Direct influence of melatonin on the thyroid and comparison with prolactin

Melatonin administered in vivo had previously been shown to inhibit thyroid cell proliferation and subsequent in vitro thyroxine (T(4)) secretion in anuran tadpoles. Melatonin in vitro also directly reduced the sensitivity of the thyroid to thyrotropin (TSH). The present work sought to determine whether melatonin directly affected baseline, unstimulated T(4) secretion, and to compare its effect with that of prolactin (PRL). Thyroids from larval Rana catesbeiana or adult Rana pipiens were incubated in control or melatonin (0.01 to 100 microg/ml) media. Melatonin directly inhibited T(4) secretion by thyroids from both tadpoles and frogs at all concentrations of melatonin used and at both prometamorphic and climax tadpole stages. PRL, used in vitro at 10 microg/ml, did not influence the response of the thyroid to TSH (0.2 microg/ml) in young tadpoles, or the baseline secretion of T(4) by thyroids at any stage of larval life except climax, when T(4) secretion was significantly decreased by the third day of culture. Thus although both melatonin and PRL have been shown to antagonize the action of T(4) in vitro, and to decrease metamorphic rate, melatonin is a much more effective thyroid gland inhibitor than PRL.     

Influence of Melatonin on the Rate of Rana pipiens Tadpole Metamorphosis In Vivo and Regression of Thyroxine-Treated Tail Tips In Vitro

Metamorphosis of Rana pipiens tadpoles may be retarded when the light phase of the light/dark (LD) cycle is shortened or when thyroxine (T₄) is given in the dark because melatonin peaks during the dark. Injection of premetamorphic tadpoles in spontaneous metamorphosis with melatonin (15 μg) retarded tail growth and hindlimb development on 18L:6D but had no significant effect on 6L:18D. During induced metamorphosis (30 μg/liter T₄), melatonin injections retarded tail resorption on 18L:6D and accelerated it on 6L:18D, but did not affect the hindlimb. When melatonin was injected during T₄ immersion at different times in the photophase on 18L:6D (L onset 0800 hr), tail regression was retarded by melatonin at 1430 or 2030 hr. At 0830 hr, shrinkage of tail length was accelerated whereas tail height was not affected. Tail tips in vitro induced to resorb by 0.2 μg/ml T₄ in Niu-Twitty solution regressed more slowly in the presence of melatonin (10 or 15 μg/ml) than with T₄ alone on both 6L:18D and 18L:6D. The findings implicate melatonin in LD cycle effects on tadpole metamorphic rate in vivo , show the importance of the time of melatonin injections, and indicate that melatonin antagonizes the metamorphic action of T₄ at the tissue level.     

Adenohypophysial-thyroid activity of the tiger salamander, Ambystoma tigrinum, as a function of metamorphosis and captivity

The aim of this study was to determine the timing of adenohypophysial activation during metamorphosis of the tiger salamander, Ambystoma tigrinum. It consisted of two parts: 1) determination of plasma thyroid hormone concentrations and analysis of thyroid gland histology as a function of metamorphic stage and 2) analysis of the time-course of uptake of 125I by the thyroids during metamorphosis as an indicator of endogenous thyrotropin (TSH) levels. Significant increases in both triiodothyronine (T3) and thyroxine (T4) first were evident at the onset of metamorphic climax (stage II). Maximum levels of both hormones were not observed, however, until the completion of gill resorption (stage VII). No changes in thyroid histology were observed that could be unambiguously related to metamorphic transformation. The thyroids accumulated 125I in a slow but linear fashion in premetamorphic larvae (stage I). However, uptake exhibited a rapid peak during early climax (stage II), before maximum concentrations of thyroid hormones were observed. In addition, uptake was maintained above premetamorphic levels at stage VII, in conjunction with maximum levels of T4 and T3. Captivity alone produced a small but significant increase in plasma concentrations of T3. It produced no significant effect on either thyroid histology or uptake of 125I. These results indicate that adenohypophysial activation occurs rapidly and is maximal at the onset of metamorphic climax.     

