Descrição morfológica dos ovários do peixe Melanotaenia boesemani em atividade reprodutiva / Morphological description of Melanotaenia boesemani ovaries in reproductive activity

Estudou-se o tecido germinativo ovariano do peixe Melanotaenia boesemani. Por meio de análises morfológicas das gônadas, realizadas após a confecção de lâminas histológicas elaboradas em parafina e coradas com hematoxilina-eosina, foram descritos os tipos celulares encontrados e realizada a classificação do estádio de maturidade dos peixes. Macroscopicamente, foi identificado que a referida espécie possui ovário único, arredondado e, na fase observada, amarelado, localizado na parte látero-anterior da cavidade celomática. As fêmeas analisadas foram classificadas na escala de desenvolvimento gonadal como maduro/em reprodução. A presença de folículos vazios e marcas de desova nas lamelas ovulígeras mostrou-se evidente. Nos ovócitos vitelogênicos deste peixe, foi constatada a presença de filamentos de adesão ovocitária ancorados à zona pelúcida. Os ovários apresentaram aspecto morfológico geral semelhante ao de outros peixes teleósteos. Seu tipo de desova foi classificado como intermitente e o padrão de desenvolvimento de ovócitos, como assincrônico.(AU)

The germinal ovarian tissue of the fish Melanotaenia boesemani was studied. By means of morphological analyses of the gonads, performed after the manufacture of histological slides made of paraffin and stained with hematoxylin-eosin, the cell types found were described and the rank of the maturity stage of the fishes was executed. Macroscopically was found that the species has only one ovary, rounded and in the observed phase yellowish, located in the lateral front part of the coelomic cavity. The observed females were classified in the gonadal development scale as mature/in reproduction. The presence of empty follicles and spawning marks on ovigerous lamellae proved to be evident. In vitellogenic oocytes of this fish, the presence of filaments of oocyte adhesion anchored to the pellucida zona was found. The ovaries showed general morphological appearance similar to that of other teleost fishes. Its spawning type was classified as intermittent and the pattern of development of oocytes as asynchronous.(AU)

Cellular development of the germinal epithelium during the gametogenic cycle of the golden mussel Limnoperna fortunei (Bivalvia: Mytilidae)

AbstractThe golden mussel Limnoperna fortunei is an invasive species that has quickly dispersed and colonized several potential different habitats distributed all over the world, causing environmental and economic impacts. Thus, in order to contribute to a better understanding of the reproductive aspects of L. fortunei, we described the cellular dynamic of the male and female germinal epithelium during the annual reproductive life history of this species, with the use of high resolution histology. An approximate of 1 200 specimens of L. fortunei were sampled periodically on the upper Paraná River floodplain (Brazil), from March 2010 to December 2012. Based on the activity of the germinal epithelium and consequent germ cell development, this study has resulted in the recognition of the following reproductive phases: Developing, Spawning Capable, Regressing and Regenerating. In the characterization of these phases, the following germ cells were described for males: spermatogonia, primary and secondary spermatocytes, spermatids and spermatozoa. Cell nests, oogonia, early prophase oocytes, previtellogenic oocytes and vitellogenic oocytes (early vitellogenic oocytes, middle vitellogenic oocytes and full-grown oocytes) were described for females. The morphological data and reproductive parameters obtained, showed the value of the cellular dynamics of the germinal epithelium, for the understanding of the cyclic gonadal events during the adult reproductive life of the mollusk in general. These results on the gametogenesis of this invasive species may be a fundamental tool for the development of control strategies and programs implementation, to reduce their proliferation and impacts in natural local environments. Rev. Biol. Trop. 64 (2): 521-536. Epub 2016 June 01.

