Development of thyroid gland and changes in thyroid hormone levels in Leptocephali of Japanese Eel (Anguilla japonica)

Development of the thyroid gland of the Japanese eel (Anguilla japonica) was studied with the use of tank-reared fish. A single thyroid follicle was first found in larvae at 29 days post hatching (dph), total length (TL) 12 mm. Until reaching 25 mm in TL (100 dph), leptocephali had one or two follicles per individual. The inner colloid of the follicles was weakly immuno-positive against the anti-thyroxine (T₄) antibody. The number of thyroid follicles and the immunoreactivity later increased as the larvae grew. Thyroid hormones (TH) T₄ and triiodothyronine (T₃) were not detected in premetamorphic larvae by radioimmunoassays, but became detectable during metamorphosis. The maximum level of T₄ was seen in fish at the end of metamorphosis and in just-metamorphosed juveniles, whereas T₃ reached the highest level during metamorphosis and declined toward the end of metamorphosis. The results indicated that the thyroid gland first became active during metamorphosis in the development of eel.     

Changes in thyroid hormone levels in eggs and larvae and in iodide uptake by eggs of coho and chinook salmon,Oncorhynchus kisutsch andO. tschawytscha

Developmental profiles of thyroxin (T₄), triiodothyronine (T₃) and radioactive iodide uptake were established for eggs and T₄ and T₃ profiles were established for larvae (whole-body, yolk-only and body-only) of coho and chinook salmon. T₄ and T₃ were consistently present in all samples. In eggs, hormone levels remained fairly constant in all cohorst for at least the first three weeks of incubation, but then fluctuated in both directions in some sample groups. Large increases in T₄ (from 9 ng/g to 245 ng/g) were seen in 1985 chinook eggs 28 days after fertilization. Radioactive iodide uptake (which was used as a possible indicator of thyroxinogenesis) increased at least 10-fold in both 1986 coho and chinook eggs from 23–30 days after fertilization. T₄ (62 ng/g) and T₃ (393 ng/g) were found in the bodies of 28-day-old 1986 chinook embryos. In whole larvae, hormone levels varied depending upon the cohort studied. In general, initial body-only concentrations of both T₄ and T₃ decreased as body weight increased, but before yolksac resorption was completed, both thyroid hormone content and concentration increased (except for chinook T₃). T₄ and T₃ content in larval yolk stayed constant as yolksac size decreased, resulting in increased thyroid hormone concentration in the yolksac. All of these data suggest that the initial source of thyroid hormones in coho and chinook salmon eggs is maternal, but that by approximately 3–4 weeks after fertilization, the developing embryos begin to produce their own thyroid hormones. After hatching, increases in tissue T₄ and T₃ concentration coupled with constant T₄ and T₃ content in diminishing yolksacs suggest that larvae also produce their own thyroid hormones; yolksac content then may reflect both the original maternal hormones and the larva-producted hormones.     

Effect of triiodothyronine (T3) in the larval development of the barber goby Elacatinus figaro

The barber goby Elacatinus figaro is an endangered fish, endemic to Brazil and very important to the aquarium trade. One of the bottlenecks for its production in captivity is the larviculture, which is characterized by high mortality rates, especially in the first week after hatching, and prior to metamorphosis. The experiment evaluated the effect of triiodothyronine hormone (T₃) on survival, growth and metamorphosis of barber goby. Larvae of 14 days after hatching (DAH) were immersed in three concentrations of T₃, in triplicate: TC (control) – 0; T01 – 0.01; T025 – 0.025 and T05 – 0.05 mg/L. An additional replicate of each treatment was performed for sampling for the histological analysis of the thyroid follicles in 24 DAH larvae. The survival rates in TC, T01, T025 and T05 were 24%, 54%, 36% and 37% respectively, without significant differences between the treatments (p > 0.05). Regarding larval growth, the highest length values were obtained in TC. In T025 and T05, larval metamorphosis was anticipated in up to 11 days in relation to TC and T01. In T01 larvae, follicles were numerous with the presence of reabsorption vesicles in the colloid periphery, indicating increased production of thyroid hormones (THs), associated with the larval metamorphosis process. In T05, a follicle reduction in number and size was observed, given evidence of the end of metamorphosis. The use of 0.025 and 0.05 mg/L of T₃ anticipated metamorphosis in barber goby larvae, shortening the larviculture period for this species.     