Hormonal Interaction in Amphibian Metamorphosis

SYNOPSIS. The climactic stages of amphibian metamorphosis constitutea period characterized by radical morphological changes thatare driven primarily by the thyroidal hormones. Radioimmunoassaysshow that levels of thyroid hormones (TH) rise to a peak duringmetamorphic climax. Accompanying peaks are reported for ACTH,adrenal corticoids (AC), insulin (I) and prolacdn (PRL). ACenhance the metamorphic action of TH by increasing their bindingto nuclei of target cells. TH, in turn, act to raise levelsof AC by stimulating the differentiation of the median eminencethus facilitating the flow of a CRF from the hypothalamus tothe adenohypophysis, by synergizing with ACTH and bystimulationof the interrenals through some other route. During the metamorphicperiod, at least as far as climax, PRL antagonizes TH, perhapsat the thyroidal level and certainly at the level of targetcells. PRL may antagonize by inhibiting induction of hydrolyticenzymes by TH, by alteration of hydromineral responses or byaltering levels of binding of TH to receptors. The antagonisticaction of PRL is mimicked by cAMP. A surge of PRL that is releasedinto the plasma during metamorphic climax seemingly producesno antagonistic effect on thyroidal actions.     

An analysis of the influence of thyroid hormone on the synthesis of proteins in the tail fin of bullfrog tadpoles

By incubation of explants of tail fin from tadpoles of Rana catesbeiana in a solution of 35S-methionine for 4 h, newly synthesized proteins were labeled isotopically. After separation by two-dimensional polyacrylamide gel electrophoresis, those proteins were visualized by fluorography. Exposure of explants to culture medium containing thyroxine (T4) (150 nM) increased the incorporation of 35S-methionine into several proteins with 48 h. Effects of T4 on the relative abundance of two of these newly synthesized proteins were detected after 8 h of hormonal treatment. Very similar patterns of newly synthesized proteins were observed when proteins from explants of tail fin removed from tadpoles at metamorphic climax and immediately incubated with 35S-methionine were compared with proteins produced in fin derived from premetamorphic animals. These results are interpreted to indicate that both treatment of explants with T4 and elevation of endogenous levels of thyroid hormones during spontaneous metamorphosis increased the relative rates of synthesis of several identical proteins. The potential involvement of those proteins in early phases of metamorphic action which eventually lead to cell death and resorption is discussed.     

Programmed cell death and heterolysis of larval epithelial cells by macrophage-like cells in the anuran small intestine in vivo and in vitro.

The degenerative processes in the larval small intestine of Xenopus laevis tadpoles during spontaneous metamorphosis and during thyroid hormone-induced metamorphosis in vitro were examined by electron microscopy. Around the beginning of spontaneous metamorphic climax (stages 59-61), both apoptotic bodies derived from larval epithelial cells and intraepithelial macrophage-like cells suddenly increase in number. The macrophage-like cells become rounded and enlarged because of numerous vacuoles containing the apoptotic bodies. Mitotic profiles of the macrophage-like cells, however, are localized in the connective tissue where different developmental stages of macrophage-like cells are present. After stage 62, the intraepithelial macrophage-like cells decrease in number, while large macrophage-like cells which include the apoptotic bodies and retain intact cell membranes and nuclei appear in the lumen. Degenerative changes similar to those during spontaneous metamorphosis described above could be reproduced in vitro. In tissue fragments isolated from the small intestine of stage 57 tadpoles and cultured in the presence of thyroid hormone, the number of intraepithelial macrophage-like cells reaches its maximum around the 3rd day of cultivation when the larval epithelial cells most rapidly decrease in number. These results suggest that the rapid degeneration of larval epithelial cells occurs not only because of apoptosis of the epithelial cells themselves but also from heterolysis by macrophages. The macrophages probably originate in the connective tissue, actively proliferate, migrate into the larval epithelium around the beginning of metamorphic climax, and are finally extruded into the lumen.     

Molecular cloning of hepatic mRNAs in Rana catesbeiana responsive to thyroid hormone during induced and spontaneous metamorphosis