ResumenEl mejillón dorado Limnoperna fortunei (Dunker, 1857) es una especie invasora que se ha dispersado rápidamente y ha colonizado diferentes hábitats potenciales distribuidos por todo el mundo, causando impactos ambientales y económicos. El control de esta especie depende del conocimiento de los aspectos reproductivos. Entre marzo 2010 y diciembre 2012 fueron muestreados periodicamente alrededor de 1 200 especímenes de L. fortunei en la planicie de inundación del Alto Rio Paraná, Brasil. Con el fin de contribuir con una mejor comprensión de los aspectos reproductivos de L. fortunei, se describió la dinámica celular del epitelio germinativo masculino y femenino durante la vida reproductiva anual de esta especie, mediante histología de alta resolución basada en la actividad del epitelio germinal y el consecuente desarrollo de las células germinales. En este estudio se reconocieron las siguientes fases reproductivas: desarrollo, capacidad de desove, regresión y regeneración. En los machos se describieron las siguientes células germinales: espermatogonias, espermatocitos primarios y secundarios, espermátidas y espermatozoides. Mientras que en las hembras se estudiaron: nidos celulares, ovogonias, ovocitos profásicos tempranos, ovocitos pre-vitelogénicos y ovocitos vitelogénicos (ovocitos vitelogénicos tempranos, ovocitos vitelogénicos y ovocitos completamente desarrollados).Los datos morfológicos y parámetros reproductivos que se muestran aquí confirman cómo la dinámica celular del epitelio germinal permite la comprensión de los acontecimientos cíclicos gonadales durante la vida reproductiva de adultos del molusco, en general. El conocimiento de la gametogénesis de esta especie invasora puede convertirse en una herramienta fundamental para el desarrollo de estrategias de control y aplicación de programas para disminuir su proliferación en ambientes naturales.

Signaling Pathways in Insulin- and IGF-I Mediated Oocyte Maturation in Lower Vertebrates.

The endocrine control of oocyte maturation in fish and amphibians has proved to be a valuable model for investigating the rapid and non-genomic steroid actions at the cell surface. Considerable progress has made over the last decade in elucidating signaling pathways in steroid-induced oocyte maturation. In addition to steroids, various growth factors have also been reported to be involved in this process and progress being made to elucidate their mechanism of actions. Exposure of fully-grown oocytes to steroids or growth factors (insulin/IGFs) initiates various signaling cascade, leading to formation and activation of maturation-promoting factor (MPF), a key enzyme that catalyzes entry into M-phase of meiosis I and II. Whereas the function of MPF in promoting oocyte maturation is ubiquitous, there are differences in signaling pathways between steroids- and growth factors-induced oocyte maturation in amphibian and fish. Here, we have reviewed the recent advances on the signaling pathways in insulin- and IGF-I-induced oocyte maturation in these two groups of non-mammalian vertebrates. New findings demonstrating the involvement of PI3 kinase and MAP kinase in induction of oocyte maturation by insulin and IGF-I are presented.

Gonad maturation of Haemulon plumieri (Teleostei: Haemulidae) in Ceará state, Northeastern Brazil / Maturação gonadal de Haemulon plumieri (Teleostei: Haemulidae) no Nordeste do Brasil

To elucidate the reproductive cycle of any animal, it is first necessary to understand the development of the germ cells and how this process occurs throughout the reproductive tract. Thus, the aim of the present study was to understand the oogenesis and spermatogenesis, as well as the ovarian and testis maturation and type of spawning, of Haemulon plumieri. A total of 119 females and 136 males were analysed. Based on micro and macroscopic characteristics of gonads, the following stages of maturation for females were determined: immature, maturing, ripe, partially spawning, spawning and resting. For males, only the stages immature, maturing, ripe and spent were observed. We can conclude that the ovarian development occurs cyclically, in which the female can spawn more than two times during the reproductive season with the spawning “synchronous in more than two groups”. Since males do not present the “rest” stage after copulation, the maturation process is continuous.

