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1.
Aquat Toxicol ; 234: 105796, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33713916

RESUMO

This study leveraged the Japanese medaka fish embryo model for the assessment of effects of select contaminants on early development in fish. Fish embryos were exposed to various pharmaceutical contaminants including synthetic hormones and non-steroidal anti-inflammatory drugs and their effects on development were observed. Initial screening determined that swim bladder inflation failure was the most common endpoint detected. Swim bladder inflation failure was first explored in a study demonstrating that medaka require access to the air-water interphase to inflate their swim bladders in a time-dependent manner, and swim bladder inflation failure was correlated with mortality. Fish embryos were exposed 24-hours post fertilization until hatch to concentration ranges of various pharmaceutical contaminants including: 17ß-estradiol, 17α-ethinylestradiol, and levonorgestrel (1 to 1000 µg/L), or diclofenac (0.32 to 100 mg/L). The main effect observed across all four compounds was a significant increase in failure of swim bladder inflation with increasing exposure concentration (24 to 72-hours post-hatch). Following single compound experiments combinatorial exposures using no-observed-effect concentrations were conducted. The main effect observed was a significant decrease in inflation success 24-hours post-hatch following a binary mixture of levonorgestrel and 17α-ethinylestradiol, as well as a significant decrease in swim bladder inflation success at all times following exposure to a quaternary mixture of all four compounds. This study demonstrated that embryonic exposure to pharmaceutical compounds, both alone and in combination, resulted in failure of swim bladder inflation in larval Japanese medaka.


Assuntos
Anti-Inflamatórios não Esteroides/toxicidade , Oryzias/crescimento & desenvolvimento , Bexiga Urinária/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Animais , Diclofenaco/toxicidade , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/fisiologia , Estradiol/toxicidade , Oryzias/fisiologia , Bexiga Urinária/fisiologia
2.
Methods Mol Biol ; 2218: 195-208, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33606233

RESUMO

For organisms to function normally, biological molecules must work at the correct time in the right cells and within the right intracellular compartments of cells. Biological research relies heavily on discovering the cellular locations at which such molecular interactions occur. A mainstay technique in this process of discovery is the visualization of locations of proteins in cells and tissues, known as immunocytochemistry and immunohistochemistry, respectively. If performed correctly, these techniques can provide detailed information regarding the endogenous locations of proteins and their ectopic locations or absence in mutants and in disease states.


Assuntos
Embrião não Mamífero/metabolismo , Embrião não Mamífero/fisiologia , Imuno-Histoquímica/métodos , Peixe-Zebra/metabolismo , Peixe-Zebra/fisiologia , Animais , Feminino , Masculino , Proteínas de Peixe-Zebra/metabolismo
3.
Methods Mol Biol ; 2218: 185-194, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33606232

RESUMO

Here, we describe a fast and straightforward methodology to in vivo detect transcriptional activity in the early zebrafish germ line. We report how fluorescently labeled morpholinos, targeted to nascent early transcripts, can be used to track the onset of transcriptional events during early embryogenesis. This method could be applied to any tagged cell line in a developing early zebrafish embryo as long as the gene of interest is expressed at high enough level for morpholino detection and is expressed at the first and main wave of genome activation, for which the protocol has been verified. The protocol, in combination with genetic manipulation, allows studies of mechanisms driving zygotic genome activation (ZGA) in individual cells. The reported procedures apply to a broad range of purposes for zebrafish embryo manipulation in view of imaging nuclear molecules in specific cell types.