Amphibian metamorphosis affords a useful experimental system in which to study thyroid hormone regulation of gene expression during postembryonic vertebrate development. In order to isolate gene-specific cDNA probes which correspond to thyroid hormone-responsive mRNAs, we employed differential colony hybridization of a cDNA library constructed from poly(A)+ RNA of thyroxine-treated premetamorphic tadpole liver. From an initial screening of about 6000 transformants, 32 "potentially positive" colonies were obtained. The recombinant cDNA-plasmids from 13 of these colonies plus two "potentially negative" colonies were purified for further study. Southern blot analysis of the plasmid DNA was employed to determine whether different cDNAs encoded for the same mRNA. The effect of thyroid hormone on the relative levels of specific mRNA species was examined by Northern analysis of liver RNA from premetamorphic tadpoles, thyroxine-treated tadpoles, and adult bullfrogs. Three independent cDNA clones were obtained which encoded thyroid hormone-enhanced mRNAs. We also obtained two independent cDNA clones encoding thyroid hormone-inhibited mRNAs and three independent clones encoding thyroid hormone-unresponsive mRNAs. The levels of two thyroid hormone-enhanced mRNAs and one thyroid hormone-inhibited mRNA were essentially the same in the thyroid hormone-treated tadpole liver and adult liver, suggesting that thyroid hormone induces stable changes in liver gene expression during spontaneous metamorphosis. Using selected cDNAs, RNA dot blot analysis of liver mRNA from tadpoles at different stages of metamorphosis showed that the level of one thyroid hormone-enhanced mRNA increased during late prometamorphosis and metamorphic climax. Similarly, a mRNA which was strongly inhibited by thyroid hormone treatment was observed to decline during prometamorphosis and reach undetectable levels during metamorphic climax. One mRNA was detected which was reproducibly inhibited by thyroid hormone treatment but which remained essentially unchanged during spontaneous metamorphosis. These results provide the first direct evidence for the coordinate and selective pretranslational regulation by thyroid hormone of several liver genes during the developmental process of metamorphosis.     

Influence of the photocycle and thermocycle on rhythms of plasma thyroxine and plasma and ocular melatonin in late metamorphic stages of the bullfrog tadpole, Rana catesbeiana

The diel fluctuations in plasma thyroxine (T(4)) and plasma and ocular melatonin entrain to the light/dark (LD) cycle in the bullfrog tadpole, although the phase of the rhythms changes during development. Previous studies on the rhythmicity of these hormones were conducted under various LD cycles, but with a constant temperature, raising the question of the role of the natural thermocycle in determining the phase of the rhythms, and the changes that occur in the hormone levels and rhythms during late metamorphosis. To study this question, tadpoles were acclimated to simulated natural conditions of 14.5L:9.5D with a corresponding thermocycle in which the thermophase was 28 degrees C and the cryophase was 18 degrees C, or to the same thermocycle under constant light (24L). On both photoregimens, the diel fluctuations changed between prometamorphosis and metamorphic climax. However, more statistically significant rhythms, as indicated by the cosinor, occurred on 14.5L:9.5D than on 24L. At climax on the LD cycle, all hormones peaked around the same time in the late scotocryophase, whereas on 24L, plasma T(4) peaked in the thermophase and plasma and ocular melatonin peaks occurred some distance from each other early in the cryophase. The earlier peaks of plasma and ocular melatonin on 24L were due to a transient rise in these hormones at the onset of the cryophase, which was not sustained in the absence of an LD cycle. On 14.5L:9.5D with a corresponding thermocycle, the hormone rhythms had nearly the same phases as was found in previous work on 12L:12D at a constant temperature of 22 degrees C, allowing for minor phase shifting due to the photocycle differences, indicating that in this species laboratory studies on constant temperature give valid results even in the absence of a thermocycle. The findings show that the phases of the hormone rhythms are determined by the LD cycle although the onset of the cryophase, in the absence of a photocycle, may exert some influence on the nighttime rise in melatonin. The developmental rise in plasma T(4), and drop in plasma melatonin, occurred on both 14.5L:9.5D and 24L, indicating, taken together with previous work, that these climactic changes were independent of temperature and light cycling.     

Energetics of metamorphic climax in the pickerel frog (Lithobates palustris)

Anuran metamorphosis, the transition from aquatic larvae to terrestrial juveniles, is accompanied by significant morphological, physiological, and behavioral changes. Timing of metamorphosis and final size, which can influence adult fitness, may depend on sufficient energy accumulated during the larval period to support metamorphosis. However, only two species of anurans have been examined for energetic costs of metamorphosis, Rana tigrina and Anaxyrus terrestris. Based on these species, it has been hypothesized that differences in energy expenditure are related to duration of metamorphosis. To compare energetic costs of metamorphosis among species and examine this hypothesis, we quantified the total energy required for metamorphosis of Lithobates palustris tadpoles by measuring oxygen consumption rates over the duration of metamorphic climax using closed-circuit respirometry. Total energy costs for L. palustris were positively related to tadpole mass and duration of metamorphic climax. However, larger tadpoles completed metamorphosis more efficiently because they used proportionally less total energy for metamorphic climax than smaller counterparts. Costs were intermediate to R. tigrina, a larger species with similar metamorphic duration, and A. terrestris, a smaller species with shorter metamorphic climax. The results supported the hypothesis that amphibian species with more slowly developing tadpoles, such as ranids, require more absolute energy for metamorphosis in comparison to more rapidly developing species like bufonids.     