Para se conhecer a biologia reprodutiva de qualquer animal, é fundamental o conhecimento prévio do desenvolvimento das células germinativas e como esse processo ocorre ao longo do trato reprodutivo. Desta forma, o objetivo do presente trabalho foi descrever a oogênese e a espermatogênese, bem como o desenvolvimento maturacional dos ovários e testículos, e o tipo de desova do Haemulon plumieri. Foram analisadas 119 fêmeas e 136 machos. A partir das características macro e microscópicas das gônadas, foram estabelecidos os seguintes estágios de maturação para fêmeas: imatura, em maturação, matura, parcialmente desovada, desovada e em repouso. Para os machos, foram identificados apenas os seguintes estágios: imaturo, em maturação, maturo e esvaziado. A partir das análises, pôde-se concluir que o desenvolvimento ovariano é cíclico, em que a fêmea pode desovar mais de duas vezes por estação, por apresentar a desova do tipo ‘sincrônica em mais de dois grupos'. Uma vez que os machos não apresentam o estágio ‘em repouso' após a cópula, o processo de maturação é contínuo.

Histología del ciclo reproductor de hembras del pejelagarto Atractosteus tropicus (Lepisosteiformes: Lepisosteidae) enTabasco, México / Histology of reproductive cycle of tropical gar Atractosteus tropicus females (Lepisosteiformes: Lepisosteidae) in Tabasco, Mexico.

En el sureste de México A. tropicus es una especie de gran importancia ecológica, económica, cultural y para acuicultura, sin embargo, sus poblaciones silvestres han disminuido a causa de diversas actividades antropogénicas, por lo que se caracterizó el ciclo reproductor de A. tropicus, a partir de los cambios en los elementos germinales y somáticos durante la ovogénesis y la estructura de toda la gónada, información valiosa para el mejor manejo y aprovechamiento de la especie. Mensualmente se capturaron con redes de arrastres, seis hembras sexualmente maduras (N=72), durante un ciclo anual (octubre 2003-septiembre 2004), en poblaciones silvestres de la laguna de Pomposu del municipio de Jalpa de Méndez; Tabasco, México. Las hembras fueron sacrificadas por decapitación y los ovarios se procesaron por métodos histológicos convencionales, se determinó el diámetro entre 10-20 ovocitos en los diferentes estadios de desarrollo al azar. Se determinó mensualmente el índice gonadosomático (IGS), volumen gonadal (gV) y el diámetro del folículo (fD). Además, se identificaron ocho estadios de desarrollo de la ovogénesis: I: cromatina nucléolo, II: perinucléolo temprano, III: perinucléolo avanzado, IV: alvéolos corticales, V: vitelogénesis temprana, VI: vitelogénesis media, VII: vitelogénesis avanzada, VIII: maduración final. El ciclo reproductor la constituye cuatro clases: I.-Proliferativa, II.-Maduración inicial, III.-Maduración tardía, IV.-Regresión-proliferación. El análisis de la información obtenida de los valores encontrados del IGS, gV, fD, estadios de la ovogénesis y clases reproductivas, indican que en diciembre se reinicia el crecimiento de los folículos (previtelogenesis), de mar-zo-julio se presenta la captación del vitelo (vitelogenesis) y en agosto-noviembre se presenta la máxima actividad reproductora, al contrastar este patrón con los factores ambientales (temperatura, precipitación) se sugiere que estos pueden funcionar como un gatillo ambiental, en el desarrollo de la ovogénesis y el desove.