Thyroid gland development in Senegalese sole (Solea senegalensis Kaup 1858) during early life stages: A histochemical and immunohistochemical approach

A key to success in the culture of marine fish species is the mass production of high quality fry, a process largely dependent on successful first feeding and normal development and growth of fish larvae. In this regard it is important to examine the structural and functional development of the endocrine system (pituitary, thyroid, interrenal glands) during early ontogeny of marine fish. Particularly, the thyroid hormones, thyroxine (T₄) and triiodothyronine (T₃), influence numerous metabolic processes, such as growth, differentiation, metamorphosis, reproduction, respiration, migratory behaviour, central nervous system activity, seasonal adaptation, etc. Therefore the aim of this study was to describe the development of the thyroid gland and the ontogeny appearance of the thyroid hormones in Solea senegalensis larvae by means of histological and immunohistochemical techniques. The first thyroid follicle was present at 4 days-post-hatch (dph) coinciding with first feeding. During metamorphosis (12–20 dph) the follicles increased in both number and size, and by 30 dph presented the same characteristics as that seen in adult fish. Tissue immunostaining of both thyroid hormones decreased during the endogenous larvae development to nearly undetectable levels at the completion of yolk-sac absorption. During larvae exogenous phase, T₃ and T₄ immunostaining was first detected by 6 dph and an increase of specific staining for both hormones was detected between 12 and 20 dph, during metamorphosis phase.     

Intrafollicular thyroid hormone staining in whole-mount zebrafish (<Emphasis Type="Italic">Danio rerio</Emphasis>) embryos for the detection of thyroid hormone synthesis disruption

Endocrine-disrupting chemicals are known to impact multiple hormonal axes of vertebrates, among which the thyroid system is crucial for multiple developmental and physiological processes. Thus, the present study focused on the semi-quantitative visualization of intrafollicular triiodothyronine (T3) and thyroxin (T4) in zebrafish embryos as a potential test system for the detection of disrupted thyroid hormone synthesis. To this end, an antibody-based fluorescence double-staining protocol for whole-mount zebrafish embryos and larvae was adapted to simultaneously detect intrafollicular T3 and T4. During normal development until 10 days post-fertilization (dpf), the number of thyroid follicles increased along the ventral aorta. Concentrations of T4 and T3, measured by fluorescence intensity, increased until 6 dpf, but decreased thereafter. Exposure of zebrafish embryos to propylthiouracil (PTU), a known inhibitor of TH synthesis, resulted in a significant decrease in the number of follicles that stained for T3, whereas a trend for increase in follicles that stained for T4 was observed. In contrast, fluorescence intensity for both thyroid hormones decreased significantly after exposure to PTU. Overall, the zebrafish embryo appears to be suitable for the simultaneous visualization and detection of changing intrafollicular TH contents during normal development and after PTU treatment.     

鳙、团头鲂和短盖巨脂鲤早期发育阶段甲状腺激素含量变化研究

鳙、团头鲂和短盖巨脂鲤胚胎与仔鱼体内三碘甲腺原氨酸和四状腺素含量的变化，采用放射免疫测定法进行测定，研究发现胚胎阶段的Ｔ３和Ｔ４含较稳定，出膜后仔鱼阶段的Ｔ３和Ｔ４含量逐渐下降。     

Coordinated expression and regulation of deiodinases and thyroid hormone receptors during metamorphosis in the Japanese flounder (<Emphasis Type="Italic">Paralichthys olivaceus</Emphasis>)

In vertebrates, thyroid hormone receptors (TRs) and deiodinases are essential for developmental events driven by the thyroid hormones (THs). However, the significance of deiodinases during the metamorphosis of the Japanese flounder (Paralichthys olivaceus) remains unclear. Moreover, regulation and response of the TRs and deiodinases to THs in this fish are poorly understood. Therefore, we detected the expression patterns of THs, deiodinases, and TRs in drug-treated larvae and untreated larvae of P. olivaceus by using enzyme-linked immunosorbent assay and quantitative real-time PCR during P. olivaceus metamorphosis. To further understand the roles of these elements, a rescue assay was performed. Our results show the importance of THs, TRs, and deiodinases in flatfish metamorphosis. Our results also confirm that D1 and D2 activate THs and D3 plays the opposite and complementary role. Moreover, we demonstrated that both TRα and TRβ have important but different roles during P. olivaceus metamorphosis.     

