Effects of fasting on maximum thermogenesis in temperature-acclimated rats

To further investigate the limiting effect of substrates on maximum thermogenesis in acute cold exposure, the present study examined the prevalence of this effect at different thermogenic capabilities consequent to cold- or warm-acclimation. Male Sprague-Dawley rats (n=11) were acclimated to 6, 16 and 26C, in succession, their thermogenic capabilities after each acclimation temperature were measured under helium-oxygen (21% oxygen, balance helium) at –10C after overnight fasting or feeding. Regardless of feeding conditions, both maximum and total heat production were significantly greater in 6>16>26C-acclimated conditions. In the fed state, the total heat production was significantly greater than that in the fasted state at all acclimating temperatures but the maximum thermogenesis was significant greater only in the 6 and 16C-acclimated states. The results indicate that the limiting effect of substrates on maximum and total thermogenesis is independent of the magnitude of thermogenic capability, suggesting a substrate-dependent component in restricting the effective expression of existing aerobic metabolic capability even under severe stress.     

Chronic mitochondrial uncoupling treatment prevents acute cold-induced oxidative stress in birds

Endotherms have evolved two major types of thermogenesis that allow them to actively produce heat in response to cold exposure, either through muscular activity (i.e. shivering thermogenesis) or through futile electro-chemical cycles (i.e. non-shivering thermogenesis). Amongst the latter, mitochondrial uncoupling is of key importance because it is suggested to drive heat production at a low cost in terms of oxidative stress. While this has been experimentally shown in mammals, the oxidative stress consequences of cold exposure and mitochondrial uncoupling are clearly less understood in the other class of endotherms, the birds. We compared metabolic and oxidative stress responses of zebra finches chronically treated with or without a chemical mitochondrial uncoupler (2,4-dinitrophenol: DNP), undergoing an acute (24 h) and a chronic (4 weeks) cold exposure (12 °C). We predicted that control birds should present at least a transient elevation of oxidative stress levels in response to cold exposure. This oxidative stress cost should be more pronounced in control birds than in DNP-treated birds, due to their lower basal uncoupling state. Despite similar increase in metabolism, control birds presented elevated levels of DNA oxidative damage in response to acute (but not chronic) cold exposure, while DNP-treated birds did not. Plasma antioxidant capacity decreased overall in response to chronic cold exposure. These results show that acute cold exposure increases oxidative stress in birds. However, uncoupling mitochondrial functioning appears as a putative compensatory mechanism preventing cold-induced oxidative stress. This result confirms previous observations in mice and underlines non-shivering thermogenesis as a putative key mechanism for endotherms in mounting a response to cold at a low oxidative cost.     

Improving cold tolerance in elderly rats by aminophylline

During severe cold exposure, old rats (23-26 months) were less capable in maintaining normal body temperature as compared to young rats (6-9 months) due to lower rate of heat production (HP). Single injection of optimal doses of aminophylline (AMPY; 10 and 18.7 mg/kg, i.p.), a phosphodiesterase inhibitor which enhances the intracellular cyclic AMP concentration, significantly increased the rate of HP in old rats to levels beyond the control values observed in young rats. Consequently, cold tolerance of the old rats was significantly improved. This AMPY-improved cold tolerance is apparently not due to increased non-shivering thermogenesis (NST) since AMPY failed to enhance norepinephrine-stimulated NST in the old rats. It is likely that AMPY increased substrate mobilization and/or conversion, thereby circumventing the limiting role of substrate availability for shivering thermogenesis. Thus, the age-dependent decrease in cold tolerance may be due to a reduced capacity for substrate mobilization when challenged by cold.     

Effects of feeding on aminophylline induced supra-maximal thermogenesis

Single injection of aminophylline (a phosphodiesterase inhibitor which prolongs cyclic AMP actions) elicited “supra-maximal thermogenesis” in cold exposed rats. The increases were 19.4, 16 and 15%, respectively, above their saline-injected self-control maxima in the overnight fasted, rationed, and ad libitum fed states. However, in the fasted state the increased thermogenesis could not be sustained. Feeding of a 5 ml substrate mixture containing carbohydrate, protein and fat (12.56 kJ/ml) prior to aminophylline injection elicited increases of 29.6, 16 and 13.7%, respectively, for the three feeding regimens and a sustained “supra-maximal thermogenesis” in the fasted state. Since substrate feeding modulated aminophylline stimulated thermogenesis only in the fasted state when endogenous substrate reserves are diminished, it is indicative that both the magnitude and duration of aminophylline induced thermogenesis are substrate dependent.     

