Respiration and the efficiency of energy during embryonic development of Asellus aquaticus L. (Crustacea) [Translation from: Vestsi Akademii Navuk Belorusskoi SSR, Seriya Biyal. 1978(5) 112-114, 1978]

Experimental studies were carried out on the transformation of matter and energy during embryonic development of the isopod Crustacea Asellus aquaticus L. under two constant temperatures. Material for the experiments was collected in the quarry lakes of the Kurasovshchin-zone (city-Minsk) at 9°C in April and at 18°C in June, 1977. The rate of consumption of oxygen per specimen by eggs and embryos of Asellus aquaticus was observed and dynamics of the basic biological characteristics during embryonic development studied.

Part I. Regulation of development in the cellular slime mold Dictyostelium discoideum. Part II. Polysomes and RNA synthesis during early development of the surf clam Spisula solidissima

Part I. The cellular slime mold Dictyostelium discoideum is a simple eukaryote which undergoes a multi-cellular developmental process. Single cell myxamoebae divide vegetatively in the presence of a food source. When the food is depleted or removed, the cells aggregate, forming a migrating pseudoplasmodium which differentiates into a fruiting body containing stalk and spore cells. I have shown that during the developmental cycle glycogen phosphorylase, aminopeptidase, and alanine transaminase are developmentally regulated, that is their specific activities increased at a specific time in the developmental cycle. Phosphorylase activity is undetectable in developing cells until mid-aggregation whereupon it increases and reaches a maximum at mid-culmination. Thereafter the enzyme disappears. Actinomycin D and cycloheximide studies as well as studies with morphologically aberrant and temporally deranged mutants indicate that prior RNA and concomitant protein synthesis are necessary for the rise and decrease in activity and support the view that the appearance of the enzyme is regulated at the transcriptional level. Aminopeptidase and alanine transaminase increase 3 fold starting at starvation and reach maximum activity at 18 and 5 hours respectively.

The cellular DNA s of D. discoideum were characterized by CsC1 buoyant density gradient centrifugation and by renaturation kinetics. Whole cell DNA exhibits three bands in CsCl: ρ = 1.676 g/cc (nuclear main band), 1.687 (nuclear satellite), and 1.682 (mitochondrial). Reassociation kinetics at a criterion of Tm -23°C indicates that the nuclear reiterated sequences make up 30% of the genome (Cot_1/2 (pure) 0.28) and the single-copy DNA 70% (Cot_1/2(pure) 70). The complexity of the nuclear genome is 30 x 10⁹ daltons and that of the mitochondrial DNA is 35-40 x 10⁶ daltons (Cot_1/2 0.15). rRNA cistrons constitute 2.2% of nuclear DNA and have a ρ = 1.682.

RNA extracted from 4 stages during developmental cycle of Dictyostelium was hybridized with purified single-copy nuclear DNA. The hybrids had properties indicative of single-copy DNA-RNA hybrids. These studies indicate that there are, during development, qualitative and quantitative changes in the portion of the single-copy of the genome transcribed. Overall, 56% of the genome is represented by transcripts between the amoeba and mid-culmination stages. Some 19% are sequences which are represented at all stages while 37% of the genome consists of stage specific sequences.

Part II. RNA and protein synthesis and polysome formation were studied during early development of the surf clam Spisula solidissima embryos. The oocyte has a small number of polysomes and a low but measurable rate of protein synthesis (leucine-³H incorporation). After fertilization, there is a continual increase in the percentage of ribosomes sedimenting in the polysome region. Newly synthesized RNA (uridine-5-³H incorporation) was found in polysomes as early as the 2-cell stage. During cleavage, the newly formed RNA is associated mainly with the light polysomes.

RNA extracted from polysomes labeled at the 4-cell stage is polydisperse, nonribosomal, and non-4 S. Actinomycin D causes a reduction of about 30% of the polysomes formed between fertilization and the 16-cell stage.

In the early cleavage stages the light polysomes are mostly affected by actinomycin.