Assuntos
Células Germinativas/fisiologia , Transcrição Genética/fisiologia , Peixe-Zebra/fisiologia , Animais , Embrião não Mamífero/metabolismo , Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário/genética , Desenvolvimento Embrionário/fisiologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento/genética , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Genoma/genética , Genoma/fisiologia , Células Germinativas/metabolismo , Masculino , Morfolinos/metabolismo , Transcrição Genética/genética , Peixe-Zebra/genética , Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Zigoto/metabolismo , Zigoto/fisiologia
4.
Methods Mol Biol ; 2218: 265-276, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33606238

RESUMO

Transgenic zebrafish in which the germline is specifically labeled with enhanced green fluorescent protein (eGFP) can be used for continuous observation of germline development during the lifetime, from the primordial germ cells (PGCs) in the early embryo to the gametes in the mature gonad. In this chapter, we describe a procedure for the generation of transgenic fish Tg(piwil1:egfp-UTRnanos3), the sample preparation for live imaging of PGCs, for high-resolution imaging of germ cells in developing gonads, and quantifying PGC numbers. The methods described in this chapter are not only applicable to the study of germ cells, but also provide general advices for researchers who are willing to generate transgenic zebrafish and do observation on live embryos as well as on fixed tissues.


Assuntos
Animais Geneticamente Modificados/fisiologia , Células Germinativas/fisiologia , Animais , Embrião não Mamífero/fisiologia , Feminino , Gônadas/fisiologia , Masculino , Peixe-Zebra/fisiologia
5.
Methods Mol Biol ; 2218: 347-354, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33606244

RESUMO

Many proteins are assumed to mediate post-transcriptional regulation of mRNAs. However, the lack of information about their target mRNAs and functional domains hampers the detailed analysis of their molecular function. Here we describe a method to analyze the post-transcriptional effects of proteins of interest by artificially tethering the protein to a reporter mRNA in zebrafish embryos.


Assuntos
Bioensaio/métodos , Embrião não Mamífero/fisiologia , Proteínas de Ligação a RNA/genética , RNA/genética , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/genética , Animais , Processamento Pós-Transcricional do RNA/genética , RNA Mensageiro/genética
6.
Methods Mol Biol ; 2218: 367-380, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33606246

RESUMO

The study of translational regulation requires reliable measurement of both mRNA levels and protein synthesis. Cytoplasmic polyadenylation is a prevalent mode of translational regulation during oogenesis and early embryogenesis. Here the length of the poly(A) tail of an mRNA is coupled to its translatability. We describe a protocol to identify translationally regulated genes and measure their translation rate in the early zebrafish embryo using genome-wide polysome profiling. This protocol relies on the isolation of mRNA by means of an rRNA depletion strategy, which avoids capture bias due to short poly(A) tail that can occur when using conventional oligo(dT)-based methods. We also present a simple PCR-based method to measure the poly(A) tail length of selected mRNAs.


Assuntos
Biossíntese de Proteínas/genética , Peixe-Zebra/genética , Animais , Citoplasma/genética , Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário/genética , Oócitos/fisiologia , Oogênese/genética , Poli A/genética , Poliadenilação/genética , RNA Mensageiro Estocado
7.
Ecotoxicol Environ Saf ; 208: 111700, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-33396031

RESUMO

Sertraline (SER) is one of the most frequently detected antidepressant drugs in aquatic environments. However, knowledge regarding SER-induced behavioral alterations in fish is insufficient, as well as the mechanisms underlying SER-induced toxicity. The present study aimed to determine behavioral and molecular responses in larval fish following SER exposure with a focus on its mode of action. Zebrafish embryos (~6 h-post-fertilization, hpf) were exposed to one of three concentrations of SER (1, 10, 100 µg/L) for 6 days, respectively. Evaluated parameters included development, behavior, transcripts related to serotonin signaling, serotonin levels, and acetylcholinesterase activity. Accelerated hatching of zebrafish embryos was observed for those fish exposed to 100 µg/L SER at 54 hpf. Locomotor activity (e.g. distance moved and mobile cumulative duration) was significantly reduced in larval zebrafish following exposure to 10 and 100 µg/L SER. Conversely, larval fish showed increased dark-avoidance after exposure to 1-100 µg/L SER. Of the measured transcripts related to serotonin signaling, only serotonin transporter (serta) and serotonin receptor 2c (5-ht2c) mRNA levels were increased in fish in response to 10 µg/L SER treatment. However, serotonin levels were unaltered in larvae exposed to SER. There were no differences among groups in acetylcholinesterase activity at any concentration tested. Taking together, the results evidenced that exposure to SER alters behavioral responses in early-staged zebrafish, which may be related to the abnormal expression of 5-ht2c. This study elucidates molecular responses to SER and characterizes targets that may be sensitive to antidepressant pharmaceuticals in larval fish.


