Leanness of Lou/C rats does not require higher thermogenic capacity of brown adipose tissue

Lou/C rats, an inbred strain of Wistar origin, remain lean throughout life and therefore represent a remarkable model of obesity resistance. To date, the exact mechanisms responsible for the leanness of Lou/C rats remain unknown. The aim of the present study was to investigate whether the leanness of Lou/C rats relies on increased thermogenic capacities in brown adipose tissue (BAT).Results showed that although daily energy expenditure was higher in Lou/C than in Wistar rats, BAT thermogenic capacity was not enhanced in Lou/C rats kept at thermoneutrality as demonstrated by reduced thermogenic response to norepinephrine in vivo, similar oxidative activity of BAT isolated mitochondria in vitro, similar levels of UCP1 mRNA and lower abundance of UCP1 protein in interscapular BAT depots. Relative abundance of β₃-adrenergic receptor mRNA was lower in Lou/C BAT while that of GLUT4, FABP or CPT1 was not altered. Activity-related energy expenditure was however considerably increased at thermoneutrality as Lou/C rats demonstrated an impressively high spontaneous running activity in voluntary running wheels. Prolonged cold-exposure (4 °C) depressed the spontaneous running activity of Lou/C rats while BAT thermogenic capacity was increased as reflected by rises in BAT mass, oxidative activity and UCP1 expression.It is concluded that the leanness of Lou/C rats cannot be ascribed to higher thermogenic capacity of brown fat but rather to, at least in part, increased locomotor activity. BAT is not deficient in this rat strain as it can be stimulated by cold exposure when locomotor activity is reduced suggesting some substitution between these thermogenic processes.     

Sex-associated differences in cold-induced UCP1 synthesis in rodent brown adipose tissue

 The effects of acute and chronic acclimation to cold on uncoupling protein 1 (UCP1) levels, as well as on GDP-binding to mitochondria, cytochrome c oxidase activity and mitochondrial protein concentration in brown adipose tissue (BAT) of intact male and female rats have been analyzed. Results reveal that females rats are more sensitive to cold because their threshold temperature for the thermogenic response is set at a higher value (around 22°C) than that of males (around 18°C), hence leading to differences in BAT UCP1 levels between the sexes at different environmental temperatures. In vitro experiments showed that steroid hormones, β-estradiol, estrone and progesterone, can reduce norepinephrine-induced UCP1 synthesis in brown adipocytes differentiated in primary culture. Thus the different sex-associated response of cold-induced thermogenesis in rats does not appear to be explained by a direct action of sex steroids upon the adipocyte, implying that other factors in the thermogenic regulatory system must be involved. Received: 23 March 1998 / Received after revision: 20 May 1998 / Accepted: 21 May 1998     

Synergic effect of overweight and cold on uncoupling proteins expression,a role of alpha(2)/beta(3) adrenergic receptor balance?

The effect of cold exposure, being overweight and their interaction was investigated on the response of uncoupling proteins UCP1, UCP2 and UCP3 and the alpha(2)/beta(3) adrenergic receptor (AR) balance in brown adipose tissue (BAT), as well as the involvement of leptin gene expression in white adipose tissues, in control and overweight male rats of the dietary obesity model known as the post-cafeteria model. UCP1, UCP2 and UCP3 mRNAs were up-regulated by cold, with a synergic effect of cold exposure and being overweight on UCP1 mRNA levels (with the related UCP1 protein response), and with UCP2 mRNA showing a parallel response. Furthermore, the BAT alpha(2)/beta(3) AR ratio was diminished in overweight rats. The results suggest that the UCP1-dependent thermogenic capacity in BAT of post-cafeteria overweight rats has a more sensitive response to cold exposure and that UCP2 and UCP3 could be somehow involved in the thermogenic response but differentially regulated. Moreover, the diminished alpha(2)/beta(3) AR ratio in BAT could be one of the factors involved in the more sensitive response of overweight rats to cold in terms of BAT thermogenesis-related parameters.     