Mussel eggs as indicators of mutagen exposure in coastal and estuarine environments

The aim of this study was to develop a short-term genotoxicity assay for monitoring the marine environment for mutagens. Based on the developing eggs and embryos of the marine mussel Mytilus edulis, an important pollution indicator species, the test employs the sensitive sister chromatid exchange (SCE) technique as its end-point, and exploits the potential of mussel eggs to accumulate mutagenic pollutants from the surrounding sea water. Mussel eggs take up to 6 months to develop while in the gonad, which provides scope for DNA damage to be accumulated over an extended time interval; chromosome damage is subsequently visualised as SCEs in 2-cell-stage embryos after these have been spawned in the laboratory. Methods which measure biological responses to pollutant exposure are able to integrate all the factors (internal and external) which contribute to the exposure. The new cytogenetic assay allows the effects of adult exposure to be interpreted in cells destined to become part of the next generation.

Cultura de tecidos desdiferenciados e embriões somáticos de Petiveria alliacea L. visando à produção de substâncias bioativas

Petiveria alliacea L. pertence à família Phytolaccacceae e é conhecida popularmente como guiné ou amansa-senhor, entre outros nomes. Tem sido muito utilizada na medicina popular como agente terapêutico, devido à diversas propriedades farmacológicas. Estudos fitoquímicos têm contribuído para a descoberta de grande variedade de substâncias biologicamente ativas produzidas em diferentes partes da planta (saponinas, alcalóides, flavonóides, sulfetos, taninos, cumarinas, entre outros). A análise química da raiz tem revelado grande quantidade de derivados sulfurados, principalmente o dibenzil trissulfeto (DTS), com atividade antifúngica, antibacteriana, antioxidante e anticancerígena. Visando avaliar a produção biotecnológica do DTS, o presente trabalho teve como objetivo, otimizar a cultura de novas linhagens de calos, células em suspensão e embriões somáticos, a partir de plantas de P. alliacea L. mantidas in vitro, com o monitoramento da capacidade biossintética das culturas. Os resultados mostraram que a produção de calos friáveis foi possível em explantes foliares inoculados em meio MS suplementado com PIC ou 2,4-D. Além da resposta calogênica, foi observada a produção de estruturas globulares caracterizadas como embriões somáticos. A ocorrência de embriogênese somática direta foi confirmada através da análise histológica do processo regenerativo. A indução de embriões somáticos gerou um processo de embriogênese secundária altamente repetitivo até 150 dias de cultura e conversão a plantas em freqüência de 5%. Em relação à cultura de células em suspensão a partir dos calos friáveis, observou-se uma diminuição do crescimento celular ao longo das subculturas. As culturas em suspensão originadas de tecido embriogênico secundário continuaram o processo repetitivo em meio líquido e apresentaram conversão a plantas em taxas mais baixas que as obtidas em meio sólido. A obtenção de plantas completas a partir dos embriões somáticos demonstrou a possibilidade de utilização desse sistema para a micropropagação dessa espécie. O monitoramento fitoquímico dos sistemas de cultura in vitro e plantas de campo mantidas em casa de vegetação durante 02 anos apresentou diferenças significativas, confirmando que a cultura de tecidos pode alterar as rotas metabólicas. A cromatografia gasosa acoplada à espectrometria de massas realizada com extrato em diclorometano de embriões secos e hexânico de embriões frescos e raízes secas de plantas provenientes de embriões somáticos, demonstrou a presença do DTS, constituindo, portanto, sistemas in vitro importantes para a modulação desta substância.

