A modulação da via do AMPc/PKA altera a morfologia da oligodendroglia e a distribuição das proteínas CNPase e MAG in vitro

A diferenciação da oligodendroglia depende de alterações coordenadas no citoesqueleto e na sua relação com a membrana plasmática, um componente importante para a formação da bainha de mielina. A 23 nucleotídeo cíclico 3 fosfodiesterase (CNPase) está relacionada com a organização do citoesqueleto, sendo uma proteína ancoradoura de microtúbulos na membrana plasmática. In vitro, a CNPase compõe, com a F-actina e os microtúbulos, as estruturas semelhantes a nervuras ou os componentes radiais. A glicoproteína associada a mielina (MAG), também é importante para a formação dos véus de membrana e está associada a CNPase e a tubulina. Além disso, as três proteínas podem ser reguladas pela via de sinalização do AMPc/PKA. Buscando avaliar os efeitos da via do AMPc/PKA na regulação da diferenciação oligodendroglial, culturas de hemisférios cerebrais com 5 dias foram tratadas por 30 min ou 24 h com o inibidor (SQ22356-SQ [1 M]) ou com o ativador (forscolina [10M]) da adenilato ciclase ou com o inibidor da PKA, H-89 [1 M]. A oligodendroglia foi identificada pelo anticorpo anti-CNPase e por sua morfologia. Com 30 min de tratamento com forscolina, as células das culturas tratadas apresentaram prolongamentos maiores e menos véus de membrana quando comparadas às culturas controle. O tratamento com SQ também causou um aumento no tamanho dos prolongamentos e o tratamento com H-89 causou a redução no tamanho dos prolongamentos e nos véus de membrana. Com 24 h, as células tratadas com forscolina apresentaram poucos prolongamentos, já as culturas tratadas com SQ apresentaram um aumento no tamanho do prolongamento e as tratadas com H-89 demonstraram redução no véu de membrana. Observamos também alterações na distribuição da CNPase, tubulina e MAG, a primeira apresentou uma concentração próxima ao núcleo depois dos dois tempos de tratamento com H-89, o mesmo ocorreu com a tubulina. A CNPase adquiriu ainda um padrão puntiforme depois de 24 h de tratamento com ambos os inibidores. A MAG apresentou um aumento na concentração próximo ao núcleo depois de 30 min de tratamento com forscolina e SQ. O tratamento com SQ também reduziu a distribuição da MAG nos véus de membrana. A mesma redução foi observada depois de 24 h de tratamento com H-89. Esses resultados reforçam a participação da via do AMPc/PKA no desenvolvimento da oligodendroglia, incluindo a formação dos prolongamentos, suas ramificações e ainda a formação dos véus de membrana, com prováveis consequências na formação e manutenção da bainha de mielina.

Avaliação morfofuncional do complexo hipocampal em ratos submetidos a um modelo de hipóxia-isquemia pré-natal

A diminuição do aporte de oxigênio e nutrientes na vida perinatal resulta em danos, como astrogliose, morte de neurônios e de células proliferativas. Déficits cognitivos podem estar relacionados a danos no hipocampo. Neste trabalho avaliamos a citoarquitetura do giro dentado (DG) durante o desenvolvimento e a memória de ratos submetidos à HI. Para tal, utilizamos a técnica de imunohistoquímica para marcador de proliferação celular (KI67), neurônio jovem (DCX), de astrócitos (GFAP) e de óxido nítrico sintase neuronal (NOSn). Para avaliar a memória de curta e de longa duração foi utilizado o teste de reconhecimento de objetos (RO). Ratas Wistar grávidas em E18 foram anestesiadas (tribromoetanol) e as quatro artérias uterinas foram ocluídas com grampos de aneurisma (Grupo HI). Após 45 minutos, os grampos foram removidos e foi feita a sutura por planos anatômicos. Os animais do grupo controle (SHAM) sofreram o mesmo procedimento, excetuando a oclusão das artérias. Os animais nasceram a termo. Animais com idades de 7 a 90 dias pós-natal (P7 a P90), foram anestesiados e perfundido-fixados com paraformaldeído a 4%, e os encéfalos submetidos ao processamento histológico. Cortes coronais do hipocampo (20m) foram submetidos à imunohistoquímica para KI67, DCX, GFAP e NOSn. Animais P90 foram submetidos ao RO. Os procedimentos foram aprovados pelo comitê de ética (CEA/019/2010). Observamos menor imunomarcação para KI67 no giro dentado de animais HI em P7. Para a marcação de DCX nesta idade não foi observada diferença entre os grupos. Animais HI em P15, P20 e P45 tiveram menor imunomarcação para DCX e Ki67 na camada granular. Animais P90 de ambos os grupos não apresentaram marcação para KI67 e DCX. Vimos aumento da imunomarcação para GFAP nos animais HI em todas as idades. A imunomarcação para NOSn nos animais HI foi menor em todas as idades. O maior número de células NOSn positivas foi visto em animais P7 em ambos os grupos na camada polimórfica. Em P15, animais HI apresentam células NOSn+ em todo o DG. Em P30 animais HI apresentam células NOSn+ nas camadas polimórfica e sub-granular. Animais adultos (P90) de ambos os grupos apresentam células NOSn positivas apenas nas camadas granular e sub-granular. Embora animais HI P90 não apresentaram déficits de memória, estes apresentaram menor tempo de exploração do objeto. Comportamento correspondente a déficits de atenção em humanos. Nossos resultados sugerem que HI perinatal diminui a população de células proliferativas, de neurônios jovens, de neurônios NOSn+, além de causar astrogliose e possivelmente déficits de atenção. O modelo demonstrou ser útil para a compreensão dos mecanismos celulares das lesões hipóxico-isquêmicas e pode ser usado para testar estratégias terapêuticas.