In Southeast Mexico, A. tropicus is a species with great ecological, economic, cultural and aquaculture value, however wild populations have diminished due to diverse anthropological causes. The objective of this study was to characterize the reproductive cycle of A. tropicus, with a description of complete gonad structure and the changes in germinal and somatic elements during oogenesis, for better management and use of this species. Six sexually mature females (N=72) were captured monthly with drag nets during one year cycle (October 2003-October 2004) in wild populations of Pomposu lagoon, municipality of Jalpa de Mendez, Tabasco, Mexico. Females were sacrificed by decapitation and the ovaries were processed by standard histological methods; the diameter of 10-20 oocytes taken at random was determined in different stages of development. Gonadosmatic index (GSI), gonadal volume (gV) and follicule diameter (fD) were determined monthly. Among results we could determine eight oogenesis developmental stages: I: chromatin nucleolus stage, II: early perinucleolus stage, III: mid perinucleolus stage, IV: advanced perinucleolus stage, V: early vitellogenesis stage, VI: mid vitellogenesis stage, VII) advanced vitellogenesis stage and VIII: final maturation stage. The reproductive cycle could be divided in four phases: I: proliferative phase, II: initial maturation phase, III: late maturation phase, IV: regressing phase. Analysis of the data obtained from the IGS, gV and fD values, oogenesis stages and reproductive phases, indicate that in December previtellogenesis phase restarts, from March through July yolk deposition (vitellogenesis) occurs, and from August to November the maximum reproductive activity takes place. Contrasting this pattern with local environmental factors such as temperature and precipitation, we suggested that these may serve as an environmental activator in the development of oogenesis and spawning of this species. Rev. Biol. Trop. 60 (4): 1857-1871. Epub 2012 December 01.

Mecanismos fisiológicos e bioquímicos envolvidos na ovogênese / Physiological and biochemical mechanism involved in ovogenesis

O artigo refere-se a uma atualização sobre os mecanismos fisiológicos e bioquímicos envolvidos na ovogênesis de mamífero.(AU)

The article concern with an up to date about physiologic and biochemical mechanism of ovogenesis in mammals.(AU)

Oocyte structure and ultrastructure in the Mexican silverside fish Chirostoma humboldtianum (Atheriniformes: Atherinopsidae)

the structural and ultrastructural features of gonads from endemic Mexican fish have received scarce attention. This study describes the histological and ultrastructural characteristics of the oocyte in Chirostoma humboldtianum. The ovary is asynchronic, and as such, most phases of oocyte development are found in the same ovary. The complete process of oogenesis was divided in five stages: oogonium and folliculogenesis, primary growth, cortical alveoli and lipid inclusions, vitellogenesis and maturation. The presence of big filaments, which appear at the end of primary growth, induces some common follicular adaptation. During primary growth, abundant ribosomes, rough endoplasmic reticulum, and mitochondria are grouped in the cytoplasm. At the end of this stage, the Z1 layer of the chorion is developed, while microvilli start to be evident as well. In the cortical alveoli and lipid droplets phase, intense PAS positive vesicles, some of them containing nucleoid material, are observed in the peripheral cytoplasm and the lipid droplets take a more central position. In vitellogenesis, the proteic yolk accumulates in a centripetal way while the chorion is completely formed. In maturation, the germinal vesicle migrates to the animal pole, meiosis is restored, and there is nuclear breakdown. The oocyte increases its size and holds some oil droplets and a big fluid mass of yolk. On the outside, filaments surround the oocyte completely. Rev. Biol. Trop. 56 (4): 1825-1835. Epub 2008 December 12.

Los aspectos estructurales y ultraestructrurales de las gónadas de peces mexicanos endémicos han sido poco estudiados. En el presente trabajo reportamos las características histológicas y ultraestructurales del ovocito de Chirostoma hulmboldtianum. El ovario es de tipo asincrónico, por ende, la mayoría de las fases del desarrollo del ovocito pueden ser encontradas en el mismo ovario. El desarrollo del ovocito fue dividido en cinco etapas: ovo-gonia y foliculogénesis, crecimiento primario del ovocito, inclusiones lipídicas y gránulos corticales, vitelogénesis y maduración. La presencia de grandes filamentos que aparecen al final de la etapa de crecimiento primario, inducen adaptaciones foliculares. Durante el crecimiento primario, en el citoplasma se encuentran abundantes ribosomas, retículo endoplásmico rugoso y agrupamientos de mitocondrias. Al final de esta etapa, inicia el desarrollo de la capa Z1 del corion, comenzando a ser evidentes las microvellosidades del ovocito. Durante la etapa de inclusiones lipídicas y gránulos corticales, vesículas PAS positivas, algunas de ellas con material nucleoide, se ubican en la periferia del ovocito, mientras que las que contienen material graso toman una posición más central en la célula. Durante la vitelogénesis se presenta una acumulación de vitelo protéico en un sentido centrípeto; durante esta etapa, el corion está completamente formado. En la maduración, la vesicular germinal migra hacia el polo animal, se reinicia la meiosis y se rompe la envoltura nuclear. El ovocito incrementa su tamaño y en el citoplasma se pueden observar algunas gotas de grasa y el vitelo se presenta como una gran masa acuosa. En el exterior, los filamentos rodean completamente al ovocito.

