Amy Warner

Effects of Manipulating Hypothalamic Triiodothyronine Concentrations on Seasonal Body Weight and Torpor Cycles in Siberian Hamsters

Siberian hamsters display photoperiodically regulated annual cycles in body weight, appetite, and reproduction. Previous studies have revealed a profound up-regulation of type 3 deiodinase (DIO3) mRNA in the ventral ependyma of the hypothalamus associated with hypophagia and weight loss in short-day photoperiods. DIO3 reduces the local availability of T(3), so the aim of this study was to test the hypothesis that decreased hypothalamic T(3) availability underlies the short-day-induced catabolic state. The experimental approach was to determine whether a local increase in T(3) in the hypothalamus of hamsters exposed to short days could reverse the behavioral and physiological changes induced by this photoperiod. In study 1, microimplants releasing T(3) were placed bilaterally into the hypothalamus. This treatment rapidly induced a long-day phenotype including increased appetite and body weight within 3 wk of treatment and increased fat mass and testis size by the end of the 10-wk study period. In study 2, hypothalamic T(3) implants were placed into hamsters carrying abdominal radiotelemetry implants. Again body weight increased significantly, and the occurrence of winter torpor bouts was dramatically decreased to less than one bout per week, whereas sham-implanted hamsters entered torpor up to six times a week. Our findings demonstrate that increased central T(3) induces a long-day metabolic phenotype, but in neither study was the molt cycle affected, so we infer that we had not disrupted the initial detection of photoperiod. We conclude that hypothalamic thyroid hormone availability plays a key role in seasonal regulation of appetite, body weight, and torpor.

Thyrotrophin-releasing hormone decreases feeding and increases body temperature, activity and oxygen consumption in Siberian hamsters.

Thyrotrophin‐releasing hormone (TRH) is known to play an important role in the control of food intake and energy metabolism in addition to its actions on the pituitary‐thyroid axis. We have previously shown that central administration of TRH decreases food intake in Siberian hamsters. This species is being increasingly used as a physiological rodent model in which to understand hypothalamic control of long‐term changes in energy balance because it accumulates fat reserves in long summer photoperiods, and decreases food intake and body weight when exposed to short winter photoperiods. The objectives of our study in Siberian hamsters were: (i) to investigate whether peripheral administration of TRH would mimic the effects of central administration of TRH on food intake and whether these effects would differ dependent upon the ambient photoperiod; (ii) to determine whether TRH would have an effect on energy expenditure; and (iii) to investigate the potential sites of action of TRH. Both peripheral (5–50 mg/kg body weight; i.p.) and central (0.5 µg/ml; i.c.v.) administration of TRH decreased food intake, and increased locomotor activity, body temperature and oxygen consumption in the Siberian hamster, with a rapid onset and short duration of action. Systemic treatment with TRH was equally effective in suppressing feeding regardless of ambient photoperiod. The acute effects of TRH are likely to be centrally mediated and independent of its role in the control of the production of thyroid hormones. We conclude that TRH functions to promote a catabolic energetic state by co‐ordinating acute central and chronic peripheral (thyroid‐mediated) function.

Effects of photoperiod on daily locomotor activity, energy expenditure, and feeding behavior in a seasonal mammal

The objective of this study was to determine whether the previously observed effects of photoperiod on body weight in Siberian hamsters were due to changes in the daily patterns of locomotor activity, energy expenditure, and/or feeding behavior. Adult males were monitored through a seasonal cycle using an automated comprehensive laboratory animal monitoring system (CLAMS). Exposure to a short-day photoperiod (SD; 8:16-h light-dark cycle) induced a significant decline in body weight, and oxygen consumption (Vo(2)), carbon dioxide production (Vco(2)), and heat production all decreased reaching a nadir by 16 wk of SD. Clear daily rhythms in locomotor activity, Vo(2), and Vco(2) were observed at the start of the study, but these all progressively diminished after prolonged exposure to SD. Rhythms in feeding behavior were also detected initially, reflecting an increase in meal frequency but not duration during the dark phase. This rhythm was lost by 8 wk of SD exposure such that food intake was relatively constant across dark and light phases. After 18 wk in SD, hamsters were transferred to a long-day photoperiod (LD; 16:8-h light-dark cycle), which induced significant weight gain. This was associated with an increase in energy intake within 2 wk, while Vo(2), Vco(2), and heat production all increased back to basal levels. Rhythmicity was reestablished within 4 wk of reexposure to long days. These results demonstrate that photoperiod impacts on body weight via complex changes in locomotor activity, energy expenditure, and feeding behavior, with a striking loss of daily rhythms during SD exposure.

