End Of Prometamorphosis Research Articles

Characterization and Developmental Expression Profile of the Steroidogenic Acute Regulatory Protein (StAR) in the Gonad-Mesonephros Complex of Lithobates sylvaticus

The steroidogenic acute regulatory (StAR) protein is responsible for the movement of cholesterol across mitochondrial membranes and is therefore a key factor in regulating the timing and rate of steroidogenesis. In this study, we characterized the coding region of the star gene in the ranid Lithobates sylvaticus and studied star mRNA levels in steroidogenic tissues during development and under natural conditions. Our results support previous research showing that the StAR sequence is well conserved. We determined that star is expressed in both the interrenal and gonadal tissues of adults and in the tadpole gonad-mesonephros complex (GMC). The mRNA levels of star in the GMC were found to increase during tadpole development, reaching a maximum between Gosner stages (Gs) 32 and 38. We observed a significant drop in star mRNA levels at the end of prometamorphosis (Gs40-41), just before the start of the metamorphic climax. Significant differences in star levels between females and males, with males presenting higher levels than females, were detected at Gs36-38. To our knowledge, this is the first study that reports transitory star sex differences in tadpoles' developing GMC. Our results suggest an involvement of StAR in anuran late male GMC formation and development that requires further investigation.

Giant Tadpole and Delayed Metamorphosis of Pseudis platensis Gallardo, 1961 (Anura, Hylidae)

Pseudis platensis, like Pseudis paradoxa, is a Neotropical aquatic frog with a giant tadpole. Despite being one of the few anuran species whose larvae grow larger than adults, knowledge of its larval development is limited to prometamorphic stages. This study describes qualitative and quantitative variation through P. platensis larval development, including descriptions of various traits (lateral line, oral disc, vent tube, limbs) and their transformations during metamorphosis. We found similarities with the development of other anurans at larval stages up to the end of prometamorphosis but not with their metamorphic stages. For this species, we described key features to characterize 13 ontogenetic stages as follows: four for prometamorphic larval morphologies, eight stages covering metamorphic changes, and one defining the posmetamorphic adult stage. At the end of prometamorphosis, tadpole body size in P. platensis is similar to adult size, but the tail shows extra development. Unlike most anurans, the disappearance of two larval traits (i.e., vent tube and tail) is delayed until later in metamorphosis. Larval development of P. platensis exhibits unique features not reported for other anurans and offers a model to study developmental and evolutionary aspects of anuran metamorphosis.

Cloning of Bullfrog Thyroid-Stimulating Hormone (TSH) β Subunit cDNA: Expression of TSHβ mRNA during Metamorphosis

A thyroid-stimulating hormone β subunit (TSHβ) cDNA encoding both signal peptide and mature TSHβ molecule was cloned from a cDNA library constructed from total RNA of the bullfrog (Rana catesbeiana) adenohypophysis. The bullfrog TSHβ mRNA was estimated by Northern blot analysis to be approximately 1 kb. The deduced amino acid sequence showed 40–61% homologies with the sequences of TSH β subunits of other vertebrates. Using the cDNA as a probe, we measured changes in mRNA expression in metamorphosing tadpoles of R. catesbeiana. The TSH β subunit mRNA level increased progressively throughout prometamorphic stages, reaching its maximum at the end of prometamorphosis. The maximum level was maintained throughout early and mid climax, declining at late climax. These results, together with previously obtained data on plasma prolactin and pituitary prolactin mRNA levels, as well as thyroid hormone levels, are discussed in relation to metamorphic changes occurring in the bullfrog larvae.

Changes of Pituitary Proopiomelanocortin mRNA Levels during Metamorphosis of the Bullfrog Larvae

Adrenal corticoids accelerate metamorphosis of amphibians by potentiating the action of thyroid hormone. Adrenal corticoid secretion is considered to be controlled mainly by adrenocorticotropic hormone generated from proopiomelanocortin (POMC) in the anterior lobe of the pituitary. In order to assess the changes in POMC mRNA levels during metamorphosis, a cDNA for POMC was isolated from a cDNA library constructed from bullfrog (Rana catesbeiana) pituitary polyadenylated RNA. Northern blot analysis using the POMC cDNA as a probe revealed that POMC mRNA levels in the anterior lobe were relatively low during premetamorphosis, rose during prometamorphosis, reached the maximum at the end of prometamorphosis and remained very high during climax. The POMC mRNA levels of the intermediate lobe, where α-melanophore-stimulating hormone is generated from POMC, were also determined in metamorphosing tadpoles. The POMC mRNA levels of the intermediate lobe increased as metamorphosis progressed and were maximal at mid-climax. High POMC mRNA levels were observed even in larvae that had adapted to a white background. The significance of these findings and their relationships to the hormonal requirements during metamorphosis are discussed.