Avaliação da indução de lesões no DNA por lasers terapêuticos de baixa potência

Na medicina regenerativa há uma crescente utilização de lasers de baixa intensidade em protocolos terapêuticos para tratamento de doenças em tecidos moles e no tecido ósseo. Lasers emitem feixes de luz com características específicas, nas quais o comprimento de onda, a frequência, potência e modo de missão são propriedades determinantes para as respostas fotofísica, fotoquímica e fotobiológica. Entretanto, sugere-se que lasers de baixa potência induzem a produção de radicais livres, que podem reagir com biomoléculas importantes, como o DNA. Essas reações podem causar lesões e induzir mecanismos de reparo do DNA para preservar a integridade do código genético e homeostase celular. Portanto, o objetivo deste trabalho foi avaliar lesões no DNA de células do sangue periférico de ratos Wistar e a expressão dos genes ERCC1 e ERCC2 em tecidos biológicos expostos a lasers de baixa intensidade em comprimentos de onda, fluências, potências e modos de emissão utilizados em protocolos terapêuticos. Para tal, amostras de sangue periférico foram expostas ao laser vermelho (660 nm) e infravermelho (808 nm) em diferentes fluências, potências e modos de emissão, e a indução de lesões no DNA foi avaliada através do ensaio cometa. Em outros experimentos, lesões no DNA foram analisadas através do ensaio cometa modificado, utilizando as enzimas de reparo: formamidopirimidina DNA glicosilase (FPG) e endonuclease III. Pele e músculo de ratos Wistar foram expostos aos lasers e amostras desses tecidos foram retiradas para extração de RNA, síntese de cDNA e avaliação da expressão dos genes por PCR quantitativo em tempo real. Os dados obtidos neste estudo sugeriram que a exposição aos lasers induz lesões no DNA dependendo da fluência, potência e modo de emissão, e que essas lesões são alvos da FPG e endonuclease III. A expressão relativa do RNAm de ERCC1 e de ERCC2 foi alterada nos tecidos expostos dependendo do comprimento de onda e fluência utilizada. Os resultados obtidos neste estudo sugerem que danos oxidativos no DNA poderiam ser considerados para segurança do paciente e eficácia terapêutica, bem como alterações na expressão dos genes de reparo do DNA participariam dos efeitos de bioestimulação que justificam as aplicações terapêutica de lasers de baixa potência.

Directly modulated wavelength-multiplexed integrated microring laser array

The first demonstration of a directly modulated microring laser array is presented for on-off keyed, wavelength- division- multiplexed fiber-optic data transmission. GaInAsP-InP microring resonators oscillating at separate wavelengths in the 1.5-μm band are vertically coupled to a common passive waveguide bus, which is fabricated on the reverse side of the InP membrane. Two microrings defined with radii for a wavelength channel separation of 6 nm have been assessed for both individual and simultaneous operation. Negligible power penalty (<0.2 dB) is observed for wavelength-division-multiplexed operation with and without transmission over a 25-km fiber span in a manner which indicates low crosstalk between the integrated sources. A device area of less than 0.12 mm2 per microring on a common passive bus allows a highly scalable solution for short-reach wavelength-multiplexed links. © 2008 IEEE.