Rates of energy substrates utilization during human cold exposure

Although it is well established in animals that acute cold exposure markedly increases the oxidation of energy substrates, the absolute quality and quantity of substrate oxidation is poorly understood in humans. This study compared the rates of substrate utilization in seven healthy young men exposed to both the warm (control exposure at 29 degrees C; semi-nude, 14 h fasted) and to the cold for 2 h (10 degrees C, 1 m.s-1 wind velocity). Substrate utilization was calculated using indirect calorimetry and the nonprotein respiratory exchange ratio, which was derived from the urinary urea nitrogen output. Cold exposure induced a 3.1 +/- 0.2 degrees C drop in mean body temperature and a body heat debt of 825.9 +/- 63.3 kJ (p less than 0.01). These parameters remained essentially unchanged in the warm. Cold exposure elevated the 2 h energy expenditure 2.46-fold in comparison to the warm (p less than 0.01). This cold-induced thermogenesis was accompanied by increases of 588% in carbohydrate oxidation (p less than 0.01) and 63% in fat oxidation (p less than 0.05), whereas protein oxidation remained unchanged. Although the greatest proportion of the energy expenditure in the warm was derived from lipid (59%), carbohydrate oxidation represented the major fuel for thermogenesis in the cold, since it accounted for 51% of the corresponding total energy expenditure. The results demonstrate that cold exposure causes a much greater increase in the utilization of carbohydrate than lipid. It is suggested that these substrates are directly utilized for thermogenesis in the shivering skeletal muscles.     

Effects of fasting and aminophylline on norepinephrine-stimulated non-shivering thermogenesis

In an attempt to further elucidate the mechanisms of fasting-depressed maximum thermogenesis and cold tolerance, norepinephrine (NE)-stimulated non-shivering thermogenesis (NST) in cold-acclimated rats was used as a functional index of possible alterations in adrenergic efficacy after fasting. Fasting decreased the magnitude of maximum NE-Stimulated NST by 18.2% [6.87±0.47 Kcal (Kg^.75.min)^?1 well-fed vs. 5.81±0.39 Kcal (Kg^.75.min)^?1 fasted], but the apparent adrenergic binding affinity was not affected [Ke=0.43 μg NE min^?1 well-fed vs 0.55 μg NE min^?1 fasted]. Pretreatment with aminophylline [15 mg Kg^?1, i.p.], a phosphodiesterase inhibitor, restored the fasting-depressed NE-stimulated NST to the fed level. The results suggest that the depression of maximum thermogenesis after fasting is not due to changes in adrenergic binding characteristics but to alteration in cAMP production/degradation, resulting in decreased substrate mobilization for thermogenesis.     

Effect of food restriction on cold adaptability of rats

To determine the role of the nutritional state in nonshivering thermogenesis during cold adaptation, cold adaptability was compared between cold-adapted (5 degrees C for 4-5 weeks) rats fed ad libitum and cold-adapted rats pair fed with warm controls having the same food intake. Cold-adapted pair-fed rats suffered a significant loss in body weight during cold exposure. However, brown adipose tissue (BAT) in both cold-adapted ad libitum fed and cold-adapted pair-fed rats was enlarged to the same extent as compared with that in control rats. Fat-free dry matter in BAT also increased in cold-adapted ad libitum fed and cold-adapted pair-fed rats to the same extent. Cold tolerance as assessed by the change in the colonic temperature at -5 degrees C was improved relative to control rats and was the same for cold-adapted ad libitum fed and cold-adapted pair-fed rats. Nonshivering thermogenesis as estimated by the noradrenaline-induced increase in oxygen consumption was significantly greater in the cold-exposed rats and there was no significant difference between cold-adapted ad libitum fed and cold-adapted pair-fed rats. These results suggest that an improved cold tolerance by means of nonshivering thermogenesis in brown adipose tissue is closely related to the low temperature itself but not the increased food intake which occurred in the cold.     