Stimulatory effect of thyroid hormones improves larval development and reproductive performance in alligator gar (Atractosteus spatula) and spotted gar (Lepisosteus oculatus)

Development of alligator gar (Atractosteus spatula) and spotted gar (Lepisosteus oculatus) larvae was evaluated after exposure of embryos to thyroid hormones (TH) by two different methods of administration. For the first method, alligator gar embryos were placed in a water bath with triiodothyronine (T₃; 1 ppm for 2 h), and control treatment embryos were placed in a water bath without T₃. For the second method, spotted gar adult males and females were injected with different TH during induced spawning: a group was injected with T₃ [20 mg kg^?1 body weight (BW)], a second group with thyroxine (T₄; 20 mg kg^?1 BW), a third group with thyroid‐stimulating hormone (TSH; 4 IU kg^?1BW) and a control group with dimethyl sulfoxide. Both methods resulted in increases in the concentrations of T₃ and T₄ in embryos of both species. An increase in the alkaline enzymatic activity of TH‐treated larvae was observed as well as an increase in the RNA:DNA, RNA:Dry weight and DNA:Dry weight ratios at hatching. Some positive effects of TH application were: a significant increase in the hatching rate of spotted gar larvae from TH‐injected parents and a higher percentage of successful spawns. Evenness of larval sizes and a higher survival rate were observed in alligator gar larvae. Snout development was accelerated by TH treatment in both species.     

Ontogeny of thyroid tissue and tissue thyroid hormone clearance in rainbow trout embryos reared at two temperatures

The development of the thyroid tissue and clearance of thyroid hormones was studied in rainbow trout embryos reared at 8.5 and 5.5 °C, in order to determine if these events are affected by temperature in the same manner as growth rate. Although the change in growth rate was found to be predictable using the degree-day model, the pattern of whole body thyroid hormone clearance was not similarly related to rearing temperature. Further, although the thyroid follicles appeared at different times in embryos reared at the two temperatures, the differences could not be predicted using the degree-day model. These results suggest that the development of the thyroid tissue and the clearance of yolk hormones are not directly related to rearing temperature. Immunohistochemical staining of the thyroid tissue indicated two phases of thyroid activity, one associated with hormonogenesis, and the other a later phase of hormone release. However, there was no increase in 'whole body' thyroid hormone level associated with the time of appearance of colloid in the follicles. Further, evidence was presented to suggest that the extraction method used did not remove thyroid hormone associated with the colloid in the follicles. There was also evidence of sporadic increases in 'whole body' thyroid hormone content, after the development of phase 2 thyroid follicles which might indicate pulsatile release of T4 and T3.     

Flounder metamorphosis: its regulation by various hormones

Metamorphosis in the flounder has often been compared with the transition of tadpoles into frogs. The dorsal fin rays of the Japanese flounder (Paralichthys olivaceus) elongate during prometamorphosis when thyroid hormone levels are low, and are resorbed during metamorphic climax when thyroid hormone levels are high. Using an in vitro system for the culture of the flounder fin rays, we have examined how various hormones affect the resorption process. Both thyroxine (T₄) and triiodothyronine (T₃) directly stimulated fin ray shortening, T₃ being more potent than T₄. Other hormones, such as prolactin, cortisol and sex steroids, did not directly affect the resorption process but modified the tissue's response to thyroid hormones. Similar observations were obtained from in vivo studies. We also monitored the changes in the whole body concentrations of various hormones during early development and metamorphosis, and related these with the thyroid hormone profiles in order to get a better picture of their interactions. The gaps in the present status of research on the role of thyroid hormones during metamorphosis in the Japanese flounder are also discussed.     