Reproductive potential of Yellowfin Tuna (Thunnus albacares) in the western Indian Ocean

The reproductive biology of Yellowfin Tuna (Thunnus albacares) in the western Indian Ocean was investigated from samples collected in 2009 and 2010. In our study, 1012 female Yellowfin Tuna were sampled: 320 fish on board a purse seiner and 692 fish at a Seychelles cannery. We assessed the main biological parameters that describe reproductive potential: maturity, spawning seasonality, fish condition, and fecundity. The length at which 50% of the female Yellowfin Tuna population matures (L50) was estimated at 75 cm in fork length (FL) when the maturity threshold was established at the cortical alveolar stage of oocyte development. To enable comparison with previous studies, L50 also was estimated with maturity set at the vitellogenic stage of oocyte development; this assessment resulted in a higher value of L50 at 102 cm FL. The main spawning season, during which asynchrony in reproductive timing among sizes was observed, was November–February and a second peak occurred in June. Smaller females (<100 cm FL) had shorter spawning periods (December to February) than those (November to February and June) of large individuals, and signs of skip-spawning periods were observed among small females. The Yellowfin Tuna followed a “capital-income” breeder strategy during ovarian development, by mobilizing accumulated energy while using incoming energy from feeding. The mean batch fecundity for females 79–147 cm FL was estimated at 3.1 million oocytes, and the mean relative batch fecundity was 74.4 oocytes per gram of gonad-free weight. Our results, obtained with techniques defined more precisely than techniques used in previous studies in this region, provide an improved understanding of the reproductive cycle of Yellowfin Tuna in the western Indian Ocean.

Earthquakes, influenza and cycles of Indian kala-azar.

It is suggested that previous data indicate 3 major epidemics of kala-azar in Assam between 1875 and 1950, with inter-epidemic periods of 30-45 and 20 years. This deviates from the popular view of regular cycles with a 10-20 year period. A deterministic mathematical model of kala-azar is used to find the simplest explanation for the timing of the 3 epidemics, paying particular attention to the role of extrinsic (drugs, natural disasters, other infectious diseases) versus intrinsic (host and vector dynamics, birth and death rates, immunity) processes in provoking the second. We conclude that, whilst widespread influenza in 1918-1919 may have magnified the second epidemic, intrinsic population processes provide the simplest explanation for its timing and synchrony throughout Assam. The model also shows that the second inter-epidemic period is expected to be shorter than the first, even in the absence of extrinsic agents, and highlights the importance of a small fraction of patients becoming chronically infectious (with post kala-azar dermal leishmaniasis) after treatment during an epidemic.

Change in Pacific Northwest coastal ecosystems. Proceedings of the Pacific Northwest Coastal Ecosystems Regional Study Workshop, August 73-14, 1996, Troutdale, Oregon

Over the past one hundred and fifty years, the landscape and ecosystems of the Pacific Northwest coastal region, already subject to many variable natural forces, have been profoundly affected by human activities. In virtually every coastal watershed from the Strait of Juan de Fuca to Cape Mendocino, settlement, exploitation and development of resou?-ces have altered natural ecosystems. Vast, complex forests that once covered the region have been largely replaced by tree plantations or converted to non-forest conditions. Narrow coastal valleys, once filled with wetlands and braided streams that tempered storm runoff and provided salmon habitat, were drained, filled, or have otherwise been altered to create land for agriculture and other uses. Tideflats and saltmarshes in both large and small estuaries were filled for industrial, commercial, and other urban uses. Many estuaries, including that of the Columbia River, have been channeled, deepened, and jettied to provide for safe, reliable navigation. The prodigious rainfall in the region, once buffered by dense vegetation and complex river and stream habitat, now surges down sirfiplified stream channels laden with increased burdens of sediment and debris. Although these and many other changes have occurred incrementally over time and in widely separated areas, their sum can now be seen to have significantly affected the natural productivity of the region and, as a consequence, changed the economic structure of its human communities. This activity has taken place in a region already shaped by many interacting and dynamic natural forces. Large-scale ocean circulation patterns, which vary over long time periods, determine the strength and location of currents along the coast, and thus affect conditions in the nearshore ocean and estuaries throughout the region. Periodic seasonal differences in the weather and ocean act on shorter time scales; winters are typically wet with storms from the southwest while summers tend to be dry with winds from the northwest. Some phenomena are episodic, such as El Nifio events, which alter weather, marine habitats, and the distribution and survival of marine organisms. Other oceanic and atmospheric changes operate more slowly; over time scales of decades, centuries, and longer. Episodic geologic events also punctuate the region, such as volcanic eruptions that discharge widespread blankets of ash, frequent minor earthquakes, and major subduction zone earthquakes each 300 to 500 years that release accumulated tectonic strain, dropping stretches of ocean shoreline, inundating estuaries and coastal valleys, and triggering landslides that reshape stream profiles. While these many natural processes have altered, sometimes dramatically, the Pacific Northwest coastal region, these same processes have formed productive marine and coastal ecosystems, and many of the species in these systems have adapted to the variable environmental conditions of the region to ensure their long-term survival.