Mechanisms underlying the supra-maximal thermogenesis elicited by aminophylline in rats

Previous studies showed that acute treatment with aminophylline (AMPY) significantly elevated maximum thermogenesis and improved cold tolerance in rats and man in severe cold. However, the exact mechanism by which AMPY enhances thermogenesis was unknown. Rats receiving enprofylline (ENPRO) (1.5 and 15 mg/kg, i.p.), a selective phosphodiesterase inhibitor, failed to show enhanced thermogenesis. In contrast, treatment with a selective adenosine receptor antagonist, 8-phenyltheophylline(8-PT; 2.5 to 10 mg/kg, i.p.), significantly increased (p less than 0.05) thermogenesis and cold tolerance. However, the maximal thermogenic effect by optimal dose of 8-PT (5 mg/kg) was significantly lower than that with optimal dose of AMPY (18.7 mg/kg, i.p.); the deficit could be eradicated by combining optimal 8-PT dose with a low dose of AMPY (1.25 mg/kg), but not with ENPRO. These results indicate that the thermogenic effect of AMPY is not by inhibition of phosphodiesterase but at least partially by antagonism of adenosine receptors. It is also apparent that older mechanisms in addition to adenosine antagonism are also involved in AMPY's thermogenic action.     

Effect of acute cold exposure on the expression of the adiponectin, resistin and leptin genes in rat white and brown adipose tissues.

Leptin, adiponectin, and resistin are key hormones produced by adipose tissue. In the present study, we have examined the effects of acute cold exposure (18 h at 6 degrees C) on the expression of the genes encoding these hormones in both brown and white fat of rats. Acute cold exposure resulted in a significant (p < 0.001) increase in the level of UCP1 and metallothionein-1 mRNAs in brown adipose tissue, indicative of an activation of thermogenesis. Leptin mRNA was decreased (p < 0.001) in brown fat in the cold, and there was also a small but statistically significant (p < 0.05) decrease in adiponectin mRNA; resistin mRNA did not change significantly (p > 0.05). In white fat, the level of leptin mRNA also fell in the cold (p < 0.05), but there was no significant change (p > 0.05) in either adiponectin or resistin mRNA. The serum concentration of adiponectin was unchanged following acute cold exposure. We conclude that while leptin gene expression is inhibited by exposure to cold, there is no major effect on the expression of either the adiponectin or resistin genes in white or brown fat despite the cold-induced stimulation of sympathetic activity and fatty acid flux. Thus, adiponectin and resistin are unlikely to play a key role in the extensive metabolic adaptations to cold.     

Effect of cold-acclimation on oxygen uptake and glucose production of perfused duckling liver

The control of hepatic metabolism by substrates and hormones was assessed in perfused liver from young Muscovy ducklings. Studies were performed in fed or 24-h fasted 5-week-old thermoneutral (25 degrees C; TN) or cold-acclimated ducklings (4 degrees C; CA) and results were compared with those obtained in rats. Basal oxygen uptake of perfused liver (LVO2) was higher after cold acclimation both in fed (+65%) and 24-h fasted (+29%) ducklings and in 24-h fasted rats (+34%). Lactate (2 mM), the main gluconeogenic substrate in birds, similarly increased LVO2 in both TN and CA ducklings and the effect was larger after fasting. Both glucagon and norepinephrine dose-dependently increased LVO2 in ducklings and rats, but cold acclimation did not improve liver response and liver sensitivity to norepinephrine in ducklings was even reduced in CA animals. Liver contribution to glucagon-induced thermogenesis in vivo was estimated to be 22% in TN and 12% in CA ducklings. Glucagon stimulated gluconeogenesis from lactate in duckling liver and the stimulation was 2.2-fold higher in CA than in TN fasted birds. These results indicate a stimulated hepatic oxidative metabolism in CA ducklings but hepatic glucagon-induced thermogenesis (as measured by LVO2) was not improved. A role of the liver is suggested in duckling metabolic acclimation to cold through an enhanced hepatic gluconeogenesis under glucagon control.     