Increased Responses to the Actions of Fibroblast Growth Factor 21 on Energy Balance and Body Weight in a Seasonal Model of Adiposity

The present study aimed to investigate the actions of fibroblast growth factor 21 (FGF21) on energy balance in a natural model of relative fatness, the Siberian hamster. Hamsters were studied under long days (LD) to promote weight gain, or short days to induce weight loss, and treated with rhFGF21 (3 mg/kg/day) via s.c. minipumps for 14 days. On days 7–9, detailed assessments of ingestive behaviour, metabolic gas exchange and locomotor activity were made. FGF21 caused substantial (P < 0.0001) weight loss in the fat LD state but not in the lean SD state: at the end of the study, FGF21‐treated hamsters in LD lost 18% of body weight compared to vehicle controls, which is comparable to the natural body weight loss observed in SD. Epididymal fat pads, a correlate of total carcass fat content, were reduced by 19% in FGF21 treated hamsters in LD, whereas no difference was found in SD. Body weight loss in LD was associated with a reduction in food intake (P < 0.001) and a decreased respiratory exchange ratio (P < 0.001), indicating increased fat oxidation. Treatment with FGF21 maintained the normal nocturnal increase in oxygen consumption and carbon dioxide production into the early light phase in hamsters in LD, indicating increased energy expenditure, although locomotor activity was unaffected. These data suggest a greater efficacy of FGF21 in hamsters in LD compared to those in SD, which is consistent with both the peripheral and possibly central actions of FGF21 with respect to promoting a lean phenotype. The observed differences in FGF21 sensitivity may relate to day length‐induced changes in adipose tissue mass.

Loss of prokineticin receptor 2 signaling predisposes mice to torpor

The genes encoding prokineticin 2 polypeptide (Prok2) and its cognate receptor (Prokr2/Gpcr73l1) are widely expressed in both the suprachiasmatic nucleus and its hypothalamic targets, and this signaling pathway has been implicated in the circadian regulation of behavior and physiology. We have previously observed that the targeted null mutation of Prokr2 disrupts circadian coordination of cycles of locomotor activity and thermoregulation. We have now observed spontaneous but sporadic bouts of torpor in the majority of these transgenic mice lacking Prokr2 signaling. During these torpor bouts, which lasted for up to 8 h, body temperature and locomotor activity decreased markedly. Oxygen consumption and carbon dioxide production also decreased, and there was a decrease in respiratory quotient. These spontaneous torpor bouts generally began toward the end of the dark phase or in the early light phase when the mice were maintained on a 12:12-h light-dark cycle and persisted when mice were exposed to continuous darkness. Periods of food deprivation (16-24 h) induced a substantial decrease in body temperature in all mice, but the duration and depth of hypothermia was significantly greater in mice lacking Prokr2 signaling compared with heterozygous and wild-type littermates. Likewise, when tested in metabolic cages, food deprivation produced greater decreases in oxygen consumption and carbon dioxide production in the transgenic mice than controls. We conclude that Prokr2 signaling plays a role in hypothalamic regulation of energy balance, and loss of this pathway results in physiological and behavioral responses normally only detected when mice are in negative energy balance.

Antibody-Mediated Inhibition of the FGFR1c Isoform Induces a Catabolic Lean State in Siberian Hamsters

Hypothalamic tanycytes are considered to function as sensors of peripheral metabolism. To facilitate this role, they express a wide range of receptors, including fibroblast growth factor receptor 1 (FGFR1). Using a monoclonal antibody (IMC-H7) that selectively antagonizes the FGFR1c isoform, we investigated possible actions of FGFR1c in a natural animal model of adiposity, the Siberian hamster. Infusion of IMC-H7 into the third ventricle suppressed appetite and increased energy expenditure. Likewise, peripheral treatment with IMC-H7 decreased appetite and body weight and increased energy expenditure and fat oxidation. A greater reduction in body weight and caloric intake was observed in response to IMC-H7 during the long-day fat state as compared to the short-day lean state. This enhanced response to IMC-H7 was also observed in calorically restricted hamsters maintained in long days, suggesting that it is the central photoperiodic state rather than the peripheral adiposity that determines the response to FGFR1c antagonism. Hypothalamic thyroid hormone availability is controlled by deiodinase enzymes (DIO2 and DIO3) expressed in tanycytes and is the key regulator of seasonal cycles of energy balance. Therefore, we determined the effect of IMC-H7 on hypothalamic expression of these deiodinase enzymes. The reductions in food intake and body weight were always associated with decreased expression of DIO2 in the hypothalamic ependymal cell layer containing tanycytes. These data provide further support for the notion the tanycytes are an important component of the mechanism by which the hypothalamus integrates central and peripheral signals to regulate energy intake and expenditure.

