Thyroid status is one of the most potent regulators of peripheral thyroid hormone metabolism in vertebrates. Despite this, the few papers that have been published concerning the role of thyroid hormones in the regulation of thyroid function in fish often offer conflicting data. We therefore set out to investigate the effects of tetraiodothyronine (thyroxine) (T4) or tri-iodothyronine (T3) supplementation (48 p.p.m.) via the food on plasma and tissue thyroid hormone levels as well as iodothyronine deiodinase (D) activities in the Nile tilapia (Oreochromis niloticus). T4 supplementation did not induce a hyperthyroid state and subsequently had no effects on the thyroid hormone parameters measured, with the liver as the sole notable exception. In T4-fed tilapias, the hepatic T4 levels increased substantially, and this was accompanied by an increase in in vitro type I deiodinase (D1) activity. Although the lack of effect of T4 supplementation could be partially explained by an inefficient uptake of T4 from the gut, our current data suggest that also the increased conversion of T4 into reverse (r)T3 by the D1 present in the liver plays an important role in this respect. In addition, T3 supplementation increased plasma T3 and decreased plasma T4 concentrations. T3 levels were also increased in the liver, brain, kidney, gill and white muscle, but without affecting local T4 concentrations. However, this increase in T3 availability remained without effect on D1 activity in liver and kidney. This observation, together with the 6-n-propylthiouracyl (PTU) insensitivity of the D1 enzyme in fish, sets the D1 in teleost fish clearly apart from its mammalian and avian counterparts. The changes in hepatic deiodinases confirm the role of the liver as an important T3-regulating tissue. However, the very short plasma half-life of exogenously administered T3 implies the existence of an efficient T3 clearing/degradation mechanism other than deiodination.

The effects of the dioxin-like polychlorinated biphenyl (PCB) 77 and the ortho-substituted PCB 153 on thyroid hormone availability were investigated during the last week of embryonic development in chicken. High doses of these PCBs (1μg PCB 77 and 20μg PCB 153) were injected into chicken eggs at day 4 of incubation. Blood and tissue samples were collected from day 14 of incubation until 1 day after hatching. We did not observe influences of PCB 153 on thyroid hormone (TH) levels. Treatment with PCB 77, on the other hand, decreased plasma total T4 concentrations but increased hepatic T4 levels at day 14 of incubation. Later in development, at stages near the process of hatching, severe decreases of T4 and T3 levels were observed in the PCB 77 group, both in plasma and tissues. PCB 77 severely reduced the TH peak that normally coincides with the stage of internal pipping. This reduction was accompanied by a considerable delay in the moment of hatching. We conclude that the dioxin-like PCB 77, but not the ortho-substituted PCB 153, can decrease TH availability towards the end of embryonic development and hence disturb the process of hatching.

Glucocorticoids as well as thyroid hormones are essential for normal brain development. Exogenous glucocorticoids stimulate 3,3′,5-triiodothyronine (T3) availability in circulation of birds and similar effects have been observed in sheep. Chicken data indicate that glucocorticoid administration also stimulates thyroid hormone metabolism in brain but the effects on local thyroid hormone concentrations are not known. Therefore, the current study: (1) determined local thyroid hormone availability in separate brain areas of 18-day-old embryonic chickens (E18) after injection of dexamethasone (DEX), and (2) investigated the impact on the thyroid hormone metabolic pathways in these brain parts and compared the results with the hepatic situation. For this, E18 chicken embryos were treated with a single intravenous dose of DEX (25μg). Despite the decreased 3,5,3′,5-tetraiodothyronine (T4) availability in the liver of the DEX treated embryos, the T3 content was strongly increased, parallel to the plasma T3 surge. This T3 surge was primarily related to a fall in hepatic T3 breakdown through a downregulation of the type III deiodinase (D3). The sulfation pathway in liver seems not to be affected by DEX. In all brain parts, DEX affects the T3 production capacity by upregulation of the type II deiodinase (D2). This enables the brain to compensate for the decrease in T4 availability, although the T3 concentrations are not consistently increased like in plasma and liver. This observation points to the existence of a fine-tuning mechanism in brain that enables the brain to keep the T3 concentrations within narrow limits.

A search of the chicken expressed sequence tag (EST) database identified 2 cDNA clones that appeared to represent members of the SULT1B and SULT1C enzyme families. These cDNAs were fully sequenced and found to contain full-length inserts. Phylogenetic analysis of the derived amino acid sequences clearly placed them as the first members of the chicken SULT1B and SULT1C families, respectively, to be identified, and we propose they be named SULT1B1 and SULT1C1. (CHICK)SULT1B1 shares approximately 60% amino acid sequence identity with mammalian SULT1B enzymes, whereas the closest neighbor to (CHICK)SULT1C1 was the ortholog (RAT)SULT1C1, with 68% identity. We cloned these cDNAs into the bacterial expression vectors from the pET series. Transformed Escherichia coli cells strongly expressed the recombinant proteins. Purification of the recombinant enzymes from E. coli was accomplished by a three-step procedure involving ammonium sulfate precipitation, anion exchange chromatography, and affinity chromatography. The purified enzymes displayed subunit molecular weights of approximately 35,000Da on SDS–PAGE, as predicted, and were both able to sulfate a wide range of compounds, including xenobiotics and endogenous substrates such as iodothyronines. Detailed kinetic analysis showed SULT1C1 was more prolific in that it was able to sulfate dopamine, tyramine, and apomorphine, which SULT1B1 was not. 2-Bromophenol was the best substrate for both enzymes. We also raised antibodies against these proteins, which were able to detect the SULTs by ELISA, and which were able to strongly inhibit the recombinant enzymes. This is the first detailed characterization of sulfotransferases from the chicken, and it demonstrates that the avian and mammalian SULT1 enzymes are closely related in both structure and function.