Thyroid hormone deficiency in abnormal larvae of the Japanese flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis>

ABSTRACT:   In large-scale rearing of juveniles of the Japanese flounder Paralichthys olivaceus, a certain morphological abnormality occurred spontaneously in 4% of the fish. These fish showed a slight but clearly different appearance from any developmental stage of this species, and did not settle when all the other juveniles in the same tank completed metamorphosis and had settled. From comparisons of external and internal structures between the normal and the abnormal fish, the abnormality was attributed to unbalanced progress of metamorphosis, mainly due to metamorphic stasis. The thyroid of the abnormal fish was apparently activated morphologically. In addition, serum thyroxine (T4) concentrations in the abnormal fish were reduced to less than 1/10 of that of normal fish. After 14 days of T4 treatment (0.1 p.p.m) of the abnormal fish, all the abnormal characteristics disappeared, and the fish recovered to normal, suggesting normal responsiveness to thyroid hormones in peripheral tissues, whereas thiourea treatment (30 p.p.m., 14 days) further delayed metamorphosis. These results suggest that these abnormal fish were suffering from thyroid hormone deficiency, and were unable to secrete a sufficient amount of thyroid hormone to complete metamorphosis.     

Insulin-like growth factor binding protein-2 (IGFBP-2) in Japanese flounder, Paralichthys olivaceus: molecular cloning, expression patterns and hormonal regulation during metamorphosis

In this study, we cloned and characterized cDNA sequences of two insulin-like growth factor binding protein-2 (IGFBP-2a and IGFBP-2b) from Japanese flounder, Paralichthys olivaceus. The full-length cDNA of IGFBP-2a is 1,046 bp long and consists an open frame (ORF) of 876 bp, a 5′-untranslated region (UTR) of 125 bp and a 3′-UTR of 45 bp. IGFBP-2b is 1,067 bp, including a 5′-UTR of 53 bp, a 3′-UTR of 198 bp and an ORF of 816 bp. Real-time quantitative PCR results revealed that IGFBP-2a -2b mRNA was expressed in all detected tissues. Interestingly, the levels of IGFBP-2a mRNA in all detected tissues were higher in female than male, but IGFBP-2b was precisely the opposite. At different embryonic stages, the levels of IGFBP-2a mRNA were typically higher than IGFBP-2b. After hatching, IGFBP-2a mRNA was gradually decreased to a relatively lower level. However, the expression of IGFBP-2b mRNA was increased after hatching, including 3, 7, 10, 14, 17, 20 and 23 days post-hatching (dph), and it presents a higher level until 29 (metamorphic climax), 36 (post-climax) and 41 dph (the end of metamorphosis). In levothyroxine sodium salt (T4, the main form of thyroid hormone in animals)-treated and thiourea (TU)-treated larvae, the expressions of IGFBP-2a had not visibly changed, except in T4-treated 17 dph larvae. The expressions of IGFBP-2b mRNA were distinctly increased from 17 to 23 dph, but suddenly dropped to a lower level in and after 29 dph. However, the levels of IGFBP-2b mRNA during metamorphosis were greatly down-regulated after TU treatment. These results provided basic information for further studies on the role of IGF system in flatfish development and metamorphosis.     

Salinity and temperature effects on whole-animal thyroid hormone levels in larval and juvenile striped bass,Morone saxatilis

Whole-animal thyroxine (T₄) and 3,5,3′-triiodothyronine (T₃) levels were measured in larval and juvenile striped bass, Morone saxatilis, reared for 10 days at one of three levels of salinity (equivalent to fresh water (FW), one-third seawater (1/3 SW), and seawater (SW) and two temperatures (15°C and 20°C). The striped bass were pre-metamorphic larvae, metamorphic larvae or juveniles. The short-term effects of seawater on plasma T₄ levels of juvenile striped bass were also measured. Higher salinities increased T₄ levels in premetamorphic larvae. In metamorphic larvae, SW and 1/3 SW increased T₄ levels and SW increased T₃ levels at 20°C. This response was eliminated in those at 15°C. Whole-animal thyroid hormone content was unaffected by salinity or temperature in juvenile striped bass, although significant fluctuations in plasma T₄ levels occurred in those transferred to 1/3 SW and SW. The thyroid axis of striped bass responds to salinity and temperature as early as in the pre-metamorphic stage. Thyroid hormones may mediate the beneficial effects of salinity on larval striped bass growth and survival.     