Short-Days Induce Weight Loss in Siberian Hamsters Despite Overexpression of the Agouti-Related Peptide Gene

Many vertebrates express profound annual cycles of body fattening, although it is not clear whether these represent differential activity of the central pathways known to mediate homeostatic control of food intake and energy expenditure, or whether the recent discovery of a major role for pars tuberalis‐ependymal signalling points towards novel mechanisms. We examined this in the Siberian hamster (Phodopus sungorus) by using gene transfection to up‐regulate a major orexigenic peptide, agouti‐related peptide (AgRP), and then determined whether this increased anabolic drive could prevent the short‐day induced winter catabolic state. Infusions of a recombinant adeno‐associated virus encoding an AgRP construct into the hypothalamus of hamsters in the long‐day obese phase of their seasonal cycle produced a 20% gain in body weight over 6 weeks compared to hamsters receiving a control reporter construct, reflecting a significant increase in food intake and a significant decrease in energy expenditure. However, all hamsters showed a significant, prolonged decrease in body weight when exposed to short photoperiods, despite the hamsters expressing the AgRP construct maintaining a higher food intake and lower energy expenditure relative to the control hamsters. Visualisation of the green fluorescent protein reporter and analysis of AgRP‐immunoreactivity confirmed widespread expression of the construct in the hypothalamus, which was maintained for the 21‐week duration of the study. In conclusion, the over‐expression of AgRP in the hypothalamus produced a profoundly obese state but did not block the seasonal catabolic response, suggesting a separation of rheostatic mechanisms in seasonality from those maintaining homeostasis of energy metabolism.

The role of histamine 3 receptors in the control of food intake in a seasonal model of obesity: the Siberian hamster.

Siberian hamsters develop hypophagia and increase catabolism of fat reserves in response to short photoperiods resulting in a natural loss of body weight in winter. We previously found that histamine 3 receptor (H3R) mRNA in the posterior hypothalamus is significantly decreased in short photoperiods. We hypothesized that this lower expression of H3R might contribute to the winter hypophagic state, therefore we examined the effects of the H3R agonist imetit and inverse agonists clobenpropit and thioperamide on food intake. We expressed the Siberian hamster H3R receptor in vitro and confirmed that imetit, clobenpropit and thioperamide are bound specifically, thus validating them as tools to investigate the role of H3R in vivo. Intracerebroventricular administration of histamine decreased food intake in hamsters in the fat summer state. Administration of imetit to hamsters in the lean state increased food intake, whereas administration of inverse agonists decreased food intake, though this was associated with decreased locomotor activity. Both H3R inverse agonists prevented the nocturnal rise in body temperature indicating additional effects on energy expenditure. In summary, our results suggest that increased availability of central histamine or the reduction of H3R activity decrease food intake. These effects are similar to those observed in hamsters in short photoperiods.

Histaminergic regulation of seasonal metabolic rhythms in Siberian hamsters

We investigated whether histaminergic tone contributes to the seasonal catabolic state in Siberian hamsters by determining the effect of ablation of histaminergic neurons on food intake, metabolic rate and body weight. A ribosomal toxin (saporin) conjugated to orexin-B was infused into the ventral tuberomammillary region of the hypothalamus, since most histaminergic neurons express orexin receptors. This caused not only 75–80% loss of histaminergic neurons in the posterior hypothalamus, but also some loss of other orexin-receptor expressing cells e.g. MCH neurons. In the long-day anabolic state, lesions produced a transient post-surgical decrease in body weight, but the hamsters recovered and maintained constant body weight, whereas weight gradually increased in sham-lesioned hamsters. VO2 in the dark phase was significantly higher in the lesioned hamsters compared to shams, and locomotor activity also tended to be higher. In a second study in short days, sham-treated hamsters showed the expected seasonal decrease in body weight, but weight remained constant in the lesioned hamsters, as in the long-day study. Lesioned hamsters consumed more during the early dark phase and less during the light phase due to an increase in the frequency of meals during the dark and decreased meal size during the light, and their cumulative food intake in their home cages was greater than in the control hamsters. In summary, ablation of orexin-responsive cells in the posterior hypothalamus blocks the short-day induced decline in body weight by preventing seasonal hypophagia, evidence consistent with the hypothesis that central histaminergic mechanisms contribute to long-term regulation of body weight.