Comparative endocrine disruptive effects of contaminants in ringed seals (Phoca hispida) from Svalbard and the Baltic Sea

We investigated variables related to thyroid, vitamin A and calcitriol homeostasis, immune function and tumour development in ringed seals (Phoca hispida) from the polluted Baltic Sea and a less polluted reference location at Svalbard, Norway. We also examined the relationships between the biological variables and the concentrations of persistent organic pollutants (POPs) and their hydroxylated (OH) metabolites. Our data show higher plasma concentrations of free triiodothyronine (T3), and ratios of free and total T3 in Baltic seals as compared to Svalbard seals. Baltic seals had also higher hepatic mRNA expressions of deiodinase-I, thyroid hormone receptor β, retinoic acid receptor α, growth hormone receptor and interleukin-1β compared to Svalbard seals. Levels of plasma retinol were lower in the Baltic seals as compared to Svalbard seals. No geographical difference was observed for other thyroid hormone levels and hepatic retinoid levels. Ratios of free and total T3 were positively correlated to OH-POPs in plasma. The results of the present study suggest that endocrine homeostasis may be affected by contaminant and metabolite exposure in the Baltic ringed seals with respect to circulating hormones and retinol and hepatic mRNA expressions. In addition, OH-POPs may putatively produce the disruption of thyroid hormone transport in plasma.     

Effects of fasting on growth hormone/insulin-like growth factor I axis in the tilapia,Oreochromis mossambicus

Effects of fasting on the growth hormone (GH)-insulin-like growth factor I (IGF-I) axis were examined in the tilapia (Oreochromis mossambicus) acclimated to fresh water. Fasting for 2 weeks resulted in significant reductions in body weight, specific growth rate and hepatosomatic index in both males and females. Significant reductions in specific growth rates were observed after 1 and 2 weeks in both sexes, although the decrease in body weight was not significant in the female. A significant reduction was also seen in the condition factor of females after 2 weeks. No change was seen in the gonadosomatic index in either sex. Two weeks of fasting also produced a significant reduction in plasma IGF-I but not in plasma GH, prolactin (PRL(188)) or cortisol. Significant reductions in the hepatic IGF-I mRNA were seen in both sexes. On the other hand, a significant increase was observed in cortisol receptor mRNA in the female liver. Plasma IGF-I levels were correlated significantly with specific growth rate, condition factor and hepatosomatic index, indicating that plasma IGF-I is a good indicator of growth in the tilapia. No change was seen in plasma glucose or osmolality after 2 weeks of fasting. During fasting, tilapia appears to convert metabolic energy from growth to basal metabolism including maintenance of ion and water balance.     

Effect of fasting on thyroid hormone levels, and TR(alpha) and TR(beta) mRNA accumulation in late-stage embryo and juvenile rainbow trout, Oncorhynchus mykiss

The accumulation of mRNA encoding for hepatic and intestinal T3-receptor (TR) and body and liver masses were measured in fed and 3-week fasted juvenile and swim up stage rainbow trout embryos. Plasma and total body thyroid hormone (TH) levels were measured for juvenile and swim up stages, respectively. Fasted juveniles exhibited a lower hepatosomatic index (HSI), liver mass and plasma T4 and T3 concentrations than fed animals, but there were no changes in body mass or the accumulation of mRNA encoding for either of the TR(alpha) or TR(beta) isoforms in liver or intestine. TR(beta) mRNA accumulation was greater than TR(alpha) mRNA accumulation in both tissues. Fasted embryos had lower whole body TH levels and body, liver and intestinal tract masses, in addition to a lower intestinosomatic index. However, there was no change in HSI. Fasting did not affect whole body or hepatic TR(alpha) and TR(beta) mRNA accumulation, although intestinal tract TR(alpha) and TR(beta) mRNA accumulation was lower in the fasted embryos. The HSI and body mass changes in fasted juvenile and embryo stages, respectively, indicated that both developmental stages were impacted by fasting. Both stages also showed evidence of decreased TH production. The lower TR gene expression in the intestinal tract of fasted embryos may suggest a role for THs in the transitional stage of intestinal development during this period of development.     