Global analysis of mRNA half-lives and de novo transcription in a dinoflagellate, Karenia brevis

Dinoflagellates possess many physiological processes that appear to be under post-transcriptional control. However, the extent to which their genes are regulated post-transcriptionally remains unresolved. To gain insight into the roles of differential mRNA stability and de novo transcription in dinoflagellates, we biosynthetically labeled RNA with 4-thiouracil to isolate newly transcribed and pre-existing RNA pools in Karenia brevis. These isolated fractions were then used for analysis of global mRNA stability and de novo transcription by hybridization to a K. brevis microarray. Global K. brevis mRNA half-lives were calculated from the ratio of newly transcribed to pre-existing RNA for 7086 array features using the online software HALO (Half-life Organizer). Overall, mRNA half-lives were substantially longer than reported in other organisms studied at the global level, ranging from 42 minutes to greater than 144 h, with a median of 33 hours. Consistent with well-documented trends observed in other organisms, housekeeping processes, including energy metabolism and transport, were significantly enriched in the most highly stable messages. Shorter-lived transcripts included a higher proportion of transcriptional regulation, stress response, and other response/regulatory processes. One such family of proteins involved in post-transcriptional regulation in chloroplasts and mitochondria, the pentatricopeptide repeat (PPR) proteins, had dramatically shorter half-lives when compared to the arrayed transcriptome. As transcript abundances for PPR proteins were previously observed to rapidly increase in response to nutrient addition, we queried the newly synthesized RNA pools at 1 and 4 h following nitrate addition to N-depleted cultures. Transcriptome-wide there was little evidence of increases in the rate of de novo transcription during the first 4 h, relative to that in N-depleted cells, and no evidence for increased PPR protein transcription. These results lend support to the growing consensus of post-transcriptional control of gene expression in dinoflagellates.

Diving behavior of immature Steller sea lions (Eumetopias jubatus)

Understanding the ontogenetic relationship between juvenile Steller sea lions (Eumetopias jubatus) and their foraging habitat is key to understanding their relationship to available prey and ultimately their survival. We summarize dive and movement data from 13 young-of-the-year (YOY) and 12 yearling Steller sea lions equipped with satellite dive recorders in the Gulf of Alaska and Aleutian Islands (n=18), and Washington (n=7) from 1994 to 2000. A total of 1413 d of transmission (x =56.5 d, range: 14.5–104.1 d) were received. We recorded 222,073 dives, which had a mean depth of 18.4 m (range of means: 5.8−67.9 m; SD=16.4). Alaska YOY dived for shorter periods and at shallower depths (mean depth=7.7 m, mean duration=0.8 min, mean maximum depth=25.7 m, and maximum depth=252 m) than Alaska yearlings (x =16.6 m, 0=1.1 min, x = 63.4 m, 288 m), whereas Washington yearlings dived the longest and deepest (mean depth=39.4 m, mean duration=1.8 min, mean maximum depth=144.5 m, and maximum depth=328 m). Mean distance for 564 measured trips was 16.6 km; for sea lions ≤10 months of age, trip distance (7.0 km) was significantly less than for those >10 months of age (24.6 km). Mean trip duration for 10 of the 25 sea lions was 12.1 h; for sea lions ≤10 months of age, trip duration was 7.5 h and 18.1 h for those >10 months of age. We identified three movements types: long-range trips (>15 km and >20 h), short-range trips (<15 km and <20 h) during which the animals left and returned to the same site, and transits to other haul-out sites. Long-range trips started around 9 months of age and occurred most frequently around the assumed time of weaning, whereas short-range trips happened almost daily (0.9 trips/day, n=426 trips). Transits began as early as 7 months of age, occurred more often after 9 months of age, and ranged between 6.5 and 454 km. The change in dive characteristics coincided with the assumed onset of weaning. These yearling sea lion movement patterns and dive characteristics suggest that immature Steller sea lions are as capable of making the same types of movements as adults.