Oocyte structure and ultrastructure in the Mexican silverside fish Chirostoma humboldtianum (Atheriniforme: Atherinopsidae)

The structural and ultrastructural features of gonads from endemic Mexican fish have received scarce attention. This study describes the histological and ultrastructural characteristics of oocyte from Chirostoma humboldtianum. The ovary is asynchronic, and as such, most phases of oocyte development are found in the same ovary. The complete process of oogenesis was divided in five stages: oogonium and folliculogenesis, primary growth, cortical alveoli and lipid inclusions, vitellogenesis, and maturation. The presence of big filaments, which appear at the end of primary growth, induces some common follicular adaptation. During primary growth, abundant ribosomes, the rough endoplasmic reticulum, and mitochondria are grouped in the cytoplasm. At the end of this stage, the Z1 layer of the chorion is developed, while microvilli start to be evident. In the cortical alveoli and lipid droplets phase, intense PAS positive vesicles, some of them containing nucleoid material, are observed in the peripheral cytoplasm and the lipid droplets take a more central position. In vitellogenesis, the proteic yolk accumulates in a centripetal way while the chorion is completely formed. During maturation, the germinal vesicle migrates to the animal pole, meiosis is restored, and there is nuclear breakdown. The oocyte increases its size and holds some oil droplets and a big fluid mass of yolk. On the outside, filaments completely surround the oocyte. Rev. Biol. Trop. 56 (3): 1371-1380. Epub 2008 September 30.

Purification, partial characterization and role in lipid transport to developing oocytes of a novel lipophorin from the cowpea weevil, Callosobruchus maculatus

Lipid transport in arthropods is achieved by highly specialized lipoproteins, which resemble those described in vertebrate blood. Here we describe purification and characterization of the lipid-apolipoprotein complex, lipophorin (Lp), from adults and larvae of the cowpea weevil Callosobruchus maculatus. We also describe the Lp-mediated lipid transfer to developing oocytes. Lps were isolated from homogenates of C. maculatus larvae and adults by potassio bromide gradient and characterized with respect to physicochemical properties and lipid content. The weevil Lp (465 kDa) and larval Lp (585 kDa), with hydrated densities of 1.22 and 1.14 g/mL, contained 34 and 56 percent lipids and 9 and 7 percent carbohydrates, respectively. In both Lps, mannose was the predominant monosaccharide detected by paper chromatography. SDS-PAGE revealed two apolipoproteins in each Lp with molecular masses of 225 kDa (apolipoprotein-I) and 79 kDa (apolipoprotein-II). The lipids were extracted and analyzed by thin-layer chromatography. The major phospholipids found were phosphatidylserine, phosphatidylcholine and phosphatidylethanolamine in adult Lp, and phosphatidylcholine, phosphatidylethanolamine and sphingomyelin in larval Lp. Hydrocarbons, fatty acids and triacylglycerol were the major neutral lipids found in both Lps. Lps labeled in the protein moiety with radioactive iodine (125I-iodine) or in the lipid moiety with fluorescent lipids revealed direct evidence of endocytic uptake of Lps in live oocytes of C. maculatus.