Brown adipose tissue response to cafeteria diet-feeding involves induction of the UCP2 gene and is impaired in female rats as compared to males

Noradrenaline-dependent brown adipose tissue (BAT) thermogenesis is activated by the cold and excess energy intake, largely depends on the activity of the uncoupling protein 1 (UCP1), and is mediated mainly through the beta3-adrenoceptor (beta3-AR). We investigated the expression of ucp2, a gene that encodes a putative UCP1-like uncoupling protein, along with that of ucp1 and beta3-ar, in the interscapular BAT (IBAT) of male and female rats chronically fed a cafeteria diet. After 3 months on this diet, male rats attained a 34% excess body mass and showed IBAT hypertrophy and increased IBAT thermogenic potential, in terms of both UCP1 and UCP2 mRNA expression (both by 1.6-fold), UCP1 protein expression (by 1.75-fold) and GDP binding to IBAT mitochondria (by 2.2-fold); female rats attained a larger excess body weight (50%) and their IBAT, although hypertrophied, showed no signs of increased thermogenic potential per gram of tissue. Interestingly, the IBAT of female rats was already activated compared to males. Treatment of mouse brown adipocytes in primary culture with noradrenaline also triggered a dose-dependent increase of the levels of UCP1 mRNA and UCP2 mRNA. Retroregulatory down-regulation of the beta3-AR mRNA levels was found in the two models used. The results support a physiological role for UCP2, along with UCP1, in rodent BAT thermogenesis.     

Foetal and lactational exposure to alcohol increases oxidative capacity of brown adipose tissue in the rat. A possible relationship to cot death

The effect was studied of chronic alcohol intake in the rat during pregnancy and lactation on the brown adipose tissue (BAT) in pups. The idea was to find a possible relationship to cot death since in some cot death victims increased amounts of BAT have been observed. Exposure to ethanol increased the relative weight of the brown adipose tissue in pups and enhanced both its total protein content and the activities of the oxidative enzymes, succinate dehydrogenase and cytochrome oxidase. In the BAT of pups sympathetic activity, as demonstrated by noradrenaline, was also increased by long-term exposure to alcohol. In theory, an increased thermogenic capacity of the BAT in the newborn together with other factors such as emotional stress and infections could lead to death from hyperthermia, in which case only non-specific morphological signs would be found in the cadaver.     

Foetal and lactational exposure to alcohol increases oxidative capacity of brown adipose tissue in the rat. A possible relationship to cot death.

The effect was studied of chronic alcohol intake in the rat during pregnancy and lactation on the brown adipose tissue (BAT) in pups. The idea was to find a possible relationship to cot death since in some cot death victims increased amounts of BAT have been observed. Exposure to ethanol increased the relative weight of the brown adipose tissue in pups and enhanced both its total protein content and the activities of the oxidative enzymes, succinate dehydrogenase and cytochrome oxidase. In the BAT of pups sympathetic activity, as demonstrated by noradrenaline, was also increased by long-term exposure to alcohol. In theory, an increased thermogenic capacity of the BAT in the newborn together with other factors such as emotional stress and infections could lead to death from hyperthermia, in which case only non-specific morphological signs would be found in the cadaver.     

Repeated immobilization stress increases uncoupling protein 1 expression and activity in Wistar rats

Repeat immobilization-stressed rats are leaner and have improved cold tolerance due to enhancement of brown adipose tissue (BAT) thermogenesis. This process likely involves stress-induced sympathetic nervous system activation and adrenocortical hormone release, which dynamically enhances and suppresses uncoupling protein 1 (UCP1) function, respectively. To investigate whether repeated immobilization influences UCP1 thermogenic properties, we assessed UCP1 mRNA, protein expression, and activity (GDP binding) in BAT from immobilization-naive or repeatedly immobilized rats (3 h daily for 4 weeks) and sham operated or adrenalectomized (ADX) rats. UCP1 properties were assessed before (basal) and after exposure to 3 h of acute immobilization. Basal levels of GDP binding and UCP1 expression was significantly increased (140 and 140%) in the repeated immobilized group. Acute immobilization increased GDP binding in both naive (180%) and repeated immobilized groups (220%) without changing UCP1 expression. In ADX rats, basal GDP binding and UCP1 gene expression significantly increased (140 and 110%), and acute immobilization induced further increase. These data demonstrate that repeated immobilization resulted in enhanced UCP1 function, suggesting that enhanced BAT thermogenesis contributes to lower body weight gain through excess energy loss and an improved ability to maintain body temperature during cold exposure.     