O envolvimento do estresse oxidativo e nitrosativo em modelo de doença pulmonar obstrutiva crônica (DPOC) induzido por elastase

Esse estudo buscou investigar o papel do estresse oxidativo e nitrosativo no enfisema pulmonar induzido por elastase. Foram utilizados camundongos machos C57BL/6 submetidos a dois modelos de indução do enfisema por elastase pancreática suína (PPE): intratraqueal (i.t.) e intranasal (i.n.). No modelo intratraqueal a PPE foi instilada nas doses de 0,05 U ou 0,5 U/camundongo para avaliação temporal do enfisema 7, 14 e 21 dias após instilação de PPE. Em outra etapa, o papel da iNOS foi avaliado através da sua inibição farmacológica por aminoguanidina (AMG) 1% na água de beber ou pela sua exclusão genética em camundongos deficientes em iNOS que tiveram o enfisema induzido por 0,5 U PPE i.t. após 21 dias. No modelo intranasal a dose de PPE foi 3 U/camundongo para avaliação temporal do enfisema (1, 7, 14 e 21 dias após PPE). O papel do estresse oxidativo e nitrosativo foi avaliado com diferentes tratamentos antioxidantes na água de beber: tempol, apocinina+alopurinol, n-acetilcisteína, vitamina C+E, e aminoguanidina durante os 21 dias de indução do enfisema. Os grupos controles foram submetidos à instilação de salina. Lavado broncoalveolar, imunoensaios, análises bioquímicas de estresse oxidativo e ensaios morfométricos foram realizados nos pulmões dos animais. O enfisema foi histologicamente alcançado em 21 dias após 0,5 U PPE i.t., evidenciado pelo aumento do diâmetro alveolar médio Lm e da densidade de volume dos espaços alveolares - Vvair em comparação ao grupo controle. TNF-α foi aumentado em 7 e 14 dias após 0,5 U PPE comparados ao controle, concomitante com a redução de IL-10 nos mesmos períodos, comparados ao controle. O estresse oxidativo foi observado na fase inicial do enfisema, com aumento dos níveis de nitrito, TBARS e superóxido dismutase no grupo 7 dias após 0,5 U PPE (i.t.) quando comparados ao controle ao passo que no modelo intranasal as alterações típicas do estresse foram vistas no grupo 1 dia após 3 U de PPE. Atividade da glutationa peroxidase foi aumentada em todos os grupos PPE (i.t.). A exposição à 0,5 U PPE induziu o aumento da iNOS, eNOS e nitrotirosina, sendo revertido no grupo PPE+AMG. Os animais tratados com AMG 1% e os deficientes em iNOS tiveram o enfisema atenuado histologicamente, mantendo o Lm e o Vvair semelhantes ao grupo controle. Os grupos tratados com n-acetilcisteína e aminoguanidina no modelo i.n. tiveram redução do Lm quando comparados ao grupo PPE. Esses resultados sugerem que as vias de estresse oxidativo e nitrosativo são disparadas pela produção de óxido nítrico via iNOS no enfisema pulmonar. A modulação da iNOS parece uma estratégia promissora no estabelecimento do enfisema pulmonar

Evaluation of marine phytoplankton toxicity by application of marine invertebrate bioassays

The dinoflagellate Alexandrium minutum and the haptophyte Prymnesium parvum are well known for their toxin production and negative effects in marine coastal environments. A. minutum produces toxins which cause paralytic shellfish poisoning in humans and can affect copepods, shellfish and other marine organisms. Toxins of P. parvum are associated with massive fish mortalities resulting in negative impacts on the marine ecosystem and large economic losses in commercial aquaculture. The aim of this work is to improve our knowledge about the reliability of the use of marine invertebrate bioassays to detect microalgae toxicity, by performing: (i) a 24- to 48-h test with the brine shrimp Artemia franciscana; (ii) a 48-hour embryo-larval toxicity test with the sea urchin Paracentrotus lividus; and (iii) a 72-h test with the amphipod Corophium multisetosum. The results indicate that A. franciscana and P. lividus larvae are sensitive to the toxicity of A. minutum and P. parvum. LC50 comparison analysis between the tested organisms reveals that A. franciscana is the most sensitive organism for A. minutum. These findings suggest that the use of different organizational biological level bioassays appears to be a suitable tool for A. minutum and P. parvum toxicity assessment.