Effects of thyroxine and thiourea on the metamorphosis of coral trout grouper Plectropomus leopardus

The contribution of the thyroid hormone to the metamorphosis of the coral trout Plectropomus leopardus was examined. Juveniles, aged 35 days, were exposed to thyroxine (T4, 0.1 p.p.m) and thiourea (TU; inhibitor of thyroid hormone synthesis, 30 p.p.m) for 13 days. All the fish in the T4-treated group had completed metamorphosis (second metamorphosis, pigmentation and resorption of fin spines) within three days after the beginning of the experiment (day 3), whereas it took 13 days for this to occur in the control fish. Moreover, the fish in the TU group had not completed metamorphosis even by day 13. Settling behavior was significantly stimulated by T4 treatment, as was the resorption of the dorsal and pelvic fin spines, the reduction of serration on the spines, and the transition of body color into an opaque reddish hue. Fish in the TU group had longer dorsal and pelvic spines, retarded pigmentation and an abnormal black coloration. These results suggest that thyroid hormone accelerates most changes during the second metamorphosis in the coral trout.     

Transpositional feeding rhythm of loach <Emphasis Type="Italic">Misgurnus anguillicaudatus</Emphasis> from larvae to juveniles and its ontogenesis under artificial rearing conditions

The diel feeding rhythm and ontogenesis during early life stage of loach Misgurnus anguillicaudatus were investigated under experimental conditions (light: L 06:00–18:00, D 18:00–06:00 h). Morphological and behavioral developments of loach from newly hatched to 40 days after hatching were observed. Larvae were able to prey on daphnia 3–4 days after hatching at 23 ± 0.5°C. As the larvae grew, they showed an increasing feeding capacity and a distinct feeding rhythm. Feeding intensity and incidence for day-4 larvae were highest at 10:00 and 16:00 h. The highest levels of feeding intensity for day-12 larvae occurred at 08:00, 12:00, and 18:00 h as did feeding incidence. By day 20, when the larvae metamorphosed, the highest levels of feeding intensity occurred at 06:00, 18:00, and 24:00 h and were concurrent with the highest feeding incidence. After metamorphosis, feeding capacity had again increased considerably and, in contrast to the earlier stages before day 20, feeding intensity for day-30 juveniles peaked at 05:00 and 20:00 h, about 1–2 h after the maximum feeding incidence. The feeding rhythm of loach juveniles at day 40 was almost the same as the day-30 juveniles. The estimated maximum daily feeding rates were 43.1%, 33.4%, 19.0%, 12.8%, and 5.8% of body weight on days 4, 12, 20, 30, and 40, respectively. Thus, loach was found to have different feeding rhythms in the pre- and post-metamorphosis stages, with the highest feeding activity in daytime during the larval planktonic stage before metamorphosis, and intensely nocturnal feeding behavior during the juvenile benthic stage after metamorphosis.     

Role of thyroid hormones in tilapia larvae (Oreochromis mossambicus): I. Effects of the hormones and an antithyroid drug on yolk absorption,growth and development

Treatment of one-day old yolksac larvae of tilapia (Oreochromis mossambicus) by immersion in 0.05 ppm T₄ or 0.01 ppm T₃ significantly accelerated the differentiation and growth of all the fins, particularly pectoral and tail fins. Both the treatments also significantly accelerated yolk absorption and transition to free-swimming activity in the larvae. The treatments also significantly accelerated the growth of the larvae, with T₃ at 0.01 ppm having a greater effect than T₄ at 0.05 ppm. The yolk conversion efficiency was found not to be significantly affected by the hormone treatments but the treated larvae exhibited an increased heart beat, suggesting metabolic stimulation by the hormones.On the other hand, yolk absorption and free-swimming activity were significantly delayed in tilapia larvae immersed in 25 ppm solution of an antithyroid drug, phenylthiocarbamide (PTC). PTC also retarded the growth of the larvae. T₄ (0.05 and 0.10 ppm) or T₃ (0.01 and 0.02 ppm) therapy removed the PTC-inhibition,albeit not completely, suggesting that thyroid hormones are involved in the larval growth and development of tilapia.     