Changes in thyroid hormone levels in chicken liver during fasting and refeeding

In chickens, fasting results in increased plasma thyroxine (T(4)) levels and decreased plasma 3,5,3'-triiodothyronine (T(3)) levels. Refeeding, in turn, restores normal plasma T(3) and T(4) levels. The liver is an important tissue for the regulation of circulating thyroid hormone levels. Previous studies demonstrated that the increase in hepatic type III deiodinase in fasted chickens plays a role in the decrease of plasma T(3). Another factor that could be important is the level of T(4) and T(3) uptake by the liver. In mammals, caloric restriction is known to diminish transport of T(4) and T(3) into tissues. The present study examines whether this is also the case in chicken. Four-week-old chickens were subjected to a 24-h starvation period followed by refeeding. Blood and liver samples were collected at the start of refeeding and at different times of refeeding. Thyroid hormone levels were measured directly in plasma and in tissues following extraction. The results demonstrate that intrahepatic T(4) levels are increased and T(3) levels are decreased in fasted compared to ad libitum fed chickens. The parallel changes in plasma and hepatic T(3) and T(4) content demonstrate that T(4) availability in liver tissue is not diminished during fasting, suggesting that in chicken thyroid hormone uptake by the liver is not affected by nutritional status.     

Differential regulation of thyrotropin-releasing hormone receptor mRNA levels by thyroid hormone in vivo and in vitro (GH3 cells).

We studied the effect of thyroid status on thyrotropin-releasing hormone receptor (TRH-R) mRNA levels both in vivo and in vitro (GH3 cells) using a cloned rat TRH-R cDNA by RT-PCR. Experimental hypothyroid rats were produced by total thyroidectomy and were then killed 7 days after the operation. TRH receptor binding in the anterior pituitary and serum TSH level were elevated approximately 2-fold and 8-fold, respectively, in 7 day thyroidectomized rats. TRH-R mRNA levels in hypothyroid rats were also increased significantly compared with those of normal rats. In GH3 cells, however, no significant change of TRH-R mRNA level was observed between cultures treated with triiodothyronine (T3, 10(-9) and 10(-7) M) and the untreated group. The present data indicate that 1) the in vivo effects of thyroid status on TRH-R mRNA levels differ from the in vitro one, and that 2) the down regulation of TRH-R binding by thyroid hormone in GH3 cells may be mediated by translational or post-translational mechanisms.     

PCB153 disrupts thyroid hormone homeostasis by affecting its biosynthesis, biotransformation, feedback regulation, and metabolism

PCB153, one of the 3 dominant congeners in the food chain, causes the disruption of the endocrine system in humans and animals. In order to elucidate the effects of PCB153 on the biosynthesis, biotransformation, regulation, metabolism, and transport of thyroid hormones (THs), Sprague-Dawley (SD) rats were dosed with PCB153 intraperitoneally (i.p.) at 0, 4, 16 and 32 mg/kg/day for 5 consecutive days and sacrificed 24 h after the last dose. Results showed that after treatment with PCB153, serum total thyroxine (TT4), total triiodothyronine (TT3), and thyrotropin releasing hormone (TRH) decreased, whereas serum thyroid stimulating hormone (TSH) concentration did not alter. The serum sodium iodide symporter (NIS), thyroid peroxidase (TPO), and thyroglobulin (Tg) levels decreased. The mRNA expressions of type 2 and 3 deiodinases (D2 and D3) reduced, but the type 1 deiodinase (D1) showed no significant change. The TSH receptor (TSHr) and TRH receptor (TRHr) levels declined. PCB153 induced hepatic enzymes, and the UDPGTs, CYP2B1, and CYP3A1 mRNA levels were significantly elevated. Taken together, the observed results from the present study indicated that PCB153 disrupted thyroid hormone homeostasis through influencing synthesis-associated proteins (NIS, TPO and Tg), deiodinases, receptors (TSHr and TRHr), and hepatic enzymes, and the decrease of D3 expression might be the compensatory response of body.     

Thyroid hormone stimulation of NADPH P450 reductase expression in liver and extrahepatic tissues. Regulation by multiple mechanisms.