Restrição proteica materna e alteração do desenvolvimento das artérias coronárias em camundongos

O desenvolvimento da programação fetal é considerado um importante fator de risco para doenças não-transmissíveis da vida adulta, incluindo doença cardíaca coronariana. Com o objetivo de investigar a associação entre nutrição materna e o desenvolvimento das artérias coronárias (AC) em embriões de camundongos estadiados; embriões de camundongos C57BL/6 nos estádios de 16-23 foram retirados de mães alimentadas com dietas de proteína normal (NP) ou de baixa proteína (LP), e as AC foram estudadas. Embora os embriões LP possuam massa corporal menor, entretanto tinham taxas de crescimento cardíaco maior, quando comparados com os embriões NP. O Plexo subepicárdico foi observado no início do período pós-somítico (estádio 16) de embriões NP, enquanto que nos embriões LP apenas no estádio 17 (P <0,01), persistindo até o estádio 18 (P <0,01). As artérias coronárias foram detectadas inicialmente no estádio18 dos embriões NP, já nos embriões LP foram encontradas a partir do estádio 19 (P <0,01). Núcleos apoptóticos foram observados em torno do anel aórtico peritruncal no estádio 18 em embriões NP e LP. Células FLK1+ (Fetal Liver Kinase 1 = VEGFr2 = Vascular Endothelial Growth Factor Receptor 2) apresentaram uma distribuição homogênea nos embriões NP já no estádio 18, enquanto uma distribuição semelhante nos embriões LP foi visto apenas nos estádios 22 e 23. A restrição proteica materna em camundongos leva a um atraso no crescimento do coração no período embrionário modificando o desenvolvimento do plexo peritruncal subepicárdica e diminuindo a taxa de apoptose na região do futuro orifício coronariano.

Estudo fenotípico da região Aorta-Gônada-Mesonefros, de embriões de galinhas Gallus gallus domesticus, com ênfase na aorta dorsal

Acredita-se que os primeiros progenitores da hematopoese definitiva surjam da diferenciação do endotélio da aorta dorsal, na altura da região da Aorta-Gônada-Mesonefros (AGM). Com o intuito de estudar esta região e o fenótipo das células do endotélio da aorta dorsal nesta posição topográfica, ovos galados de Gallus gallus domesticus L. foram incubados em chocadeira, classificados em estádios de E16 a E25 e processados histotecnologicamente para obtenção de secções seriadas na altura da região AGM. Estas passaram por coloração por Hematoxilina-Eosina, histoquímica para PAS, PAS-diastase e Alcian Blue pH 1.0 e pH 2.5, histoquímica por lectinas fluoresceinadas e imunofluorescência para moléculas de superfície, citoesqueleto e matriz extracelular. Foi observada hipertrofia endotelial no assoalho da aorta nos estádios observados, o qual se apresentava positivo ao PAS, com ocorrência frequente de vacuolizações basais PAS negativas, e o surgimento ocasional de grupamentos celulares intravasculares. Nestes, as células que se destacavam da membrana basal do endotélio expressavam progressivamente mais material PAS positivo, o qual, no entanto, em nenhum momento pareceu se tratar de glicogênio. Em relação às glicosaminoglicanas, notamos a presença predominante de ácido hialurônico por todo o mesênquima da região e em outras estruturas como periferia da notocorda, tubo neural e mesoderma lateral. Ocorreu co-expressão de fibronectina e α-actina de músculo liso em células circunjacentes à aorta, na face ventral do vaso. GFAP e BMP-4 são expressas entre as células do tubo neural e em sua periferia, assim como na notocorda do embrião. As lectinas Abrus precatorius, Lens culinarise Ricinus communis mostraram-se positivas principalmente na região subedotelial do assoalho da aorta nos estádios observados neste trabalho. Bandeiraea simplicifolia exibiu pouca marcação na aorta dorsal e a Arachis hypogeae foi negativa. Outras estruturas da região AGM também expressaram resíduos de açúcares revelados por estas lectinas, tais como: notocorda, tubo neural, mesênquima, intestino primitivo e saco vitelínico. Estes resultados acrescentam elementos morfológicos e bioquímicos ao conhecimento sobre a região AGM de embriões de galinha e sobre o endotélio, possivelmente hemogênico, da aorta dorsal.