Effects of different step‐wise temperature increment regimes during egg incubation of Atlantic cod (Gadus morhua L.) on egg viability and newly hatched larval quality

We carried out an experiment to determine how rapidly the early incubation temperature of Atlantic cod eggs can be increased without affecting normal embryonic development and hatching. Atlantic cod eggs were incubated at a constant low temperature (4.5 ± 0.5°C; T5 – control) and four temperature increment treatments where the temperatures were increased stepwise from 4.5°C at zygote stage to 9.5 ± 05°C (T1‐8 h, T2‐32 h, T3‐64 h and T4‐96 h). Embryonic cell symmetry, embryonic mortality, hatching success and larval skeletal abnormalities, length and yolk sac volume were recorded. Larval samples were also taken at hatch for histological analysis. Except for higher egg mortality and lower hatching success in the T1, the differences among experimental groups were minor. Cell asymmetries and embryo mortalities were not significantly different between the control and T2–T4 treatment groups. Control larvae were significantly longer and had smaller yolk reserves at hatch than T1–T4 larvae and larvae from T2 had the largest yolk reserves. Tissue and organ histology of hatched larvae were similar. Considering embryonic cleavage pattern, hatching success and larval morphology and histology, a gradual increment of temperature in 32 h seems to be the better choice for future developmental programming studies in Atlantic cod.     

牙鲆NR4A1基因在甲状腺素诱导仔鱼变态中的表达调控分析

NR4A1基因是一个重要的转录因子,在信号转导早期通过对靶基因的特异性转录调控发挥着重要的作用。从牙鲆(Paralichthys olivaceus)中克隆了NR4A1基因c DNA序列,检测了其在牙鲆变态中和成鱼各组织中的表达模式,分析了其在外源甲状腺素(TH)以及硫脲(TU)处理仔鱼中的表达变化。结果表明,牙鲆NR4A1 c DNA全长3264 bp,编码568个氨基酸;牙鲆NR4A1基因与其它物种具有很高的同源性,在系统进化树中与鱼类聚为一支;牙鲆NR4A1基因在成鱼心、肌肉和鰓中高表达;在变态过程中,NR4A1水平逐渐升高,且在25 dph达到最高,之后逐渐降低;TH组中的NR4A1基因水平在变态早期和高峰期显著低于正常组,TU组的水平在变态中后期和结束期显著高于正常组;TU组中变态被抑制的仔鱼在正常海水和0.1 mg·L~(-1)TH海水中饲养6 d后,能够顺利变态,且NR4A1基因在拯救组(TU抑制变态仔鱼在正常海水和0.1 mg·L~(-1)TH海水中饲养)中的表达水平与TU对照组相比显著降低,与正常组中同时期的水平一致。结果表明,牙鲆NR4A1基因可能在仔鱼变态调控中扮演着非常重要的角色。     

Pigmentation and eye migration in Atlantic halibut (Hippoglossus hippoglossus L.) larvae: new findings and hypotheses

Atlantic halibut juveniles, which have been fed Artemia during larval development, frequently demonstrate malpigmentation and impaired eye migration. This is in contrast to the high percentage of normally developed larvae fed copepods, reared under similar conditions. Nutrition is therefore an important component influencing larval development. Analyses of the nutrient composition of Artemia and copepods show that Atlantic halibut larvae fed Artemia probably receive sufficient amounts of vitamin A by converting canthaxanthin, while iodine may be deficient, possibly leading to interrupted thyroid hormone synthesis. An unbalanced fatty acid composition, such as high levels of arachidonic acid and low levels of docosahexaenoic acid, can be another limiting factor in Artemia. Vitamin A, fatty acids and thyroid hormones have all been shown to affect pigmentation in flatfish. They are ligands to nuclear receptors, thyroid hormone receptors, retinoic acid receptors, retinoic X receptors and peroxisomal proliferator‐activated receptors, which are members of the superfamily of steroid hormone receptors. The receptors interact with each other to promote gene expression that modulates proliferation and differentiation of cells. Our hypothesis is that these interactions are important for development during flatfish metamorphosis. Very little data exist on the topic of impaired eye migration. However, energy limitation, iodine deficiency and an unbalanced fatty acid composition have been proposed as possible explanations. Here, we review the literature on development of pigment cells and the possible mechanisms behind the effects of vitamin A, fatty acids and thyroid hormone on pigmentation and eye migration during development of Atlantic halibut larvae.