The intracellular thyroid hormone (TH) availability is influenced by different metabolic pathways. We investigated the relationship between tissue and plasma TH levels as well as the correlation with changes of deiodination and sulfation during chicken embryonic development. From day 14 until day 19, T3 remains unchanged in liver and kidney in spite of increasing plasma T4 and T3 levels and a slightly increased T4 availability in these tissues. During this period, the T3 breakdown capacity by type III deiodinase (D3) is high in liver but low in kidney. The TH inactivation capacity of type I deiodinase (D1), with production of inactive rT3 instead of T3, in kidney seems to be potentiated by the sulfation pathway. A sharp rise in T3 and T4 is detected in all tissues examined when the embryo switches to lung respiration. The same day, T4 content in liver is sharply enhanced and sulfation activity is decreased. So, T4 availability in liver is increased while a declined D3 activity allows for the accumulation of hepatic T3. The increase in renal T3 and T4 are more closely related to plasma TH profiles and a lack of correlation with the changes in renal D1 and D3 activity suggests that T4 and T3 content in this organ is strongly dependent on direct uptake from the blood. Despite much lower T4 levels, T3 levels in brain are in the same range as in liver and kidney and intracellular T3 even exceeds the T4 levels towards the end of development. The rise in TH content coincides with a drop in D3 activity, low sulfation activity and an increased T3 production capacity via type II deiodinase (D2). In conclusion, the current study describes the dynamics of intracellular TH concentrations in liver, kidney, and brain during chicken development and investigates their relationship with circulating TH levels and changes of deiodinases and sulfotransferases. The clear differences in intracellular TH profiles among the different tissues demonstrate that circulating levels are not necessarily representative for the local TH changes. Some of the changes in intracellular TH availability can be linked to changes in local deiodination and sulfation capacities, but the importance of these enzyme systems in relation to other factors, such as hormone uptake, differs between liver, kidney, and brain.

The relationships among avian uncoupling protein (avUCP) mRNA expression, heat production, and thyroid hormone metabolism were investigated in 7–14-day-old broiler chicks (Gallus gallus) exposed to a low temperature (cold-exposed chicks, CE) or a thermoneutral temperature (TN). After 7 days of exposure, CE chicks exhibited higher heat production (+83%, P<0.01), avUCP mRNA expression (+20%, P<0.01), and circulating triiodothyronine (T3) levels (+104%, P=0.07) for non-statistically different body weights and feed intake between 3 and 7 days of exposure as compared to TN chicks. Plasma thyroxine (T4) concentration was clearly decreased in CE chicks (−33%, P=0.06). The lower hepatic inner-ring deiodination activity (−47%) and the higher renal outer-ring deiodination activity (+75%) measured in CE compared to TN chicks could partly account for their higher plasma T3 concentrations. This study describes for the first time the induction of avUCP mRNA expression by low temperature in chickens, as it has been previously shown in ducklings, and supports the possible involvement of avUCP in avian thermogenesis.

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.

Next to deiodination, glucuronidation and sulfation of the phenolic hydroxyl group are important pathways of iodothyronine metabolism. Although in general conjugation serves to facilitate the urinary and biliary excretion of lipophilic substances, sulfation of thyroid hormones has another important function. Sulfation protects the tissues against an excess of active thyroid hormone and it can also be used as a reservoir from which active hormone is released by sulfatases when and where it is required. Interaction between sulfation and deiodination has already been described in several mammalian tissues. After an initial characterization of the sulfotransferases we determined the changes in sulfation during chicken embryonic development and induced metamorphosis of the axolotl, a neotenous amphibian. Sulfation assays demonstrate the presence of sulfotransferases in liver, kidney and brain cytosol in one-day-old chicken. Similar to the situation in mammals, the enzyme(s) show a substrate preference for 3,3'-diiodothyronine (3,3'-T2) over 3,5,3'triiodothyronine (T3) and 3,3',5'-triiodothyronine (rT3) over thyroxine (T4). Measurements of sulfation using 3,3'-T2 as substrate in tissues from 16-day-old embryonic chickens (E16) to one-day-old posthatch chicks (C1) demonstrate that sulfation activity varies independently in liver, kidney and brain, suggesting that the activity is regulated in a tissue specific manner. We also detected sulfation activity in liver, kidney and brain of the axolotl. The highest activity was found in the liver, hence we used this tissue to study the profile of activity during induced metamorphosis. We found that the activity in the liver remains stable during the full length of metamorphosis.