Effects of maternal age and size on embryonic energy reserves, developmental timing, and fecundity in quillback rockfish (Sebastes maliger)

Maternal effects on the quality of progeny can have direct impacts on population productivity. Rockfish are viviparous and the oil globule size of larvae at parturition has been shown to have direct effects on time until starvation and growth rate. We sampled embryos and preparturition larvae opportunistically from 89 gravid quillback rockfish (Sebastes maliger) in Southeast Alaska. Because the developmental stage and sampling period were correlated with oil globule size, they were treated as covariates in an analysis of maternal age, length, and weight effects on oil globule size. Maternal factors were related to developmental timing for almost all sampling periods, indicating that older, longer, and heavier females develop embryos earlier than younger, shorter, or lighter ones. Oil globule diameter and maternal length and weight were statistically linked, but the relationships may not be biologically significant. Weight-specific fecundity did not increase with maternal size or age, suggesting that reproductive output does not increase more quickly as fish age and grow. Age or size truncation of a rockfish population, in which timing of parturition is related to age and size, could result in a shorter parturition season. This shortening of the parturition season could make the population vulnerable to fluctuating environmental conditions.

Ontogenetic patterns and temperature-dependent growth rates in early life stages of Pacific cod (Gadus macrocephalus)

Pacific cod (Gadus macrocephalus) is an important component of fisheries and food webs in the North Pacific Ocean and Bering Sea. However, vital rates of early life stages of this species have yet to be described in detail. We determined the thermal sensitivity of growth rates of embryos, preflexion and postflexion larvae, and postsettlement juveniles. Growth rates (length and mass) at each ontogenetic stage were measured in three replicate tanks at four to five temperatures. Nonlinear regression was used to obtain parameters for independent stage-specific growth functions and a unified size- and temperature-dependent growth function. Specific growth rates increased with temperature at all stages and generally decreased with increases in body size. However, these analyses revealed a departure from a strict size-based allometry in growth patterns, as reduced growth rates were observed among preflexion larvae: the reduction in specific growth rate between embryos and free-swimming larvae was greater than expected based on body size differences. Growth reductions in the preflexion larvae appear to be associated with increased metabolic rates and the transition from endogenous to exogenous feeding. In future studies, experiments should be integrated across life transitions to more clearly define intrinsic ontogenetic and size-dependent growth patterns because these are critical for evaluations of spatial and temporal variation in habitat quality.

Maturity and growth of female dusky rockfish (Sebastes variabilis) in the central Gulf of Alaska

The dusky rockfish (Sebastes variabilis) has recently been resurrected as a distinct species in the genus Sebastes. Reproductive biology and growth were examined for this redescribed species in the central Gulf of Alaska. Age and length at 50% maturity were 9.2 years and 365 mm fork length, respectively, which are lower than previously reported. Fertilized ova and eyed embryos were observed in April and evidence of postparturition was not observed until May. The gonadosomatic index decreased with the onset of postparturition in May. Von Bertalanffy growth parameters for female dusky rockfish, estimated from the maturity samples, were significantly different from growth parameters derived from Gulf of Alaska fishery-independent survey data.