Thermogenetic mechanisms involved in Man's fitness to resist cold exposure

People whose evolution has taken place in contrasting climates, appear to have an almost identical critical temperature (27–29°C). This does not exclude the existence of minor variations within and between populations with regard to individual fitness to resist cold exposure. The main factor by which Man's fitness to resist cold exposure can be varied, appears to be thermogenesis. The biological variation of BMR, shivering, a possible non-shivering thermogenesis, and the maximal aerobic power is reviewed. BMR of an individual can vary with diet, general health, habitual physical activity, as well as various environmental conditions, conceivably including cold exposure. A definite inter- and intra-individual variation in shivering threshold exists. The underlying mechanism of these differences is poorly understood, however. Firm evidence in favour of a non-shivering thermogenesis in Man is lacking. Man's capability to raise his metabolism in muscular exercise is an important part of his fitness to resist cold exposure. This capability can be assessed by measurement of maximal oxygen uptake. Maximal oxygen uptake is influenced by age, sex, health, diet and habitual physical activity. It is questionable if evolution in contrasting climates brings about variation in maximal oxygen uptake.     

Acute and chronic cold exposure differentially affects the browning of porcine white adipose tissue

Piglets are characteristically cold intolerant and thus susceptible to high mortality. However, browning of white adipose tissue (WAT) can induce non-shivering thermogenesis as a potential strategy to facilitate the animal’s response to cold. Whether cold exposure can induce browning of subcutaneous WAT (sWAT) in piglets in a similar manner as it can in humans remains largely unknown. In this study, piglets were exposed to acute cold (4°C, 10 h) or chronic cold exposure (8°C, 15 days), and the genes and proteins of uncoupling protein 1 (UCP1)-dependent and independent thermogenesis, mitochondrial biogenesis, lipogenic and lipolytic processes were analysed. Interestingly, acute cold exposure induced browning of porcine sWAT, smaller adipocytes and the upregulated expression of UCP1, PGC1α, PGC1β, C/EBPβ, Cidea, UCP3, CKMT1 and PM20D1. Conversely, chronic cold exposure impaired the browning process, reduced mitochondrial numbers and the expression of browning markers, including UCP1, PGC1α and PRDM16. The present study demonstrated that acute cold exposure (but not chronic cold exposure) induces porcine sWAT browning. Thus, browning of porcine sWAT could be a novel strategy to balance the body temperature of piglets, and thus could be protective against cold exposure.     

Npvf: Hypothalamic Biomarker of Ambient Temperature Independent of Nutritional Status

The mechanism by which mice, exposed to the cold, mobilize endogenous or exogenous fuel sources for heat production is unknown. To address this issue we carried out experiments using 3 models of obesity in mice: C57BL/6J+/+ (wild-type B6) mice with variable susceptibility to obesity in response to being fed a high-fat diet (HFD), B6. Ucp1-/- mice with variable diet-induced obesity (DIO) and a deficiency in brown fat thermogenesis and B6. Lep-/- with defects in thermogenesis, fat mobilization and hyperphagia. Mice were exposed to the cold and monitored for changes in food intake and body composition to determine their energy balance phenotype. Upon cold exposure wild-type B6 and Ucp1-/- mice with diet-induced obesity burned endogenous fat in direct proportion to their fat reserves and changes in food intake were inversely related to fat mass, whereas leptin-deficient and lean wild-type B6 mice fed a chow diet depended on increased food intake to fuel thermogenesis. Analysis of gene expression in the hypothalamus to uncover a central regulatory mechanism revealed suppression of the Npvf gene in a manner that depends on the reduced ambient temperature and degree of exposure to the cold, but not on adiposity, leptin levels, food intake or functional brown fat.     

Absence of adaptive nonshivering thermogenesis in a marsupial,the fat-tailed dunnart (<Emphasis Type="Italic">Sminthopsis crassicaudata</Emphasis>)