The role of thyroid hormone in regulating the expression of the flavoprotein NADPH cytochrome P450 reductase was studied in adult rats. Depletion of circulating thyroid hormone by hypophysectomy, or more selectively, by treatment with the anti-thyroid drug methimazole led to a 75-85% depletion of hepatic microsomal P450 reductase activity and protein in both male and female rats. Thyroxine substantially restored P450 reductase activity at a dose that rendered the thyroid-depleted rats euthyroid. Microsomal P450 reductase activity in several extrahepatic tissues was also dependent on thyroid hormone, but to a lesser extent than in liver (30-50% decrease in kidney, adrenal, lung, and heart but not in testis from hypothyroid rats). Hepatic P450 reductase mRNA levels were also decreased in the hypothyroid state, indicating that the loss of P450 reductase activity is not a consequence of the associated decreased availability of the FMN and FAD cofactors of P450 reductase. Parallel analysis of S14 mRNA, which has been studied extensively as a model thyroid-regulated liver gene product, indicated that P450 reductase and S14 mRNA respond similarly to these changes in thyroid state. In contrast, while the expression of S14 and several other thyroid hormone-dependent hepatic mRNAs is stimulated by feeding a high carbohydrate, fat-free diet, hepatic P450 reductase expression was not increased by this lipogenic diet. Injection of hypothyroid rats with T3 at a supraphysiologic, receptor-saturating dose stimulated a major induction of hepatic P450 reductase mRNA that was detectable 4 h after the T3 injection, and peaked at approximately 650% of euthyroid levels by 12 h. However, this same treatment stimulated a biphasic increase in P450 reductase protein and activity that required 3 days to reach normal euthyroid levels. T3 treatment of euthyroid rats also stimulated a major induction of P450 reductase mRNA that was maximal (12-fold increase) by 12 h, but in this case no major increase in P450 reductase protein or activity was detectable over a 3-day period. Together, these studies establish that thyroid hormone regulates P450 reductase expression by pretranslational mechanisms. They also suggest that other regulatory mechanisms, which may involve changes in P450 reductase protein stability and/or changes in the translational efficiency of its mRNA, are likely to occur.     

Triiodothyronine increases rat apolipoprotein A-I synthesis and alters high-density lipoprotein composition in vivo

The effects of altered serum 3,3',5-triiodothyronine levels on rat lipoprotein metabolism were examined. Daily injections of the hormone (50 micrograms/100 g body mass) over a period of six days led to an increase of 6.4-fold in the hepatic mRNA level for apolipoprotein(apo)A-I, and a 21% increase in serum apoA-I levels. 12h after a single injection of 3,3',5-triiodothyronine the rate of [14C]leucine incorporation into apoA-I increased 2.1 fold. Conversely, in hypothyroid rats there was a decrease in hepatic mRNA levels for apoA-I and a decreased rate of [14C]leucine incorporation into apoA-I. The increase in hepatic apoA-I mRNA levels following 3,3',5-triiodothyronine treatment occurred prior to significant changes in serum triacylglycerol levels. High-density lipoprotein (HDL) particles isolated from the serum of hyperthyroid rats were smaller and enriched in apoA-I compared to apoA-IV and apoE. Similar changes in HDL composition were observed following in vitro incubations of normal rat serum with purified rat apoA-I. The results suggest that during altered thyroid status, changes in serum HDL size and composition occur in association with significant changes in apoA-I gene expression.     

NPY and MC4R signaling regulate thyroid hormone levels during fasting through both central and peripheral pathways

Fasting-induced suppression of the hypothalamic-pituitary-thyroid (HPT) axis is an adaptive response to decrease energy expenditure during food deprivation. Previous studies demonstrate that leptin communicates nutritional status to the HPT axis through thyrotropin-releasing hormone (TRH) in the paraventricular nucleus (PVN) of the hypothalamus. Leptin targets TRH neurons either directly or indirectly via the arcuate nucleus through pro-opiomelanocortin (POMC) and agouti-related peptide/neuropeptide Y (AgRP/NPY) neurons. To evaluate the role of these pathways in vivo, we developed double knockout mice that lack both the melanocortin 4 receptor (MC4R) and NPY. We show that NPY is required for fasting-induced suppression of Trh expression in the PVN. However, both MC4R and NPY are required for activation of hepatic pathways that metabolize T₄ during the fasting response. Thus, these signaling pathways play a key role in the communication of fasting signals to reduce thyroid hormone levels both centrally and through a peripheral hepatic circuit.     

Age and sex differences in the timing of moult in the common seal, Phoca vitulina

This study followed the progress of the annual moult within a population of common seals in Orkney, Scotland. Moulting seals were seen over a three-month period, from 7 June until 16 September. Yearlings were first to start moulting. Amongst older seals, females completed their moult an average of seven days earlier than immature males and 19 days earlier than mature males. Differences in the timing of moult appeared to be related to the age or reproductive status of the animals, and may be the result of differential changes in levels of the sex hormones.     