Thyroxine-binding proteins in liver and tail of Rana catebeiana undergoing metamorphosis.

Presence of a thyroxine-binding protein was demonstrated in vivo in cell sap of tail and liver of metamorphosing Rana catesbeiana tadpoles. Thyroxine-binding protein was not present in tail of prematamorphic tadpoles while it appeared during progressing metamorphosis roughly coinciding with the beginning of tail resorption. Susceptibility to pronase indicates that this thyroxine-binding macromolecule is protein in nature. Thyroxine-binding in liver was already present during premetamorphic stages and increased further during metamorphosis. A further difference between tail and liver thyroxine-binding protein was evidenced by molecular sieve chromatography on Sephadex G-200 indicating a molecular weight of thyroxine-binding protein in the tail of 60 000 as opposed to 42 000 for liver. Scatchard analysis of tail cell sap of tadpoles in metamorphic climax revealed a high affinity thyroxing binding site (Kd of 2 - 10(-10) M) of low capacity (1.7 pmol per mg protein) while tadpoles in premetamorphic stage had a thyroxine-binding site of lower affinity (9 - 10(-10) M) and higher capacity (4.8 pmol per mg protein). Thus affinity of thyroxine binding is 4-fold in metamorphic climax and appears to reflect the appearance of thyroxine binding observed in vivo.     

Developmental and diel profiles of plasma corticosteroids in the bullfrog,Rana catesbeiana

Corticosteroids synergize with the thyroid hormone (TH) at late metamorphic stages and might have a role in the hormonal regulation of amphibian metamorphosis. This role could be influenced by diel fluctuations, particularly if the peak of the plasma corticoids changed in relation to the TH peaks. Diel variation in plasma corticosteroids was studied in Rana catesbeiana prometamorphic and climax tadpoles on 18:6, 12:12 and 6:18 light:dark (LD) cycles. Cortisol (hydrocortisone; HC) and aldosterone (ALDO) exhibited different, but LD cycle-specific, circadian fluctuations at prometamorphosis, whereas corticosterone (CORT) was undetectable (less than 1.18 ng/ml). HC, ALDO and CORT rhythms became synchronous at early metamorphic climax on all LD cycles, although the cosinor-derived acrophases, which occurred around the time of the dark:light transition, shifted approximately 6 h earlier from 18L:6D to 6L:18D. On both 18L:6D and 12L:12D, the acrophase of HC changed little from prometamorphosis to climax, whereas that of ALDO underwent a major phase shift. On 6L:18D, both the ALDO and the HC acrophases shifted at climax. These LD cycle-specific phase shifts of the diel rhythms placed the acrophases of the corticoids in different phase relationships to that of the previously determined thyroxine (T(4)) acrophase at climax, and may partially explain the influence of the light regimen on metamorphic timing. The pronounced diel variations in the corticoid concentrations from the troughs to the peaks show that hormone levels are a function of the time of day and the environmental lighting regimen, which need to be taken into account in measuring the level of plasma hormones in amphibians. The 24-h means calculated from the data of all the sampling times showed that only plasma ALDO and CORT, but not HC, rose markedly at climax, although there were significant LD cycle-related differences in the mean levels of both HC and ALDO at prometamorphosis, and in HC at climax. Additional work sampling at mid-light showed that plasma CORT peaked at Stage XXIII, decreased at the end of climax, and remained low in the postmetamorphic froglet at 2.1 ng/ml. In the adult bullfrog, CORT was clearly the predominant corticosteroid at 34.3 ng/ml, whereas HC and ALDO levels were only approximately 1.3 ng/ml.     

The fat body of bullfrog (Lithobates catesbeianus) tadpoles during metamorphosis: changes in mass, histology, and melatonin content and effect of food deprivation

The fat body of Lithobates catesbeianus (formerly Rana catesbeiana) tadpoles was studied during metamorphosis and after food deprivation in order to detect changes in its weight, adipocyte size, histology, and melatonin content. Bullfrog tadpoles have large fat bodies throughout their long larval life. Fat bodies increase in absolute weight, and weight relative to body mass, during late stages of prometamorphosis, peaking just before climax, and then decreasing, especially during the latter stages of transformation into the froglet. The climax decrease is accompanied by a reduction in size of adipocytes and a change in histology of the fat body such that interstitial tissue becomes more prominent. Food deprivation for a month during early prometamorphosis significantly decreased fat body weight and adipocyte size but did not affect the rate of development. However, food restriction just before climax retarded development, suggesting that the increased nutrient storage in the fat body before climax is necessary for metamorphic progress. Melatonin, which might be involved in the regulation of seasonal changes in fat stores, stayed approximately at the same level during most of larval life, but increased sharply in the fat body during the late stages of climax. The findings show that the rate of development of these tadpoles is not affected by starvation during larval life as long as they can utilize fat body stores for nourishment. They also suggest that the build up of fat body stores just before climax is necessary for progress during the climax period when feeding stops.     