Identification of the stages of ovarian maturation of the Argentine hake Merluccius hubbsi Marini, 1933 (Teleostei: Merlucciidae): advantages and disadvantages of the use of the macroscopic and microscopic scales

The Argentine hake Merluccius hubbsi is a demersal-pelagic species on which few studies have been undertaken, despite its importance for the fisheries of the South-Southeastern Brazilian region, . The species is the most important commercial fishery resource in Uruguay and Argentina, where several studies have permitted the proper monitoring of the species. The ovarian maturation of the Argentine hake is analysed in this study. A scale of maturation is presented in the light of the oocyte development and the use of macro and microscopic scales of ovarian maturation are compared. It was detected that the oocyte types and derived structures present in the ovaries are similar to those already described both for the species and for teleosts in general; group synchronous development and multiple spawning are typical. The identification of ovarian maturation based only on external morphological characteristics of the gonads presents many errors, mainly in the characterization of the stages of maturation, which makes imperative the parallel analysis of the oocyte development, undertaken through histological ovarian cross-sections.

A merluza Merluccius hubbsi é uma espécie demerso-pelágica sobre a qual, apesar de sua importância nas pescarias da Região Sudeste-Sul do Brasil, poucos estudos foram realizados. Neste aspecto, destacam-se o Uruguai e a Argentina, países onde ela é o principal recurso pesqueiro e onde a existência de inúmeros estudos tem permitido o monitoramento da espécie. Neste estudo a maturação ovariana da merluza é analisada. Através do desenvolvimento ovocitário, apresenta-se uma escala de maturação e compara-se o emprego de escalas macro e microscópicas de maturação dos ovários. Constatou-se que os tipos de ovócitos e estruturas derivadas presentes nos ovários são semelhantes aos já descritos para a espécie e para teleósteos; o desenvolvimento dos ovócitos é sincrônico em grupos e a desova é parcelada. A identificação da maturação ovariana com base apenas em características morfológicas externas das gônadas apresenta muitos erros, principalmente na caracterização do estádio em maturação, sendo imprescindível a análise paralela do desenvolvimento ovocitário, realizada através de cortes histológicos dos ovários.

Oogenesis and egg development in triatomines: a biochemical approach

Em triatomíneos, assim como em outros insetos, o acúmulo de vitelo é um processo no qual um tecido extraovariano, o corpo gorduroso, produz proteínas que são empacotadas no interior de um ovo. A principal proteína, sintetizada pelo corpo gorduroso, que é acumulada no interior de um ovócito, é a vitelogenina. Este processo é também conhecido por vitelogênese. Existem crescentes evidências em triatomíneos, que além do corpo gorduroso, o ovário também produz proteínas de vitelo. A forma como estas proteínas de vitelo entram nos ovócitos será aqui comentada. O vitelo é um material complexo composto por proteínas, lipídeos, carboidratos e outros compostos minoritários que são empacotados de uma maneira organizada no interior dos ovócitos. A fertilização dispara a embriogênese, um processo que culmina com o desenvolvimento do embrião. Durante a embriogênese o vitelo será utilizado para a construção de um novo indivíduo, a ninfa de primeiro estádio. O desafio para a próxima década é entender onde e como estas proteínas de vitelo são utilizadas junto com os seus componentes não protéicos, em compasso com o programa genético do embrião, que comanda a diferenciação celular (fase inicial da embriogênese) e diferenciação do embrião (fase final da embriogênese) no interior do ovo.

Seasonal ovarian cycle in freshwater teleost, Labeo rohita (Ham.) in Tarai region of Uttaranchal.