Sexual dimorphism in the adrenergic control of rat brown adipose tissue response to overfeeding

Gender-related differences in the brown adipose tissue (BAT) response to overfeeding rats on a cafeteria diet were studied by assessing the balance between the expression of beta-adrenoceptors (beta1-, beta2-, beta3-AR) and alpha2A-AR and their relation to the expression of uncoupling proteins (UCP1, UCP2, UCP3). Cafeteria diet feeding for 15 days, which involved a similar degree of hyperphagia in both sexes, led to a greater body weight excess in females than in males and a lower activation of thermogenesis. Gender-related differences were found for different adrenoceptor expression and protein levels, which might explain, in part, sex differences in the thermogenic parameters. The lower expression of alpha2A-AR in females than in males could be responsible for the higher expression of UCP1 and thermogenic capacity under non-hyperphagic conditions. However, in a situation of high adrenergic stimulation--as occurs with overfeeding--as there is a preferential recruitment of the beta3-AR by noradrenaline compared with other adrenergic receptors, the higher levels of beta3-AR in males rats than in females could be responsible for the greater thermogenic capacity and the lesser weight gain in males. Thus, the alpha2/beta3 balance in BAT could be a key in the thermogenic control.     

Short- and long-term influences of hypoxia during early postnatal period of development on behavioral and hormonal responses in rats

Hypoxia is a common neonatal stress that leads to essential long-lasting complications in the brain development. The aim of this study was to determine short- and long-term effects of early postnatal hypoxia (on day 7) on depression- and pain-related behaviors and the plasma corticosterone levels. The plasma corticosterone levels increased after 3-h severe hypoxia in 7-day-old rat pups (hypoxic rats) as compared to basal corticosterone in naïve pups and corticosterone levels in pups removed from the experimental chamber without hypoxia (normoxic rats). Adult rats (110-day-old) that were exposed to hypoxia at the postnatal day 7 showed increased corticosterone levels as compared to the basal corticosterone in naïve adult rats. The “direction” of hormonal reaction in response to the forced swimming depended on age and differed in hypoxic and normoxic rats. The forced swimming increased plasma corticosterone in naïve and normoxic adults and decreased in pups but failed to alter it in hypoxic adults and in naïve and normoxic 7-day-old. Thus, short- and long-term effects of early postnatal hypoxia revealed themselves in the decrease of responsiveness of the HPA axis to the forced swimming. The hypoxia reduced the time of immobility in the forced swim test and enhanced pain-related response in pups in the formalin test as compared to these indices in normoxic pups. Hypoxic adult rats demonstrated the increased time of immobility during the forced swim test as compared to immobility in normoxic rats. Early postnatal hypoxia disturbed interrelations between depression- and pain-related indices.     

Chronic l-menthol-induced browning of white adipose tissue hypothesis: A putative therapeutic regime for combating obesity and improving metabolic health

IntroductionObesity constitutes a serious global health concern reaching pandemic prevalence rates. The existence of functional brown adipose tissue (BAT) in adult humans has provoked intense research interest in the role of this metabolically active tissue in whole-body energy balance and body weight regulation. A number of environmental, physiological, pathological, and pharmacological stimuli have been proposed to induce BAT-mediated thermogenesis and functional thermogenic BAT-like activity in white adipose tissue (WAT), opening new avenues for therapeutic strategies based on enhancing the number of beige adipocytes in WAT.HypothesisRecent evidence support a role of l-menthol cooling, mediated by TRPM8 receptor, on UCP1-dependent thermogenesis and BAT-like activity in classical WAT depots along with the recruitment of BAT at specific anatomical sites. l-Menthol-induced BAT thermogenesis has been suggested to occur by a β-adrenergic-independent mechanism, avoiding potential side-effects due to extensive β-adrenergic stimulation mediated by available beta receptor agonists. l-Menthol has been also linked to the activation of the cold-gated ion channel TRPA1. However, its role in l-menthol-induced UCP1-dependent thermogenic activity in BAT and WAT remains undetermined. White adipose tissue plasticity has important clinical implications for obesity prevention and/or treatment because higher levels of UCP1-dependent thermogenesis can lead to enhanced energy expenditure at a considerable extent. We hypothesize that chronic dietary l-menthol treatment could induce TRPM8- and TRPA1-dependent WAT adaptations, resembling BAT-like activity, and overall improve whole-body metabolic health in obese and overweight individuals.ConclusionsThe putative impact of chronic l-menthol dietary treatment on the stimulation of BAT-like activity in classical WAT depots in humans remains unknown. A detailed experimental design has been proposed to investigate the hypothesized l-menthol-induced browning of WAT. If our hypothesis was to be confirmed, TRPM8/TRPA1-induced metabolic adaptations of WAT to BAT-like activity could provide a promising novel therapeutic approach for increasing energy expenditure, regulating body weight, and preventing obesity and its related co-morbidities in humans.     

