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The aim of this study was to describe the time-course of estrogen-induced gene expression, corresponding plasma protein detection and histological alterations after cessation of octylphenol (OP) exposure of Cichlasoma dimerus, to test differential responses of biomarkers suitable for environmental monitoring. Male fish were exposed to a nominal concentration of 150 µg/L OP for 28 days, and later transferred to OP-free water aquaria for 1, 3, 7, 14, 21 or 28 days. Blood and mucus samples were obtained in order to analyze vitellogenin (VTG) and zona pellucida (ZP) proteins by Western blot; liver samples were used for gene expression and to assess tissue damage and further recovery of all the analyzed endpoints. Partial sequences of C. dimerus VTG and Na⁺/K⁺-ATPase were obtained. Comparison with VTGs of several teleosts supports that the partial sequence obtained for C. dimerus belongs to VTGAb type. ZP and VTG expression was highly up-regulated by OP. Immunoreactive (ir-) bands of 62, 52 and 50 kDa for ZP and 140, 103, 75 and 64 kDa for VTG, were detected after 28 days of OP exposure in plasma and mucus samples. After transfer of treated fish to clean water, ZP ir-bands in plasma disappeared rapidly (day 3), while VTG ir-bands decreased gradually; no ir-bands were detected on day 28 of recovery. Similarly, ZPB transcripts abruptly returned to background levels (day 3), earlier than for ZPC (day 7) or VTG (day 14). Liver from OP treated fish showed tissue disarrangement, eccentric and euchromatic hepatocytes nuclei and intense perinuclear basophilia. After the recovery period, these changes were still evident though less pronounced, accounting for irreversibility of tissue damage or the requirement for a longer period of depuration. The present results confirm that for biochemical and molecular biomarkers, such as induction of female proteins in male fish exposed to OP, complete recovery is achieved after adequate time of depuration (28 days). Male ZPB expression reflects a recent exposure to estrogenic contaminants, while VTG may reveal past exposures. The combination of biomarkers with different temporal responses such as C. dimerus ZP and VTG provides a more comprehensive interpretation of pollution status.

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The organochlorine insecticide endosulfan (ES) is widely used despite its high toxicity to fish (96-h LC(50) median value of 2.6 µg L(-1)). This study aimed to assess the acute toxicity, histological and physiological parameters after exposure to 0; 0.25; 1; 2; 3; 4 and 16 µg L(-1) ES for 96 h under semi-static conditions in a freshwater perciform fish, Cichlasoma dimerus. Prior to death, fish exhibited behavior indicative of neurotoxicity. No difference was found in brain AChE activity. A decrease in erythrocyte mean corpuscular volume, mean corpuscular hemoglobin and mean corpuscular hemoglobin concentration in exposed fish suggests a state of anemia. Histological alterations observed in exposed fish included hyperplasia of the interlamellar epithelium, blood congestion in secondary lamellae, and mucous cells hyperplasia and hypertrophy in gills; pycnotic nuclei and hydropic degeneration in liver; testicular damage. These moderate pathological responses in major organs could become crucial during reproduction and under prolonged exposure periods.

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We detected a close morphological association between melanin-concentrating hormone (MCH)-immunoreactive (ir) fibers and somatolactin (SL)-ir cells in the pars intermedia of the cichlid fish Cichlasoma dimerus by double-label immunofluorescence. Male pituitaries obtained from adult C. dimerus were incubated with 0.1-10 microM salmon MCH, and the amount of SL released into the culture medium was semi-quantified by Western blot. This assay showed an increase of SL release in a dose-dependent manner (linear regression: P<0.05). A close association of GnRH-ir fibers with SL-ir cells was also detected at the pars intermedia level. Male pituitaries were also incubated with 0.1-10 microM of mammalian GnRH, and SL release was semi-quantified by Western blot, showing an increase of released SL levels in a dose-dependent manner (linear regression: P<0.05). In contrast, SL release was unaffected from female pituitaries incubated with salmon MCH; however, an increasing tendency was observed when mammalian GnRH was used. Hypothalamic close association of MCH-ir perikarya and GnRH-ir fibers was found by double-label immunofluorescence indicating a possible relationship between them. These results suggest that SL, like other pituitary hormones, is under hypothalamic control and is involved in diverse physiological processes including background adaptation and reproduction. This study has also shown that the in vitro culture of a single C. dimerus pituitary is a feasible method for studying the control of SL release and other pituitary hormones.