An Immunohistochemical and Morphometric Analysis of Insulin, Insulin-like Growth Factor I, Glucagon, Somatostatin, and PP in the Development of the Gastro-entero-pancreatic System ofXenopus laevis

The ontogeny of the classical islet hormones insulin (INS), glucagon (GLUC), somatostatin (SOM), and pancreatic polypeptide (PP) as well as insulin-like growth factor I (IGF-I) in the gastro-entero-pancreatic (GEP) system ofXenopus laevis(stages 41–66) was studied using double immunofluorescence and morphometric analysis. As early as stage 41, clustered INS-immunoreactive (-IR) and isolated GLUC-IR cells occurred in the pancreas. The first SOM-IR cells appeared at stage 43, followed by PP-IR cells at stage 46. About 79% of the PP immunoreactivity was confined to a subpopulation of the GLUC-IR cells. Both the GLUC/PP-IR cells and the PP-IR cells were located in a distinct area of the pancreas. The first islets occurred in premetamorphosis (around stage 50) and comprised mainly INS-IR and GLUC-IR cells. The majority of SOM-IR, PP-IR, and GLUC/PP-IR cells was dispersed. The numbers of hormone cells remained quite constant until the end of prometamorphosis (stage 58). Around stages 60–62, the islets were partly disintegrated and the numbers of islet cells slightly decreased. At stage 63, the cell number began to increase and reached the levels typical for the adult around stage 66. After metamorphic climax, the islets were reformed. In the gastrointestinal tract, transient INS-IR cells occurred prior to the adaptation of the gastrointestinal tract to feeding (stages 41–44) and during metamorphosis when there is remodeling of the gastrointestinal tract (stages 60–63). Therefore, INS released from the transient mucosal INS-IR cells may be involved in the temporary proliferation of mucosal epithelial cells. The first GLUC-IR and SOM-IR cells were seen at stage 41. PP-IR cells followed at stage 46. In contrast to the islets, GLUC-IR and PP-IR cells constituted different cell populations. Around stage 46, the first IGF-I immunoreactions appeared in the GEP-system. In pancreas, IGF-I immunoreactivity was found in the GLUC/PP-IR cells (85–99%) but was absent from INS-IR, GLUC-IR, and SOM-IR cells. The IGF-I-IR gastro-entero-endocrine cells, however, seemed to contain none of the classical islet hormones.

Pigment cell localizations in anuran ventral skin at climactic metamorphosis

In anuran amphibians, the specific color pattern of the skin is expressed after metamorphosis, and its formation involves pigment cell migrations. Pigment cells are differently distributed in the tadpole, larval, and froglet skin. To learn more about their fate during metamorphic climax and in the young froglet, we focused our attention on the different localizations of larval melanophores and iridophores in the ventral skin of Rana esculenta before and during skin homing. Localizations of melanophores and iridophores can be elucidated at the developmental stages suggested by Taylor and Kollros (TK stages). At TK stage II (during early premetamorphosis), large melanophores beneath the larval skin are detected. At TK stage X, dispersed melanophores lie under bundles of muscular striated fibrils near the larval skin; they are also observed at the vascular level. At TK stage XVII (prometamorphosis), melanophores are extended on the inner side of the basement lamellar collagen. At the end of prometamorphosis, iridophores are located with melanophores in the separating space between attached basement collagen and derived basement collagen. At TK stage XX (earlier climax), melanophores and iridophores are detected inside the upper extremities of fractures opened in the derived basement collagen. At TK stage XXIV (later climax), both types of larval pigment cells are observed in the inner extremities of breaks derived from the fractures. During climax, these pigment cells occupy the well-formed breaks. At TK stage XXV in young froglet, the pigment cells remain alone in the breaks formed in the derived basement collagen. Briefly, breaks in the basement lamellar collagen are opened by invading cell processes of mesenchymal cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Thyroxine surge in metamorphosing flounder larvae

The tissue concentration of thyroxine (T 4) in larval flounder ( Paralichthys olivaceus) was studied at various stages of their metamorphosis using a specific radioimmunoassay developed for samples obtained by quantitative extraction of the hormone from fish eggs and larvae. T 4 concentrations were below the limit of detection in extracts of fertilized eggs and larvae until the end of prometamorphosis. Several dorsal fin rays continued growing from premetamorphosis until the end of prometamorphosis. The hormone became detectable at the onset of metamorphic climax when the elongated dorsal fin rays stopped growing. A sharp increase in tissue T 4 concentration was observed in midclimax of metamorphosis, and the high level of T 4 (11–13 ng/g body weight) was maintained until the end of the climax. The period of elevated T 4 levels coincided with resorption of the elongated dorsal fin rays. The right eye completed translocation from the right to the left side of the body during metamorphic climax. The tissue T 4 level declined to about a half of the peak value during postclimax, and then increased moderately in later adult-type juveniles. The results of an additional experiment, in which fertilized eggs and newly hatched larvae were treated with T 4 and/or thiourea, suggested the presence of negative feedback regulation of pituitary-thyroid axis, implying an involvement of thyroid hormones even in early premetamorphic larvae. These results suggest that metamorphic climax is induced by a surge of thyroid hormone, and that thyroid hormone may also regulate development before and after the metamorphosis.