Metamorphic rate as a function of the light/dark cycle in Rana pipiens larvae

1. The rate of development of Rana pipiens tadpoles in spontaneous and thyroxine (T4)-induced metamorphosis was studied on light/dark (LD) cycles in which the photophase was held constant while the scotophase was progressively extended or vice versa. 2. Metamorphic rate fluctuated in both types of experiments as the LD cycle lengthened. However, the pattern of resonance differed with the length of the photophase. For example, with an 8 hr light phase, development rate slowed and then increased as the cycle was extended from 24 to to 36 hr, whereas with a 12 hr photophase the reverse took place. 3. The findings are compatible with the occurrence of a rhythm of light sensitivity in photoperiodic time measurement in this amphibian. 4. From the viewpoint of hormonal mechanisms, it is suggested that photoperiodic effects on metamorphic rate result from different patterns of melatonin secretion under the various LD cycles, since melatonin can modify the action of T4 in metamorphosis.     

Ontogeny of the Thyroid Glands During Larval Development of South American Horned Frogs (Anura,Ceratophryidae)

The role of thyroid hormone (TH) in anuran metamorphosis has been documented from a variety of approaches, but the sequence of morpho-histological development of the thyroid glands that produce the secretion of the hormone was assumed invariant from studies of relatively few species even when the effects of environmental influences on larval development and metamorphosis have been largely documented. There are anurans in which developmental and growth rates diverge, and the resulting heterochrony in growth and development produces giant/miniature tadpoles, and or rapid/delayed metamorphosis suggesting changes of the activity of the thyroid glands during larval development. Herein, we analyze the morpho-histological variation of the thyroid glands in larval series of Ceratophrys cranwelli, Chacophrys pierottii, Lepidobatrachus laevis and L. llanensis that share breeding sites along semiarid environments of the Chaco in South America, belong to a monophyletic lineage, and present accelerated patterns in growth and development in order to have a morphological evidence about a possible shift of TH physiology. We describe gross morphology and histology of the thyroid glands and find features shared by all studied species such as the presence of supernumerary heterotopic follicles; changes in the volume and number of follicles towards the metamorphic climax, and cuboidal epithelia with occasional intra-cellular vacuoles as signs of low glandular activity without a manifest peak at the climax as it was assumed for anurans. We discuss different lines of evidence to interpret sources of extra supplement of TH to support the rapid metamorphosis. These interpretations highlight the necessity to design a research program to investigate the endocrine variation during development of ceratophryids taking in account their morphology, physiology and ecology in order to learn more about the effects of environmental and developmental interactions involved in the anuran evolution.     

Thyroid hormone-dependent repression of α1-microglobulin/bikunin precursor (AMBP) gene expression during amphibian metamorphosis

 A Xenopus AMBP (xAMBP) cDNA clone was isolated from a subtracted liver cDNA library by differential hybridization screening. The deduced amino acid sequence shared 50–60% identity with its mammalian counterparts, which are the precursors of the plasma glycoproteins, α1-microglobulin and bikunin. Both peptide structures were well conserved in xAMBP. Northern and in situ hybridization revealed that the xAMBP gene was specifically expressed in liver parenchymal cells. The gene was activated around embryo hatching and repressed at the metamorphic climax stage. During adult life the mRNA level remained low. Treating the tadpoles with thyroid homone prematurely reduced the mRNA level. Furthermore, thyroid hormone acted on larval hepatocytes in primary culture and reduced the mRNA level. Thus, xAMBP gene expression appears to be repressed through the direct action of thyroid hormone on the hepatocytes at the metamorphic climax stage. On the other hand, adult hepatocytes in thyroid hormone-free culture medium expressed mRNA at a low level, which was not reduced in response to thyroid hormone, suggesting that the repressed xAMBP gene expression in adult hepatocytes was maintained in a thyroid hormone-independent manner. The unique expression profile suggested that the xAMBP gene plays a biological role in the progression of amphibian metamorphosis. Received: 12 April 1996 / Accepted: 19 September 1996     