The presence of nonshivering thermogenesis in marsupials is controversially debated. Survival of small eutherian species in cold environments is crucially dependent on uncoupling protein 1 (UCP1)-mediated, adaptive nonshivering thermogenesis that is executed in brown adipose tissue. In a small dasyurid marsupial species, the fat-tailed dunnart (Sminthopsis crassicaudata), an orthologue of UCP1 has been recently identified which is upregulated during cold exposure resembling adaptive molecular adjustments of eutherian brown adipose tissue. Here, we tested for a thermogenic function of marsupial brown adipose tissue and UCP1 by evaluating the capacity of nonshivering thermogenesis in cold-acclimated dunnarts. In response to an optimal dosage of noradrenaline, cold-acclimated dunnarts (12°C) showed no additional recruitment of noradrenaline-induced maximal thermogenic capacity in comparison to warm-acclimated dunnarts (24°C). While no differences in body temperature were observed between the acclimation groups, basal metabolic rate was significantly elevated after cold acclimation. Therefore, we suggest that adaptive nonshivering thermogenesis does not occur in this marsupial species despite the cold recruitment of oxidative capacity and UCP1 in the interscapular fat deposit. In conclusion, the ancient UCP orthologue in marsupials does not contribute to the classical nonshivering thermogenesis, and may exhibit a different physiological role.     

Lower critical temperature and cold-induced thermogenesis of lean and overweight humans are inversely related to body mass and basal metabolic rate

It is colloquially stated that body size plays a role in the human response to cold, but the magnitude and details of this interaction are unclear. To explore the inherent influence of body size on cold-exposed metabolism, we investigated the relation between body composition and resting metabolic rate in humans at thermoneutrality and during cooling within the nonshivering thermogenesis range. Body composition and resting energy expenditure were measured in 20 lean and 20 overweight men at thermoneutrality and during individualized cold exposure. Metabolic rates as a function of ambient temperature were investigated considering the variability in body mass and composition. We observed an inverse relationship between body size and the lower critical temperature (LCT), i.e. the threshold where thermoneutrality ends and cold activates thermogenesis. LCT was higher in lean than overweight subjects (22.1 ± 0.6 vs 19.5 ± 0.5 °C, p < 0.001). Below LCT, minimum conductance was identical between lean and overweight (100 ± 4 vs 97 ± 3 kcal/°C/day respectively, p = 0.45). Overweight individuals had higher basal metabolic rate (BMR) explained mostly by the higher lean mass, and lower cold-induced thermogenesis (CIT) per degree of cold exposure. Below thermoneutrality, energy expenditure did not scale to lean body mass. Overweight subjects had lower heat loss per body surface area (44.7 ± 1.3 vs 54.7 ± 2.3 kcal/°C/m²/day, p < 0.001). We conclude that larger body sizes possessed reduced LCT as explained by higher BMR related to more lean mass rather than a change in whole-body conductance. Thus, larger individuals with higher lean mass need to be exposed to colder temperatures to activate CIT, not because of increased insulation, but because of a higher basal heat generation. Our study suggests that the distinct effects of body size and composition on energy expenditure should be taken in account when exploring the metabolism of humans exposed to cold.     

Cold‐Induced Adaptive Thermogenesis in Lean and Obese

On entering a cold environment, people react by increasing insulation and energy expenditure (EE). However, large interindividual differences exist in the relative contribution of each mechanism. Short‐term studies revealed that obese subjects increase EE (i.e., adaptive thermogenesis) less than lean subjects, which might have implications for the predisposition to obesity. In this study, we validate the differences in adaptive thermogenesis between lean and obese upon midterm mild cold exposure. Therefore, 10 lean and 10 obese subjects were exposed for 48 h to mild cold (16 °C) in a respiration chamber. The preceding 36 h they stayed in the same chamber at a neutral temperature (22 °C) for the baseline measurements. EE, physical activity, skin temperature, and core temperature have been measured for the last 24 h of both parts. Mean daytime EE increased significantly in the lean subjects (P < 0.01), but not in the obese. Physical activity decreased significantly in the lean (P < 0.01) and the obese (P < 0.001) subjects. The change in EE was related to the change in physical activity in both groups (respectively R² = 0.673, P < 0.01 and R² = 0.454, P < 0.05). Upon mild cold exposure, lean subjects decreased proximal skin temperature less, but distal skin temperature more than obese. In conclusion, the interindividual differences in cold‐induced thermogenesis were related to changes in physical activity in both lean and obese, pointing at the existence of individual variation in physical activity to compensate for cold‐induced thermogenesis. Furthermore, although a large part of the lean subjects counteracted the cold by increasing EE, most obese subjects changed temperature distribution, and therefore, increased insulation.     