Age and growth of the blue shark (Prionace glauca) in the North Atlantic Ocean

Age and growth estimates for the blue shark (Prionace glauca) were derived from 411 vertebral centra and 43 tag-recaptured blue sharks collected in the North Atlantic, ranging in length from 49 to 312 cm fork length (FL). The vertebrae of two oxytetracycline-injected recaptured blue sharks support an annual spring deposition of growth bands in the vertebrae in sharks up to 192 cm FL. Males and females were aged to 16 and 15 years, respectively, and full maturity is attained by 5 years of age in both sexes. Both sexes grew similarly to age seven, when growth rates decreased in males and remained constant in females. Growth rates from tag-recaptured individuals agreed with those derived from vertebral annuli for smaller sharks but appeared overestimated for larger sharks. Von Bertalanffy growth parameters derived from vertebral length-at-age data are L∞ = 282 cm FL, K = 0.18, and t0 = –1.35 for males, and L∞ = 310 cm FL, K = 0.13, and t0 = −1.77 for females. The species grows faster and has a shorter life span than previously reported for these waters.

Movement of American lobster (Homarus americanus) in the southwestern Gulf of St. Lawrence

A total of 42,445 American lobsters (Homarus americanus) were tagged in thirty-one sites throughout the southwestern Gulf of St. Lawrence between 1980 and 1997. Results from the recapture of 8503 tagged lobsters showed small distances traveled between the release and the recapture position for animals ranging in size from 51 to 152 mm carapace length. The average distance traveled ranged from 2 km in parts of Baie des Chaleurs and western Cape Breton to 19 km in central Northumberland Strait. Lobsters moved generally along the shore (93% of the dispersion was in areas between the shore and the 20-m bathymetric contour). As a result, lobsters traveled longer distances in sites characterized by a gradually sloping bottom where the distance between the shore and the 20-m contour line was extensive in contrast to areas characterized by rapidly changing depths and by a relatively small amount of habitat shallower than 20 m. In the majority of sites (14 of 19) there was no significant difference between males and females in the average distance they traveled. In four of the five sites females moved farther than males. In general, the average distance traveled by berried females was shorter than that traveled by males or nonberried females. No relationship was observed between the distance traveled and the size of the animal. There was no strong evidence of a relationship between the average distance traveled and the number of days at liberty. In general, lobsters in the southwestern Gulf of St. Lawrence traveled short distances and dispersion was restricted to the nearshore habitat. Further, the distance traveled was not correlated to size, sex, or years at large. These findings show that there is little interaction between American lobsters from different fishing areas at the benthic level and that American lobster movements should have minimal consequences for management of the species in the southwestern Gulf of St. Lawrence.

Recruitment season, size, and age of young American eels (Anguilla rostrata) entering an estuary near Beaufort, North Carolina

Net catches from 1985–86 to 1994–95 at Pivers Island, North Carolina, indicated that glass-eel stage American eels (Anguilla rostrata) were recruited to the estuary from November to early May, with peak numbers in January, February, and March. There was no declining trend in recruitment over the years of sampling. Except for one year, there was no clear seasonal decrease in mean length. But shorter glass eels were older than longer glass eels, as judged by age within the glass eel growth zone of the otolith, suggesting that smaller fish took longer to arrive. The mean age of glass eels collected from the lower estuary and a freshwater site 9.5 km upriver differed by 8.4 d (36.2 vs. 44.6, respectively). Outer increments (30–35) of the otolith growth zone of glass eels from North Carolina were significantly wider than corresponding increments of otoliths from New Brunswick. Mean total ages of North Carolina, New Jersey, and New Brunswick elvers were 175.4, 201.2, and 209.3 d, corresponding to mean lengths of 55.9, 60.9, and 58.1 mm TL, respectively. The mean durations of glass-eel growth zones (44.6, 62.3, and 69.8) were in close agreement with those from previous studies, but total ages were not. This suggested that perhaps some finer (leptocephalus stage) increments were not detected by light microscopy, differences occurred in seasonal increment deposition, or absorption of the otolith material may have taken place during metamorphosis, rendering the aging of larvae inaccurate. Judging from the long recruitment period and seasonal uniformity in both mean age and length found in our study, the spawning period of American eels may be somewhat more protracted than previously considered.

Inter-regional correlation in tree-growth variations in the western United States, 1513-1964, at ENSO, sunspot, and longer frequency bands

Cross-spectral analysis of regional tree-ring data suggests the spatial pattern of correlation between moisture variations in the Sierra Nevada of central California and in other parts of the western United States is frequency dependent. Short wavelengths (2.8 to 10.7 years), perhaps associated with El Niño/Southern Oscillation, are strongly coherent both to the north (Oregon) and to the south (Southern California). Longer wavelengths (45 to 75 years) are strongly coherent only to the north. Frequency bands corresponding to annual sunspot series were associated with relatively weak patterns of spatial correlation.