Independence of ultrasonic vocalization and thermogenic responses in infant rats

Ultrasonic vocalizations (USV) normally accompany brown adipose tissue (BAT) thermogenesis in infant rats exposed to cold. BAT activation (measured by implanted thermistors) was pharmacologically blocked with hexamethonium (20 mg/kg ip) in 12-13-day-old pups, but they nevertheless showed normal USV responses to cold. Activation of BAT in warm pups by norepinephrine (800 micrograms/kg sc) failed to elicit USV. It is concluded that BAT activation is neither necessary nor sufficient for USV production. To evaluate how tightly the two responses may be coupled centrally, rat pups deprived of nutrients for 24 hr, in which sympathetic activation is known to be inhibited centrally (Bignall, Heggeness, & Palmer, 1975), were studied. These pups vocalized with the same latency in response to cold as normals but failed to show evidence of concurrent BAT activation. It is concluded that USV and BAT thermogenesis are normally elicited together by cold but are not tightly linked physiologically.     

Ontogeny of behavioral inhibition induced by unfamiliar adult male conspecifics in preweanling rats.

Previous studies showed that when socially isolated at 22 degrees C, postnatal day 14 rats, but not younger day 7 rats, reduce their emission of ultrasonic vocalizations when exposed to an unfamiliar adult male rat, a naturalistic threat. Because ultrasound production is associated with factors such as age and body temperature, this study examined in age-appropriate thermoneutral temperature ranges whether preweanling rats of different ages are equally capable of inhibiting their emission of ultrasounds when threatened. In Experiment 1, 7- and 14-day-old rats were socially isolated and exposed to unfamiliar anesthetized adult male rats in a thermoneutral environment. Only 14-day-old rats significantly reduced their emission of ultrasounds. This reduction in ultrasound production was accompanied by freezing. In Experiment 2, additional ages were examined under identical test conditions. At 3, 6, and 9 days of age, pups frequently emitted ultrasounds when exposed to the anesthetized male rat. However, at 12 days of age, rat pups responded to the anesthetized male rat by freezing and significantly reducing their emission of ultrasounds. Results indicate clearly that under the present testing conditions the ability of rat pups to inhibit ultrasounds and freeze when threatened is not present at birth but emerges by the end of the second postnatal week.     

Fictive motor activity in rat after 14 days of hindlimb unloading

The goal of the present study was to examine the effects of chronic hindlimb unloading on fictive motor patterns which can be developed in hindlimb nerves of adult rats. The animals were divided into two groups. The first group was submitted to hindlimb unloading for 2 weeks by tail suspension. The second group served as controls. After this initial phase, the animals of both groups were acutely decorticated, paralysed and electroneurographic efferent activity was recorded from hindlimb muscle nerves under conditions of "fictive locomotion" in order to evaluate variations in central locomotor command. Fictive rhythmic motor episodes were either spontaneous or evoked by electrical stimulation of the mesencephalic locomotor region. Only the second ones were recognised as locomotor-like activities. The motor pattern was not fundamentally affected by unloading except that, after the unloading period, extensor muscle nerves were significantly more frequently activated and their burst durations were increased compared to activity in control animals, despite the fact that the phasic sensory afferent inputs were suppressed. This suggests that unloading induces plastic modifications of the central networks of neurons implicated in the locomotor command. The origin of this extensor hyperactivity is discussed. It is proposed that it could be the consequence of either changes in motoneuronal properties or of an increase in afferent input to motoneurones. Electronic Publication     

Concomitant food intake and adipose tissue responses under chronic insulin infusion in rats

Body weight (BW), food intake (FI), and activity of white adipose tissue (WAT) and brown adipose tissue (BAT) were studied in adult male rats under chronic insulin infusion. Insulin was infused for 4, 7 or 10 days via implanted minipumps. Insulin-treated rats gained more BW than control rats until 7th day of infusion. At 10 days, the difference in BW decreased. The average cumulative FI was significantly higher after 4, 7 and 10 days of insulin infusion. Feed efficiency (FE) was increased in insulin-treated rats after 4 and 7 days. An increase in WAT weight was observed in insulin-treated rats together with an increased activity of lipogenic enzymes. BAT weight was augmented after 4 days of insulin infusion. This was due mainly to lipid accumulation. Specific mitochondrial guanosine diphosphate (GDP) binding was significantly decreased by 58% in insulin-treated rats after 4 days of infusion. This reduced thermogenic activity, along with the increased FI and FE were responsible for the rapid BW gain observed during the first 7 days of insulin infusion.     

