Mouthbrooding in the black-chinned tilapia, Sarotherodon melanotheron (Pisces: Cichlidae): the presence of eggs reduces androgen and estradiol levels during paternal and maternal parental behavior.

The black-chinned tilapia (Sarotherodon melanotheron) is considered a paternal mouthbrooder in which the male churns the eggs in his mouth for 14-18 days after they are fertilized. We previously showed that plasma androgen and estradiol concentrations increased significantly between the beginning and end of mouthbrooding. Here we provide evidence that androgen and estradiol are relatively high in prespawning male tilapia, decrease significantly with the initiation of mouthbrooding, and return to and slightly surpass prespawning levels by the end of mouthbrooding. We then test the hypothesis that the initiation of mouthbrooding causes a decrease in androgen and estradiol. We raised single pairs of tilapia in seawater aquaria and conducted two experiments. In the first, we removed eggs from males on the morning of Day 1 of mouthbrooding, followed changes in plasma androgen and estradiol in the afternoon of Day 1 and on Days 6, 11, and 16, and compared them to levels in control males who kept their eggs. Removing eggs from mouthbrooding males on the morning of Day 1 eliminated the decrease in plasma androgen and estradiol observed in the afternoon of Day 1 of mouthbrooding. This demonstrates that the eggs are necessary for the decrease. We observed that, infrequently, the female of the pair would pick up the eggs to brood them in her mouth. In a second experiment, in the afternoon of Day 1, we sampled both the male and female of pairs in which the female brooded eggs. In female-brooding pairs, the females had significantly lower androgen and estradiol levels than prespawning females. The black-chinned tilapia in captivity exhibits both paternal and maternal parental behavior. In both sexes the initiation of mouthbrooding triggers a decrease in androgen and estradiol concentrations. We conclude that the presence of eggs inhibits the pituitary-gonadal axis in both males and females and hypothesize that a chemical signal from the eggs is delaying the initiation of the next brood.

Paternal mouthbrooding in the black-chinned tilapia, Sarotherodon melanotheron (Pisces: cichlidae): changes in gonadal steroids and potential for vitellogenin transfer to larvae.

The black-chinned tilapia (Sarotherodon melanotheron) is a paternal mouthbrooder. Pairs of adult black-chinned tilapia were raised in freshwater and the males were sampled during the mouthbrooding cycle. Sampling also occurred 10 days after release of the free-swimming fry for comparison. During the first week of incubation of the eggs, total androgens and estradiol were low (<5 and <0.3 ng/ml, respectively). During the second week of brooding, when the eggs have hatched and they are called newly hatched embryos, plasma levels of gonadal steroids increased (13-38 ng androgen/ml and >0.6 ng estradiol/ml). The plasma concentrations of vitellogenin (VTG) in male parents changed during mouthbrooding, with decreases occurring between egg pickup and hatching of the embryo (Day 6 of mouthbrooding). The pattern of change in concentrations of VTG in surface mucus of male parents differed from the pattern in plasma, with peak concentrations occurring at the time of hatching. The amount of VTG in mucus was similar to that measured in the female Oreochromis mossambicus during mouthbrooding of embryos. The appearance of peak VTG levels in the mucus at the time of hatching when plasma levels have declined and the availability of comparable amounts of mucus VTG in both maternal and paternal mouthbrooding tilapia, despite unequivalent plasma levels, support the possibility that parental provisioning of the young occurs during mouthbrooding in tilapia.

Metamorphosis in the summer flounder, Paralichthys dentatus: thyroidal status influences gill mitochondria-rich cells.

Metamorphosis in the summer flounder (Paralichthys dentatus) is mediated by thyroid hormones (TH) and is accompanied by changes in gill mitochondria-rich cells (MRCs) and in salinity tolerance. Altered thyroid status during larval development and metamorphosis in this species influences salinity tolerance, though the influence of any hormone on MRCs of larval marine teleosts is not known. This study characterized the effect of altered thyroid status on MRC intracellular membranes, mitochondria size and ultrastructure, immunoreactive (ir)-Na(+),K(+)-ATPase, and cell size and density during metamorphosis in summer flounder. Inhibition of metamorphosis with thiourea (30 ppm) (TU, an inhibitor of TH synthesis) inhibited changes in MRCs, producing large "larval" type MRCs with weak reactivity to osmium; large, electron-lucent mitochondria; and weak ir-Na(+),K(+)-ATPase. Replacement of TH with TU + thyroxine-Na salt (100 ppb) rescued the fish from developmental inhibition, producing smaller "juvenile" type MRCs with strong reactivity to osmium; smaller, electron-opaque mitochondria; and strong ir-Na(+), K(+)-ATPase. The findings suggest that TH are necessary for MRCs to change from larval to juvenile form during metamorphosis.

Immunocytochemical and immunogold localization of two prolactin isoforms in the same pituitary cells and in the same granules in the tilapia (Oreochromis mossambicus).