Enhancement of maximal thermogenesis by reducing endogenous adenosine activity in the rat

Adenosine has been shown in vitro to be a potent antilipolytic agent and an inhibitor of insulin-stimulated glucose utilization in skeletal muscle. To test whether endogenously produced adenosine (e.g., from ATP hydrolysis) shares these deleterious effects on substrate mobilization and utilization and thus limits maximum thermogenesis in vivo, adenosine deaminase (converts adenosine to inosine) was given to rats 15 min before cold exposure. Significant (P less than 0.05) increases in thermogenesis were observed under both well-fed (100 units/kg ip) and food-rationed (200 units/kg ip) states. Significant (P less than 0.05) increases in thermogenesis and cold resistance were also observed after pretreatment with selective adenosine receptor antagonists [8-cyclopentyltheophylline (1 microgram/kg ip) greater than 1,3-dipropyl-8-p-sulfophenylxanthine (1.25 mg/kg ip) greater than aminophylline (18.7 mg/kg ip)], indicating an A1-receptor-mediated effect. These results indicate that endogenously released adenosine can indeed attenuate the thermogenic capacity in severe cold and that adenosine antagonists, especially those selective for A1-receptor, are useful in improving cold resistance under varying nutritional states.     

Changes in plasma cortisol during acute cold exposure in euthermic European hedgehogs: thermoregulatory role of the hypothalamo-pituitary-adrenal axis

Summary During acute cold exposure regulatory heat production of European hedgehogs was significantly increased with a simultaneous rise in the plasma cortisol level. Soon after cold exposure, at the time when standard metabolic rate was measured, the plasma cortisol level was reduced again. This finding indicates a highly reactive hypothalamo-pituitary-adrenal axis in euthermic hedgehogs exposed to a cold environment.Two effects of the adrenocorticostatic agent metopirone ditartrate were observed: (1) A metopirone-induced increase in nonshivering thermogenesis (NST) starting 18 min after application, reaching a maximum after 43 min and disappearing after 90 min (Fig. 1, Table 1). Simultaneously a profound long lasting hyperglycemia was observed (Fig. 2). (2) In contrast to untreated hedgehogs, animals which were given metopirone did not show an increase in the plasma cortisol level, when exposed to cold within 2 to 3 h (Fig. 6).When animals were pretreated with dexamethasone the metopirone-induced NST was suppressed, indicating that the effect of metopirone is mediated by the hypothalamo-pituitary-adrenal system (Fig. 4).The effect of metopirone at doses used in this study was reversible.It is suggested that a combined action of corticosteroids and catecholamines is involved in the control of regulatory heat production.Abbreviations SMR standard metabolic rate - NST nonshivering thermogenesis Supported by the Deutsche Forschungsgemeinschaft Wu 63/5     

Human skeletal muscle mitochondrial uncoupling is associated with cold induced adaptive thermogenesis

Background

Mild cold exposure and overfeeding are known to elevate energy expenditure in mammals, including humans. This process is called adaptive thermogenesis. In small animals, adaptive thermogenesis is mainly caused by mitochondrial uncoupling in brown adipose tissue and regulated via the sympathetic nervous system. In humans, skeletal muscle is a candidate tissue, known to account for a large part of the epinephrine-induced increase in energy expenditure. However, mitochondrial uncoupling in skeletal muscle has not extensively been studied in relation to adaptive thermogenesis in humans. Therefore we hypothesized that cold-induced adaptive thermogenesis in humans is accompanied by an increase in mitochondrial uncoupling in skeletal muscle.

Methodology/Principal Findings

The metabolic response to mild cold exposure in 11 lean, male subjects was measured in a respiration chamber at baseline and mild cold exposure. Skeletal muscle mitochondrial uncoupling (state 4) was measured in muscle biopsies taken at the end of the respiration chamber stays. Mild cold exposure caused a significant increase in 24h energy expenditure of 2.8% (0.32 MJ/day, range of −0.21 to 1.66 MJ/day, p<0.05). The individual increases in energy expenditure correlated to state 4 respiration (p<0.02, R² = 0.50).

Conclusions/Significance

This study for the first time shows that in humans, skeletal muscle has the intrinsic capacity for cold induced adaptive thermogenesis via mitochondrial uncoupling under physiological conditions. This opens possibilities for mitochondrial uncoupling as an alternative therapeutic target in the treatment of obesity.