Effects of leptin replacement on hypothalamic-pituitary growth hormone axis function and circulating ghrelin levels in ob/ob mice

Leptin-deficient obese mice (ob/ob) have decreased circulating growth hormone (GH) and pituitary GH and ghrelin receptor (GHS-R) mRNA levels, whereas hypothalamic GH-releasing hormone (GHRH) and somatostatin (SST) expression do not differ from lean controls. Given the fact that GH is suppressed in diet-induced obesity (a state of hyperleptinemia), it remains to be determined whether the absence of leptin contributes to changes in the GH axis of ob/ob mice. Therefore, to study the impact of leptin replacement on the hypothalamic-pituitary GH axis of ob/ob mice, leptin was infused for 7 days (sc), resulting in circulating leptin levels that were similar to wild-type controls (approximately 1 ng/ml). Leptin treatment reduced food intake, body weight, and circulating insulin while elevating circulating n-octanoyl ghrelin concentrations. Leptin treatment did not alter hypothalamic GHRH, SST, or GHS-R mRNA levels compared with vehicle-treated controls. However, leptin significantly increased pituitary GH and GHRH-R expression and tended to enhance circulating GH levels, but this latter effect did not reach statistical significance. In vitro, leptin (1 ng/ml, 24 h) did not affect pituitary GH, GHRH-R, or GHS-R mRNA but did enhance GH release. The in vivo effects of leptin on circulating hormone and pituitary mRNA levels were not replicated by pair feeding ob/ob mice to match the food intake of leptin-treated mice. However, leptin did prevent the fall in hypothalamic GHRH mRNA and circulating IGF-I levels observed in pair-fed mice. These results demonstrate that leptin replacement has positive effects on multiple levels of GH axis function in ob/ob mice.     

Food deprivation explains effects of mouthbrooding on ovaries and steroid hormones, but not brain neuropeptide and receptor mRNAs, in an African cichlid fish

Feeding behavior and reproduction are coordinately regulated by the brain via neurotransmitters, circulating hormones, and neuropeptides. Reduced feeding allows animals to engage in other behaviors important for fitness, including mating and parental care. Some fishes cease feeding for weeks at a time in order to provide care to their young by brooding them inside the male or female parent's mouth. Maternal mouthbrooding is known to impact circulating hormones and subsequent reproductive cycles, but neither the full effects of food deprivation nor the neural mechanisms are known. Here we ask what effects mouthbrooding has on several physiological processes including gonad and body mass, brain neuropeptide and receptor gene expression, and circulating steroid hormones in a mouthbrooding cichlid species, Astatotilapia burtoni. We ask whether any observed changes can be explained by food deprivation, and show that during mouthbrooding, ovary size and circulating levels of androgens and estrogens match those seen during food deprivation. Levels of gonadotropin-releasing hormone 1 (GnRH1) mRNA in the brain were low in food-deprived females compared to controls and in mouthbrooding females compared to gravid females. Levels of mRNA encoding two peptides involved in regulating feeding, hypocretin and cholecystokinin, were increased in the brains of food-deprived females. Brain mRNA levels of two receptors, GnRH receptor 2 and NPY receptor Y8c, were elevated in mouthbrooding females compared to the fed condition, but NPY receptor Y8b mRNA was differently regulated by mouthbrooding. These results suggest that many, but not all, of the characteristic physiological changes that occur during mouthbrooding are consequences of food deprivation.     

Variation in plasma leptin levels in response to fasting in Antarctic fur seals (Arctocephalus gazella)

Plasma leptin levels were determined in 8 lactating female and 20 pup Antarctic fur seals (Arctocephalus gazella) during fasting periods of normal duration. Plasma leptin levels ranged from 1.35 3.19 ng x ml(-1) in lactating females and 1.79-4.80 ng x ml(-1) in pups and were not positively correlated with body mass or condition. A negative trend, however, was observed between plasma leptin levels and body condition in lactating females upon their arrival at the colony following a foraging trip (beginning of fast). In accordance with findings in other species, plasma leptin levels dropped significantly (P < 0.02) in response to the 17-19% drop in body mass experienced by pups during fasting. In contrast, plasma leptin levels in lactating females increased during the first 24 h of fasting before decreasing throughout the remaining 48 h of the fast. This unexpected result could be due to the high level of energy expenditure by seals as they swim back to the colony (i.e. post-exercise response) or may be influenced by the intense suckling activity experienced by females during the onshore fasting periods. The results of this study support recent findings in other carnivore species which suggest the primary physiological role of leptin in these species may not necessarily be as a signal of the magnitude of body energy reserves.     