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Somatolactin (SL) is a pituitary hormone present exclusively in fish that is involved in different physiological processes. The role of SL was evaluated in Cichlasoma dimerus (Teleostei, Perciformes) exposed for 10 days to a black and white background (BB and WB). Changes in alpha-melanophore stimulating hormone (alphaMSH) and melanin concentrating hormone (MCH) cells were also analyzed for comparison with SL. A melanin dispersing effect was observed in fish exposed to a BB, while a concentrating one was observed in those exposed to a WB. By Western blot, three SL-immunoreactive (ir) bands (32, 28 and 23.5 kD) were evidenced. Pituitary SL-ir levels were 2.66- and 2.67-fold greater in the 32 Kd and 28 kD bands, respectively, in BB fish compared with those of WB fish. The SL-ir 23.5 Kd band was not included in the analysis because of its unknown identity. In addition, SL-ir cell number and area were significantly higher in the BB condition (BB 22.73+/-1.46, WB 7.37+/-0.54 and BB 27.39+/-1.00 microm2; WB: 16.61+/-0.65 microm2). No significant differences were observed in the number of the hypothalamic MCH-ir neurons. However, a significant difference was observed in their nuclear area (BB 11.61+/-0.42 microm2, WB 17.80+/-0.84 microm2). alphaMSH-ir cells showed a marked increased in number (BB 35.96+/-1.22, WB 24.36+/-1.04), but no significant differences were observed in the cell area. In conclusion, this study presented clear evidence towards a possible involvement of SL in the adaptation to background colors in teleost together with alphaMSH and MCH.

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The distribution of cells that express three prepro-gonadotropin-releasing hormones (GnRH), corresponding to salmon GnRH, sea bream GnRH (sbGnRH), and chicken II GnRH, was studied in the brain and pituitary of the South American cichlid fish, Cichlasoma dimerus. Although the ontogeny and distribution of GnRH neuronal systems have previously been examined immunohistochemically with antibodies and antisera against the various GnRH decapeptides, we have used antisera against various perciform GnRH-associated peptides (GAPs) and riboprobes to various perciform GnRH+GAPs. The results demonstrate that: (1) the GnRH neuronal populations in the forebrain (salmon and sea bream GAPs; sGAP and sbGAP, respectively) show an overlapping pattern along the olfactory bulbs, nucleus olfacto-retinalis, ventral telencephalon, and preoptic area; (2) projections with sGAP are mainly located in the forebrain and contribute to the pituitary innervation, with projections containing chicken GAP II being mainly distributed along the mid and hindbrain and not contributing to pituitary innervation, whereas sbGAP projections are restricted to the ventral forebrain, being the most important molecular form in relation to pituitary innervation; (3) sbGnRH (GnRH I) neurons have an olfactory origin; (4) GAP antibodies and GAP riboprobes are valuable tools for the study of various GnRH systems, by avoiding the cross-reactivity problems that occur when using GnRH antibodies and GnRH riboprobes alone.

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During the last decade, special attention has been focused on the consequences of exposure to environmental estrogens on reproduction in wild fish species. For this reason, characterization of biomarkers of such exposures could result in useful tools for these studies. The detection of vitellogenin (Vtg), a precursor of yolk proteins, is being intensely studied since its synthesis in the liver is regulated by the estradiol-17beta and is influenced by other estrogenic compounds. The aim of this work was to assess the presence of Vtg in the surface mucus of males of Cichlasoma dimerus (Teleostei, Perciformes), a typical South American freshwater cichlid, after hormonal treatment with estradiol-17beta (intraperitoneal injections of 10 microg E(2)/g fish). Plasma and mucus from females and treated males analyzed by Western blot revealed that different heterologous antisera against Vtg bind to putative protein of 180 & 120 kDa and 120 & 110 kDa, respectively, whereas no reaction was found in samples of untreated males. The same profile was observed in mucus samples using Dot blot, a very easy and direct technique. Using immunocytochemistry techniques, immunoreactive Vtg (ir-Vtg) producing cells in the liver of females and treated males were detected. Testes and liver tissues were also assessed by histological techniques. Marked alterations in both organs were observed, such as lower sperm production, presence of immature germ cells in the lobular lumen of testes, and some morphology changes in the hepatocytes due to the accumulation of Vtg. This is the first report about the effects of an estrogen in the Vtg synthesis and their consequences on liver and gonads of a South American fresh water cichlid. These results also support the possibility of using Vtg from surface mucus as a potential biomarker for estrogenic compounds through a noninvasive, useful and easy assay to monitor the presence of endocrine disrupting chemicals in the environment.