Histological changes in the pituitary-thyroid axis during spontaneous and artificially-induced metamorphosis of larvae of the flounder Paralichthys olivaceus

Summary Histological changes in the pituitary TSH cells and in the thyroid gland of flounder (Paralichthys olivaceus) larvae during spontaneous or artificially induced metamorphosis were studied. Activity of the immunoreactive TSH cells (IrTSH cells) gradually increased during premetamorphosis, reaching the highest level in prometamorphic larvae, and the cells were degranulated in metamorphic climax. The IrTSH cells were most inactive at the post-climax stage. The thyroid gland was morphologically the most active in metamorphic climax when the degranulation occurred in the pituitary IrTSH cells, and appeared inactive at post-climax. A few weeks after metamorphosis, both the IrTSH cells and the thyroid gland appeared to be activated again in the benthic, juvenile flounder. Administration of thyroxine or thiourea revealed negative feedback regulation of the pituitary-thyroid axis in flounder larvae. These results indicate that activation of the pituitary-thyroid axis induces metamorphosis in the flounder.     

Retardation of thyroxine-induced metamorphosis by Amphenone B in toad tadpoles

The effect of Amphenone B, an inhibitor of corticoid synthesis, on thyroxine (T4)-induced metamorphosis was studied in toad tadpoles kept in thiourea. Amphenone injections retarded T4-induced tail resorption markedly. The effect of Amphenone was nullified by aldosterone and corticosterone added to the water in which tadpoles were kept. Steroidogenic cells of adrenals in Amphenone-injected animals were enlarged markedly as compared with those in the saline-injected tadpoles or the Amphenone-injected tadpoles which were supplemented with corticoids. The results strongly suggest that endogenous corticoids act together with thyroid hormone to accelerate metamorphosis.     

Developmental implications of differential effects of calcium in tail and body skin of Anuran tadpoles

The effects of external Ca(++) on metamorphosis of Rana catesbeiana tadpoles were assessed. Treatment of tadpoles with Ca(++) (0.05 mM) during early prometamorphic stages induced precocious metamorphic events such as tail regression, shortening of the intestine, forelimb emergence, and keratinization of body epidermis within 23 days of treatment compared to control tadpoles still in mid-prometamorphic stages. These effects of Ca(++) are probably mediated by the thyroid gland, as indicated by histological features of the gland at the light and electron microscopic levels. Calcium levels of tail and body skin were measured at various stages of development by atomic absorption spectrophotometry. In control and experimental groups, body skin had significantly higher Ca(++) concentrations than tail skin. There were no statistically significant effects of developmental stage on Ca(++) levels of tail or body skin. Experimental Ca(++) treatment significantly increased Ca(++) concentration in tail but not body skin. Ultrastructure studies and gel electrophoresis indicated that calcium induced keratinization of body skin, but not tail epidermis. Ca(++)-treated tail epidermis showed various autolysing figures in apoptotic cells. In summary, calcium treatment accelerated metamorphosis and induced the following region-dependent cellular events: keratinization of body skin-a characteristic of adult epidermis-and programmed cell death in the tail. Whatever signal elicited by calcium in this experimentally induced accelerated metamorphosis is probably mediated via the thyroid gland.     

Early larval development and metamorphosis in the summer flounder: changes in per cent whole-body water content and effects of altered thyroid status

At the end of premetamorphosis, summer flounder Paralichthys dentatus larvae had 84·1% whole-body water content (WBW), which decreased to the lowest levels (8·5%) at the start of metamorphic climax (MC). During mid- and late MC, %WBW was slightly higher (82·1%) then returned to the lowest levels at the juvenile stage. In fish treated with thyroxine (T₄-Na salt, 100 ng ml⁻¹) beginning at premetamophosis, %WBW never differed from controls of the same age throughout metamorphosis, despite an earlier start of metamorphic climax and transitional settling behaviour. This suggests that thyroid hormones do not mediate the drop in %WBW which accompanies natural metamorphosis. Thiourea (TU, 30 μg ml⁻¹) treatment of fish over the same period induced a developmental stasis in early MC which was accompanied by initially higher %WBW than controls at 33 days post-hatch, followed by a progressive decrease to abnormally low %WBW by 42 and 45 days post-hatch. Since concurrent treatment with TU+T₄ rescued the fish from both the TU-induced developmental stasis and abnormally low %WBW, these findings suggest that thyroid hormones, or thyroid hormone-mediated developmental progression, are necessary for regulating %WBW.