Spring fasting behavior in a marine apex predator provides an index of ecosystem productivity

The effects of declining Arctic sea ice on local ecosystem productivity are not well understood but have been shown to vary inter‐specifically, spatially, and temporally. Because marine mammals occupy upper trophic levels in Arctic food webs, they may be useful indicators for understanding variation in ecosystem productivity. Polar bears (Ursus maritimus) are apex predators that primarily consume benthic and pelagic‐feeding ice‐associated seals. As such, their productivity integrates sea ice conditions and the ecosystem supporting them. Declining sea ice availability has been linked to negative population effects for polar bears but does not fully explain observed population changes. We examined relationships between spring foraging success of polar bears and sea ice conditions, prey productivity, and general patterns of ecosystem productivity in the Beaufort and Chukchi Seas (CSs). Fasting status (≥7 days) was estimated using serum urea and creatinine levels of 1,448 samples collected from 1,177 adult and subadult bears across three subpopulations. Fasting increased in the Beaufort Sea between 1983–1999 and 2000–2016 and was related to an index of ringed seal body condition. This change was concurrent with declines in body condition of polar bears and observed changes in the diet, condition and/or reproduction of four other vertebrate consumers within the food chain. In contrast, fasting declined in CS polar bears between periods and was less common than in the two Beaufort Sea subpopulations consistent with studies demonstrating higher primary productivity and maintenance or improved body condition in polar bears, ringed seals, and bearded seals despite recent sea ice loss in this region. Consistency between regional and temporal variation in spring polar bear fasting and food web productivity suggests that polar bears may be a useful indicator species. Furthermore, our results suggest that spatial and temporal ecological variation is important in affecting upper trophic‐level productivity in these marine ecosystems.     

Histidine decarboxylase gene expression in rat fundus is regulated by gastrin

The conversion of histidine to histamine by histidine decarboxylase (HDC) is of central importance in the control of vertebrate acid secretion. We have used PCR-generated probes to study the regulation of HDC gene expression in rat fundic mucosa. When circulating gastrin levels were lowered by fasting or elevated by treatment with omeprazole, there were parallel changes in HDC mRNA abundance. However, when animals with elevated gastrin levels were concurrently treated with the gastrin/CCK-B receptor antagonist PD 134308, HDC mRNA levels were not increased. These data are consistent with the hypothesis that HDC gene expression is regulated by gastrin, over the physiological range of circulating hormone concentrations.     

Thyroid hormone-dependent gene expression as a biomarker of short-term 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE) exposure in European common frog (Rana temporaria) tadpoles.

The effects on thyroid hormone-dependent gene biomarker responses of the persistent organochlorine pesticide metabolite 1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene (DDE) were investigated after exposure of 4-week-old European common frog (Rana temporaria) (stage 36) tadpoles to two (0.001 and 0.01 ppm) DDE concentrations. Total body weight, total length, and tail length and width increased after 3-day exposure to DDE. Expression patterns of genes encoding for growth hormone, thyroid-stimulating hormone (TSHbeta) and thyroid hormone receptor (TRalpha and TRbeta) isoforms were evaluated in the head, body and tail regions using a validated real-time polymerase chain reaction (PCR) method. The mRNA expression of growth hormone in the body, and TSHbeta in the head showed significant DDE concentration-dependent decreases. While DDE caused variable effects on TRalpha mRNA steady-state, the expression of TRbeta was significantly decreased in the tail by DDE in a concentration-specific manner. The effect of DDE exposure on TRbeta mRNA expression showed a negative correlation with tail length and width during the exposure period. The unique pattern of a DDE-induced decrease of tail TRbeta expression probably reflects the significant role of this thyroid hormone receptor isoform in tail re-absorption and overall metamorphosis in anuran species. Therefore, the present study shows that the evaluation of thyroid hormone-dependent genes may represent quantitative biomarkers of acute exposure to organochlorine pesticides in anuran species during critical developmental periods such as metamorphosis. Given the widespread environmental levels of DDT and its metabolites, these pollutants will remain a subject of concern and their effects on anuran species should be studied in more detail.