Cold-induced thermogenesis in hypothyroid rats

Hypothyroidism was induced in adult rats by oral administration of methimazole. Euthyroid and hypothyroid rats were maintained at 23 °C or exposed at 6 °C for 3 weeks. Both euthyroid and hypothyroid rats maintained at 23 °C had similar interscapular brown adipose tissue (BAT) composition and thermogenic activity. Cold-exposed hypothyroid rats showed the same interscapular BAT mass and gross tissue composition as cold-exposed euthyroid animals, but the interscapular BAT of cold-exposed hypothyroid rats did not show the characteristic increase in GDP binding, and the increase in mitochondrial mass was lower than in euthyroid rats. From these results we conclude that thyroid hormones do not influence BAT significantly when thermogenic requirements are moderate, but they participate in the trophic response of the tissue when thermogenic requirements are intense. This thyroid hormone participation in the BAT trophic response occurs at the mitochondrial level, both in quantitative (mitochondrial mass) and qualitative (GDP-binding) aspects.     

2-Mercaptoacetate does not stimulate chow intake in periweanling rats

In adult rats, administration of drugs that suppress oxidation of fatty acids, like mercaptoacetate (MA), produces increases in food intake. During development, the consequences of administration of MA are more varied. For example, in very young pups, intake of milk diets is unaffected by MA, while pups aged 12 to 15 days demonstrate increases in intake. However, in 18- and 21-day-old rats, milk intake is suppressed by administration of MA. Typically, the paradigms used to test rat pups differ significantly from those used to assess intake in adult rats. The present experiments were designed to examine whether 18-day-old pups tested with adultlike paradigms showed adultlike responses to administration of MA. In the first experiment, rat pups aged 18 days were injected with 0 or 68.4 mg/kg MA, then given 60-min tests while consuming either milk or chow diets that were novel, or to which they had previously been exposed. The results demonstrated that chow intake was not affected by administration of MA, but milk intake in experienced animals was suppressed by MA. Experiment 2 demonstrated that in contrast to administration of MA, 18-day-old pups deprived of food overnight showed increases in intake of chow and milk diets. In Experiment 3, when the effects of a range of doses of MA (22.8, 45.6, 68.4 and 91.2) on chow intake over a 4-h period were assessed, all doses of MA produced a significant suppression of chow intake in 18-day-old pups. Taken together, the data suggest that alterations in fatty acid oxidation produced by administration of MA do not stimulate chow intake in periweanling pups tested in an adultlike fashion.     

Development of the circadian rhythm of neuronal activity in suprachiasmatic nucleus of rat hypothalamic slices

Development of the circadian rhythm of neuronal activity in the suprachiasmatic nucleus (SCN) was studied in rat hypothalamic slices. The firing rate of SCN neurons of 7- and 11-day-old rat pups was low during all of the day, whereas that of 14- and 21-day-old pups was high during daytime and low during nighttime. The present results suggest that the circadian rhythm of SCN neuronal activity is established between 11 and 14 days of age, corresponding closely to the time of onset of the other various hormonal and behavioral circadian rhythms.     

Rat behavioral thermoregulation integrates with nonshivering thermogenesis during postnatal development

Rat pups are capable of behavioral thermoregulation, both in the nest and on a thermocline, as early as the 1st week of postnatal life, and these pups can also produce heat metabolically without shivering. The rat pup's primary source of nonshivering thermogenesis is the sympathetically mediated metabolism of brown adipose tissue (BAT). BAT is well formed in newborns and functions shortly after birth. While infant behavioral thermoregulation and BAT thermogenesis have been extensively studied, little is known about the extent to which thermoregulatory behavior can be influenced by BAT thermogenesis. In the present study, 2-, 7-, and 14-day-old pups were observed on a thermal gradient following pharmacological stimulation or inhibition of BAT thermogenesis, and their thermal preferences were quantified. The authors found that 7- and 14-day-old pups treated with norepinephrine (NE), which increases BAT thermogenesis, preferred cooler portions of the gradient than saline-treated controls, whereas 2-day-olds failed to show a similar NE-induced behavioral adjustment. These findings indicate that the ability to adjust thermoregulatory behavior to compensate for enhanced metabolic thermogenesis develops during the 1st week of postnatal life.