The tilapia pituitary secretes two forms of prolactin (tPRL) and a single growth hormone (tGH). The tPRLs share only 69% sequence identity and are designated tPRL177 and tPRL188 to indicate the number of amino acid residues in each isoform. Our aim was to develop specific antisera for detection of these three related polypeptides. Ten peptides corresponding to unique epitopes on the tPRLs and two peptides of tGH were synthesized using solid-phase methods, conjugated to carrier proteins, and used as immunogens for antibody production in rabbits. Select antisera for the tPRLs were highly specific, exhibiting only 1% cross-reactivity to the alternate tPRL under noncompetitive ELISA conditions at dilutions used in immunocytochemical analysis. The anti-tGH specifically bound to cells in the proximal pars distalis. Production of both tPRLs by a single cell type was indicated by the binding of both anti-tPRL177 and anti-tPRL188 to the same cells in the rostral pars distalis. Ultrastructural analysis of PRL-producing cells stained sequentially using the two different anti-tPRL antibodies labeled with immunogold of two size classes indicated that both tPRLs appear in the same granules. These findings suggest that the biological significance of two forms of PRL in the adult tilapia is not a function of differential regulation of two different classes of PRL cells or differential release of unique secretory granules.

Induction by beta-estradiol of vitellogenin in striped bass (Morone saxatilis): characterization and quantification in plasma and mucus.

Striped bass (Morone saxatilis) were implanted with beta-estradiol to induce the production of vitellogenin, the egg yolk precursor produced by the liver. Electrophoretic analysis revealed that beta-estradiol caused marked production of a plasma protein of apparent molecular mass 170 kDa. Size exclusion chromatography suggested that the estradiol-induced protein circulated as a dimer. This protein was purified from the plasma of estradiol-treated fish by DEAE-agarose column chromatography and used to induce antibodies in rabbits and goats. Western blots revealed that the antiserum bound to the putative vitellogenin in plasma from estradiol-treated fish and adult females, but not with any proteins in male plasma. Western blot of ovarian extract revealed several smaller immunoreactive protein bands and supported the identity of the purified protein as vitellogenin. A competitive ELISA was developed with sensitivity in a range from 8 to 1000 ng/ml. Plasma concentrations of adult females during their spawning migration ranged from 100 to 600 micrograms/ml. Western blot of mucus extract revealed the presence of a 170-kDa protein in vitellogenic female fish along with several minor bands ranging from 50 to 110 kDa. Positive immunoreactivity was present in the surface mucus of all females and in none of the males collected during a spawning migration in the Hudson River.

Aromatase is concentrated in the proximal pars distalis of tilapia pituitary.

Aromatase has been identified in the telostean, avian, and mammalian pituitaries, although its cellular location(s) is not yet certain. To address this question, experiments were performed in tilapia (Oreochromis mossambicus), a species which has been well characterized with respect to the intraglandular distribution of the different pituitary cell types. To estimate aromatase, glands were microdissected into rostral pars distalis (RPD), proximal pars distalis (PPD), and neurointermediate lobe (NIL) and organs were cultured in the presence of [3H]androstenedione for 16-24 hr. [3H]Estrogen products were isolated and quantified after ether extraction, hydrolysis with glucuronidase-sulfatase, thin-layer chromatography, and phenolic partition. Authentic estrone or estradiol-17 beta were produced by all pituitary regions and also by the urophyseal region of the spinal cord. Aromatase was two to five times higher in PPD than in RPD or NIL and similar to activity in adjacent hypothalamus-preoptic area (HPOA). Much lower estrogen yields were obtained in cultures of cerebellum, urophysis, and other cord regions. Since the PPD contains most of the somatotropes, these data are consistent with earlier studies implicating GH3/GH4 cell strains as an enriched enzyme source, although its presence in other cell types cannot be ruled out. The unusually high and localized aromatase in tilapia pituitary renders this species a useful model for studying the targets and functional importance of estrogen as a parahormone in the pituitary.

Complete amino acid sequences of a pair of fish (tilapia) prolactins, tPRL177 and tPRL188.

The complete amino acid sequences of a pair of tilapia (Oreochromis mossambicus) prolactins (PRLs) were determined. The larger PRL of molecular mass 20,836 Da consists of 188 amino acid residues. The smaller PRL of molecular mass 19,584 Da is 11 residues shorter. On alignment of the two sequences, the 19.6-kDa PRL (tPRL177) has two conspicuous deletions on the NH2-terminal side of the disulfide bond which connects the first and second cysteine residues. The degree of similarity between the two PRL sequences is unexpectedly low (130 identical residues, 69%) compared with that between the variants of other teleostean PRLs. Circular dichroism spectra and hydropathy profiles suggest structural similarity of the two PRLs. The sequence of the 20.8-kDa PRL (tPRL188) has 69% identity with that of salmon PRL. The sequence of tPRL177 is 56% identical with that of salmon PRL. Each tilapia PRL is equally similar to mammalian PRLs (about 30% identical residues). Regions highly conserved among teleostean and mammalian PRLs were identified on the COOH-terminal side of the disulfide bond connecting the first and second cysteine residues.

Thyroid function in adult rough-skinned newts (Taricha granulosa): effects of hypophysectomy and hormone replacement.

This study examines the regulation of thyroid activity in adult rough-skinned newts, Taricha granulosa. Aquatic, breeding-condition newts were hypophysectomized and were injected for 23 days with the mammalian adenohypophysial hormones: ACTH, PRL, and GH. During a fourth week of treatment, these newts also received TSH. Plasma T4 concentrations were lower in hypophysectomized newts than in sham-hypophysectomized newts 23 days after surgery. Both ACTH and PRL increased T4 titers above these in saline-injected control newts; GH alone had no effect. TSH effectively increased the plasma T4 concentrations in newts from all five groups. GH, and possibly PRL, depressed the response to TSH. Plasma T4 concentration was positively correlated with body fluid loss, suggesting that hormone concentration/dilution effects may occur generally in animals such as these that undergo substantial seasonal changes in degree of hydration.