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Synbranchus marmoratus (Synbranchidae), commonly known as the swamp eel, is a protogynous diandric teleost fish widely distributed throughout South America. The purpose of this work was to study the ovarian anatomy and to describe oocyte developmental stages in the swamp eel, Synbranchus marmoratus. S. marmoratus has a unique sacular ovary. It is covered by a conspicuous muscular wall, probably involved in an egg-releasing system acting as a peristaltic-like mechanism. The internal ovarian anatomy shows a U-shaped ovarian lamella delimiting a dorsal ovarian lumen. The microscopic study shows evidence of the existence of a germinal epithelium in the inner surface of the lamella, which contains germinal cells, pre-follicular cells and epithelial cells. The complete oogenesis process is divided into four stages: oogonia, primary growth, cortical alveoli and vitellogenesis. Besides, the ovulated oocytes, and atretic structures were described. The structure of the micropyle was studied by scanning electron microscopy (MEB). Near the animal pole the vitelline envelope forms crests that fuse together becoming furrow-like structures with a slightly spiraled direction that converge into the micropyle pit where is located the micropylar canal. Although the sex reversal process of Synbranchids has been subject of many studies, this is the first complete description of the ovarian anatomy and oogenesis.

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Synbranchus marmoratus (Synbranchidae), commonly known as the swamp eel, is a protogynous diandric teleost fish widely distributed throughout South America. The purpose of this work was to study the ovarian anatomy and to describe oocyte developmental stages in the swamp eel, Synbranchus marmoratus. S. marmoratus has a unique sacular ovary. It is covered by a conspicuous muscular wall, probably involved in an egg-releasing system acting as a peristaltic-like mechanism. The internal ovarian anatomy shows a U-shaped ovarian lamella delimiting a dorsal ovarian lumen. The microscopic study shows evidence of the existence of a germinal epithelium in the inner surface of the lamella, which contains germinal cells, pre-follicular cells and epithelial cells. The complete oogenesis process is divided into four stages: oogonia, primary growth, cortical alveoli and vitellogenesis. Besides, the ovulated oocytes, and atretic structures were described. The structure of the micropyle was studied by scanning electron microscopy (MEB). Near the animal pole the vitelline envelope forms crests that fuse together becoming furrow-like structures with a slightly spiraled direction that converge into the micropyle pit where is located the micropylar canal. Although the sex reversal process of Synbranchids has been subject of many studies, this is the first complete description of the ovarian anatomy and oogenesis.

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O ovário do mantídeo Coptopteryx viridis foi estudado por meio de técnicas histológicas e histoquímicas. Cada ovaríolo (tipo panoístico) contém oócitos em diferentes estádios de desenvolvimento, envolvidos por células foliculares. Estas células modificam suas estruturas durante os distintos estágios de oogênese. Descrevem-se as diferentes fases meióticas e os fenômenos das emissöes de nucléolos. A formaçäo de membrana vitelina começa quado o oócito encontra-se em um estádio avançado de vitelogênese. O endocórion e o exocórion säo formados como resultado da atividade do epitélio folicular quando a vitelogênese está terminada

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Descreve-se a ultra-estrutura e a complexidade localizada das membranas da casca do ovo de Coptopteryx viridis. Säo observadas três camadas: exocórion, endocório e membrana vitelina. Das três camadas, o exocórion é a mais espessa (4,8 a 26,4 micronm) e compreende quatro áreas morfologicamente difeentes; o endocórion cuja espessura média é de 2 micronm apresenta três subcamadas, a mais interna e homogênea de 0.05 micronm, e a mais externa de 1 micronm, estando separadas por pilares de 0,95 micronm de altura, distanciados um de outro por 0,14 micronm. A membrana vitelina, com uma espessura de 0,60 micronm, apresenta-se com um aspecto granuloso e irregular sem qualquer padräo reconhecível. Propöe-se um possível mecanismo respiratório baseado em nossas observaçöes

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Several years of observations of the behavior of the mantid Coptoteryx viridis suggested evidence of parthenogenesis in this species. C. viridis is a solitary, sedentary animal, where the female often kills the male before copulation takes place, and the average male adult life is half that of the female. Virgin females were reared in our laboratory from their hatching to the end of their lives; these laid oothecas. From these oothecas, parthenogenetic nymphs were born, all of the female sex and with a very low viability. The karyo-type of the non-parthenogenetic individuals of this species was found to be XO-XX with a diploid number of 27 chromosomes for the male and 28 for the female. The autosomes were acrocentric or "t" type while the X chromosome was subtelocentric or "st" type, according to Levan's classification.