Metabolism of thyroxine in Rana catesbeiana tadpoles during metamorphic climax.

Previous studies have indicated that premetamorphic tadpoles do not convert T4 to T3 to a measurable extent (1). The present study was performed to determine whether a T4 5'-monodeiodinating system is acquired at later stages of development. [125I]T4 metabolism in vivo was determined in tadpoles at most stages of prometamorphosis and metamorphic climax and, for comparison, in premetamorphic tadpoles. The conversion of [125I]T4 to [125I]T3, as indicated by the presence of an 125I-labeled product in serum and liver preparations that cochromatographed with carrier T3, was sometimes observed in tadpoles near the end of prometamorphosis and was always evident in tadpoles that were either undergoing or had completed metamorphic climax. However, during this phase, no correlation could be drawn between the extent of T3 production and morphological development. The formation of T3 from T4 in vivo was significantly decreased in tadpoles pretreated with propylthiouracil. The T45'-monodeiodinating system could be induced in premetamorphic tadpoles by injecting them with either T4 or T3. This finding together with the observation that normal acquisition of this system occurs at the time when endogenous T4 and T3 levels are rising rapidly suggest that its development is under the control of the thyroid hormones.

Changes in thyroidal and plasma iodine compounds during and after metamorphosis of the bullfrog, Rana catesbeiana.

Thyroidal thyroxine (T4), 3,5,3′-triiodothyronine (T3), diiodotyrosine (DIT), monoidotyrosine (MIT), and iodide (I−) in metamorphosing tadpoles, froglets, and adult frogs were measured chemically by cation-exchange resin chromatography. Plasma T4, T3, DIT, and 3,3′,5′-triiodothyronine (rT3) were also measured by radioimmunoassay. Plasma protein-bound iodine (PBI) was determined by a modified ultramicro PBI method. T4, T3, MIT, and DIT levels in thyroids tended to increase both during and after metamorphosis. Plasma T4 increased at the end of prometamorphosis, and a sharp increase was found at the onset of the metamorphic climax. At stage XXI, the T4 level reached a peak of about 500 ng/100 ml. Thoreafter the amount of circulating T4 decreased slowly, but even at 7 and 14 days after metamorphosis it remained at a higher level than in the premetamorphic stage. The level of plasma T4 in adult frogs was only 40 ng/100 ml and was undetectable in 4-month-old froglets. Similar changes were found in circulating plasma T3 and its level was less than one-third of T4, especially at the metamorphic climax. Throughout all metamorphic stages, plasma T4 and T3 levels fluctuated remarkably only at the metamorphic climax. Plasma DIT was at the low level of about 200 ng/100 ml and did not change during metamorphosis. Moreover, a small amount of rT3, less than T4 values, was detectable only in tadpoles during the metamorphic climax and in adult frogs. PBI values were at the level of about 2.0 μg/100 ml from stage XV to stage XXII, and thereafter decreased gradually. The PBI level was much higher than the sum of plasma T4, T3, and DIT, and did not correlate with the circulating hormone levels. From these results a relationship between thyroidal and plasma hormone levels and an extrathyroidal monodeiodination of T4 were discussed.

Induction of Vitellogenin Synthesis in Xenopus laevis Tadpoles

Oestradiol induces vitellogenin synthesis in vitro in liver taken from Xenopus laevis tadpoles that are in late metamorphosis. Inducibility first appears at the end of prometamorphosis, and the response to oestradiol increases during the completion of metamorphosis. Oestradiol continuously present during development does not influence the stage at which tadpole liver becomes inducible. It seems that the acquisition of inducibility is part of the normal development of the liver, and independent of both the supply of oestrogen and the sex of the tadpole.

Measurement of several intestinal lysosomal hydrolase activities, through larval development of discoglossus pictus otth, anuran amphibian

The whole specific activities of 5 lysosomal enzymes (acid phosphatase, N-acetylβ-glucosaminidase,β-glucuronidase, cathepsin and aryl sulphatase) were measured in the guts of Discoglossus tadpoles, before and during metamorphosis.For each hydrolase, the activities show the same variation pattern. They significantly increase at the end of prometamorphosis and the beginning of the climax, while there is histolysis of the first (primary) epithelium of the gut.The measured activities decrease at the end of the climax, while a secondary, enzyme-negative epithelium is developing.The free specific activity of the acid phosphatase was measured in these same guts. It significantly increases at the end of prometamorphosis and the beginning of the climax, during the epithelial histolysis. The total activity increases more slowly than the free one.In the same way, an increase is obtained for the specific activity of the bound (intralysosomal) acid phosphatase.The changes in whole enzymatic activities may result from accelerated synthesis of lysosomal hydrolases due to the primary epitheliocytes, and induced by the simultaneous increase in the plasmatic level of the thyroid hormones. Furthermore the free activity of the acid phosphatase may produce an increased permeability or a breaking of the lysosomal membrane, by the same hormones.