |
6.4 (3.4) | 17.0 (5.2) | 18.2 (4.2) | 75.1 (5.8) | 79.5 (4.6) | 1.4 (0.2) | 1.4 (0.2) |
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AC | 8 | 12.7 (3.3) | 10.4 (1.4) | 12.0 (1.9) | 75.4 (4.2) | 86.4 (1.9) | 1.4 (0.2) | 1.6 (0.2) |
SR | 7 | 15.9 (4.1) | 8.1 (1.2) | 9.6 (1.2) | 74.4 (3.5) | 88.4 (1.2) | 1.7 (0.2) |
2.0 (0.2) |
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AC | 8 | 12.5 (5.2) | 22.4 (5.0) | 25.7 (5.9) | 64.0 (6.7) | 73.1 (5.9) | 1.1 (0.2) | 1.3 (0.2) |
SR | 7 | 13.9 (5.1) | 18.2 (2.1) | 21.2 (2.8) | 66.9 (5.6) | 77.6 (2.7) | 1.1 (0.1) | 1.2 (0.1) |
Values are means (SD);
N = 5 for % lipid free mass;
The first phase of the present study replicated the failed growth and extraordinary food and water consumption observed during repeated exposure to sleep restriction without a period of extended recovery
Sleep that is disrupted and fragmented is the most plausible explanation for the metabolic effects observed. The Bergmann-Rechtschaffen paradigm has been repeatedly validated for the selective deprivation of sleep and minimization of potential confounding factors (reviewed in
Thirst became progressively abnormal after initial increases in intake during the 2-day
Changes to viscera resulting from sleep restriction and its associated energy deficit are not like those resulting from caloric restriction. The changes reported for internal organs of chronically sleep-restricted rats all appeared to be related to high energy production and demand
The results indicate that lipid was diminished only in the liver by 22% and, therefore, lipid amount did not appear to be overly or selectively drawn from the vital organs. This implies the preservation of lipids as local messengers and substrate sources (reviewed in
The undiminished organ masses in these hypercatabolic sleep-restricted rats are expected to reflect tissue-specific metabolic rates and energy demands at the cellular level (reviewed in
Plasma leptin concentrations in sleep-restricted rats without extended recovery were consistent with a large body of empirical evidence indicating that leptin is low during energy deficiency and is positively related to the amount of body fat, which appeared exhausted in these animals. In sleep-restricted rats with extended recovery, elevated leptin was associated with a persistent elevation of food intake by 20% throughout the nearly 4-month observation period, despite catch-up growth and normal adipocyte morphometrics. This is abnormal because elevated leptin is expected to suppress appetite, except in individuals with insensitivity to leptin effector functions
Cholesterol levels in sleep-restricted rats were opposite to those expected during an energy deficit produced by calorie restriction; food restriction leads to an increase in circulating cholesterol
Corticosterone concentrations in sleep-restricted rats without extended recovery were opposite to the 5-fold elevations reported for caloric deficiency produced by food restriction
Humans whose sleep is restricted for more than 24 hours show signs consistent with those of laboratory rats, suggesting comparable underlying factors and evolving adaptations and maladaptations. For example, citations of hunger in sleep-restricted humans are fairly abundant
In light of these many similarities, hypermetabolism and weight loss in sleep-restricted laboratory rodents seem at odds with results from human studies that have pointed to glucose intolerance and insulin insensitivity as consequences of sleep restriction that would promote obesity and pose a risk for type II diabetes (see Refs.
To conclude, the present outcomes point to dynamic and fundamental physiological adjustments in response to repeated exposure to inadequate sleep. Peripheral organs and systems are largely ignored by most recent approaches to the study of sleep, which typically are brain-centered and infer that the brain is the sole recipient of benefits conferred by sleep
The outcomes of the present studies show that recuperation from chronic sleep restriction takes a long time and that some of the physiological adaptations and potential maladaptations that arise in response to repeated exposure to sleep loss are long lasting. A period of extended recovery lasting nearly 4 months after a long bout of repeated exposure to limited sleep produced a return of an overall healthy-appearing countenance that was belied by signs of imbalance. Prominent among these signs were elevated food and water intake indicative of elevated metabolism; elevated leptin, which has many effector functions; and signs of altered substrate demands.
The present evidence prompts us to speculate that sleep enables or facilitates certain cellular processes to take place efficiently or effectively during relative immobilization because, without normal sleep, metabolic imbalance and pathological outcomes result. This speculation has its origins in long-standing ideas and theories about sleep
We express our appreciation to Nichole Nellessen, Nicholas Kampa, Aletha Champine, and Bennett Hiner for technical assistance during live animal experimentation and tissue harvesting; Anne Folley and Christopher Henchen for carrying out biochemical, hormonal, and morphometric analyses; and Thomas Gardiner for the creation of a computerized program of master platform rotations to produce reduced and fragmented sleep or control conditions in rats. Facilities were provided by the Medical College of Wisconsin and the Department of Veterans Affairs.
A portion of this work was presented at a joint meeting of the American Academy of Sleep Medicine and the Sleep Research Society in 2010.