Changes in body composition and fatty acid profile during embryogenesis of quillback rockfish (Sebastes maliger)

We investigated developmental changes in the body compositions and fatty acid (FA) profiles of embryos and preparturition larvae of the quillback rockfish (Sebastes maliger). Comparisons of proximate composition data from early-stage embryos with data from hatched preparturition larvae taken from wild-caught gravid females indicated that embryos gain over one-third their weight in moisture while consuming 20% of their dry tissue mass for energy as they develop into larvae. Lipid contributed 60% of the energy consumed and was depleted more rapidly than protein, indicating a protein-sparing effect. Oil globule volume was strongly correlated with lipid levels, affirming its utility as an indicator of energetic status. FA profiles of early embryos differed significantly from those of hatched larvae. Differences in the relative abundances of FAs between early embryos and hatched larvae indicated different FA depletion rates during embryonic development. We conclude that some metabolically important FAs may prove useful in assessing the condition of embryos and preparturition larvae, particularly 20:4n-6, which cannot be synthesized by many marine fish and which is conserved during embryogenesis. Variability in body composition and energy use among rockfish species should be considered when interpreting any measures of condition.

All their eggs in one basket: a rocky reef nursery for the longnose skate (Raja rhina Jordan & Gilbert, 1880) in the southern California Bight

Skates (family Rajidae) are oviparous and lay tough, thick-walled eggs. At least some skate species lay their eggs in spatially restricted nursery grounds where embryos develop and hatch (Hitz, 1964; Hoff, 2007). After hatching, neonates may quickly leave the nursery grounds (Hoff, 2007). Egg densities in these small areas may be quite high. As an example, in the eastern Bering Sea, a site <2 km2 harbored eggs of Alaska skate (Bathyraja parmifera) exceeding 500,000/km2. All skate nursery grounds have been identified over soft sea floors (Lucifora and García, 2004; Hoff, 2007).

A nursery site of the Alaska skate (Bathyraja parmifera) in the eastern Bering Sea

A nursery site for the Alaska skate (Bathyraja parmifera) was sampled seasonally from June 2004 to July 2005. At the small nursery site (~2 km2), located in a highly productive area near the shelf-slope interface at the head of Bering Canyon in the eastern Bering Sea, reproductive males and females dominated the catch and neonate and juvenile skates were rare. Seasonal samples showed summertime (June and July) as the peak reproductive time in the nursery although some reproduction occurred throughout the year. Timeseries analysis of embryo length frequencies revealed that three cohorts were developing simultaneously and the period of embryonic development was estimated at 3.5 years and average embryo growth rate at 0.2 mm/day. Estimated egg case deposition occurred mainly during summertime and hatching occurred during winter months. Protracted hatching times may be common for oviparous elasmobranch species and may be directly correlated with ambient temperatures as evident from a meta-data analysis. Evidence indicates that the Alaska skate uses the eastern Bering Sea outer continental shelf region for reproduction and the middle and inner shelf regions as habitat for immature and subadults. Skate nurseries may be vulnerable to disturbances because they are located in highly productive areas and because embryos develop slowly.

Exploitation and reproduction of the spotted eagle ray (Aetobatus narinari) in the Los Frailes Archipelago, Venezuela

We studied a small artisanal fishery for the spotted eagle ray (Aetobatus narinari) off Margarita Island in northeastern Venezuela. We analyzed data from 413 fishing trips directed at A. narinari over a 29-month sampling period (August 2005–December 2007). These trips yielded 55.9 metric tons and 1352 individuals from which a subsample of 846 females and 321 males was used for biological data. Maximum fishing effort and landings occurred between February and May, and catch per unit of effort was highest between December and February and between July and October with an overall average of 3 individuals and 133 kg per trip. The overall sex ratio was significantly different from 1:1 with a predominance of females. Females ranged in size with disc widths (DW) from 64 to 226 cm. Males ranged in size between 97 and 190 cm DW. There was no statistically significant difference between male and female length-weight relationships. Mean fecundity was estimated at 3.09 embryos per female, and the largest embryo measured 44.5 cm DW. Females in different maturity stages were found in all months, except November 2007, the month when all females were immature. Postgravid females occurred mainly during the periods of January–May and July–October. Mean length (L50) at maturity was estimated at 129.2 cm DW for males and 134.9 cm DW for females. This study provides much needed information on the biology and life history of A. narinari for the management of an intensive, directed, small-scale fishery for this little known species in northeastern Venezuela.