Seasonal ovarian cycle, based on histophysiological changes, and correlative variations in liver were studied in an Indian major carp, Labeo rohita reared in captivity under ambient environmental conditions in Tarai region of Uttaranchal. The oocyte features used for marking ovarian cycle included nuclear-ooplasmic ratio, yolk vesicle incorporation into ooplasm, number and size of nucleolus and differentiation of follicular layer. Based on these features the ovarian cycle in L. rohita was divided into five phases i.e. resting, preparatory, pre-spawning, spawning and post-spawning/regression. The resting and preparatory phases were characterized by the dominating presence of chromatin-nucleolar and yolk-vesicle stage oocytes. The pre-spawning phase ovary was marked by the prevalence of yolk globule stage oocytes having cortical alveoli and differentiating follicular wall. The oocytes containing densely packed yolk-granules and large masses of yolk vesicles and also exhibiting germinal vesicle movement (GVM) signified the ovary of spawning phase. The ovary in regression phase contained pre-ovulatory atretic oocytes and significant number of oogonia. The histophysiological changes in liver showing augmented biosynthetic activity during preparatory and pre-spawning phases seemed well correlated with the gonadal development. The increasing water temperature and day-length apparently was favourable for the gonadal development in L. rohita.

Maturation of pig oocytes in vitro in a medium with pyruvate

The aim of in vitro maturation oocyte systems is to produce oocytes of comparable quality to those derived in vivo. The present study was designed to examine the surface morphological changes of the cumulus-oocyte complex (COC) and nuclear maturation in a culture system containing pyruvate. Ovaries were obtained from a slaughterhouseand transported to the laboratory within 2 h at 35-39°C,and rinsed three times in 0.9 percent NaCl. The COCs were harvested from the ovaries and in vitro maturation was evaluated in San Marcos (SM) medium, a chemically defined culture system containing 22.3 mM sodium pyruvate. Oocytes were cultured in SM, SM + porcine follicular fluid (pFF) and in SM + pFF + gonadotropins (eCG and hCG) for 20-22 h and then without hormonal supplements for an additional 20-22 h. After culture, the degree of cumulus expansion and frequency of nuclear maturation were determined. Oocytes matured in SM (40.9 percent) and SM + pFF (42.9 percent) showed moderate cumulus expansion, whereas oocytes matured in SM + pFF + gonadotropins (54.6 percent) showed high cumulus expansion. The maturation rate of cultured oocytes, measured in function of the presence of the polar corpuscle, did not differ significantly between SM (40.9 ± 3.6 percent) and SM + pFF (42.9 ± 3.7 percent). These results indicate that pig oocytes can be successfully matured in a chemically definedmedium and suggest a possible bifunctional role of pyruvate as an energy substrate and as an antioxidant protecting oocytes against the stress of the in vitro environment.

Vitelline envelope formation during oogenesis in Bufo arenarum

The formation of vitelline envelope (VE) during the oogenesis of Bufo arenarum (Amphibia Anura) is described. At the stage of early vitellogenesis, the first structures appear: the number of oocyte microvilli increases, and many cross sections of them are observed between the follicle cells and the oocyte. A filamentous material is observed inside the follicle cells and between the follicle cells and the oocyte. Multivesicular bodies are also found in the follicle cells, and in the perivitelline space. The micrographs also suggest the participation of the oocyte in the process of VE formation: large vesicles are present in the cortex of the oocyte, filled with an amorphous material of low and uniform electron density. Some of them are in the process of releasing their content to the perivitelline space. Many vesicles (probably resulting from microvilli fragmentation) are also observed in the perivitelline space. During late vitellogenesis the VE is a continuous structure between the layer of follicle cells and the oocyte. The filamentous material is aggregated in bundles, forming a net, and the spherical components are now either included in the orifices of the net, or free near the oocyte's surface. At the end of oogenesis, when the VE is completely formed, it is difficult to distinguish its components independently. Immunolocalization with antibodies against VE, show a positive reaction in follicle cells and oocytes in previtellogenic and full grown ovarian follicles. This analysis suggests that both the oocyte and the follicle cells are directly involved in the synthesis and secretion of the components of the vitelline envelope in Bufo arenarum.