Assuntos
Antidepressivos/toxicidade , Comportamento Animal/efeitos dos fármacos , Sertralina/toxicidade , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/fisiologia , Animais , Antidepressivos/análise , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/fisiologia , Locomoção/efeitos dos fármacos , Serotonina/metabolismo , Sertralina/análise , Transdução de Sinais/efeitos dos fármacos , Poluentes Químicos da Água/análise , Peixe-Zebra/crescimento & desenvolvimento , Peixe-Zebra/metabolismo
8.
Ecotoxicol Environ Saf ; 208: 111763, 2021 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-33396083

RESUMO

Sulfate occurs naturally in the aquatic environment but its elevated levels can be toxic to aquatic life in freshwater environments. We investigated the toxicity of sulfate in humic, soft freshwater to whitefish (Coregonus lavaretus) from fertilization of eggs to hatching i.e. during the critical phases of whitefish early development. Anadromous Kokemäenjoki whitefish eggs and sperm during fertilization, embryos and larvae were exposed in the long-term 175-day incubation to seven different sodium sulfate (Na2SO4) concentrations from 44 to 2 000 mg SO4 L-1. Endpoint variables were the fertilization success, offspring survival and larval growth. Egg fertilization and early embryonic development were the most sensitive developmental stages of whitefish to sulfate, although the fertilization success and survival of embryos decreased only in the highest concentration of 2 000 mg SO4 L-1. The survival during late embryonic period, hatching and the 5-day larval period was high and no difference between the control and sulfate treatments were observed. LC50-values of sulfate for early embryonic period and for the entire embryonic and larval period was 1 413 and 1 161 mg L-1, respectively. The NOEC (No-observed Effect Concentration) of sulfate for the both periods was 1 207 mg L-1. The tolerance of whitefish early stages to sulfate toxicity seems to be on the same level as the tolerance of other salmonids' early stages.


Assuntos
Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário/efeitos dos fármacos , Salmonidae/embriologia , Sulfatos/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Água Doce/química , Larva , Dose Letal Mediana , Masculino , Salmonidae/crescimento & desenvolvimento , Espermatozoides
9.
Ecotoxicol Environ Saf ; 211: 111920, 2021 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-33497861

RESUMO

Azoxystrobin is a broad-spectrum strobilurin fungicide for use on a wide range of crops available to end-users as formulated products. Due to its extensive application, it has been detected in aquatic ecosystems, raising concerns about its environmental impact, which is still poorly explored. The objective of this work was to study the effects of a commercial formulation of azoxystrobin in the zebrafish embryo model. Sublethal and lethal effects were monitored during the exposure period from 2 h post fertilisation (hpf) to 96 hpf after exposure to azoxystrobin concentrations (1, 10 and 100 µg L-1). The responses of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR)) as well as detoxifying enzymes (glutathione-s-transferase (GST) and carboxylesterase (CarE)) were evaluated at 96 hpf. Similarly, glutathione levels (reduced (GSH) and oxidised (GSSG) glutathione), neurotransmission (acetylcholinesterase (AChE)) and anaerobic respiration (lactate dehydrogenase (LDH)) -related enzymes were assayed. At 120 hpf, larvae from each group were used for behaviour analysis. Results from this study showed concentration-dependent teratogenic effects, particularly by increasing the number of malformations (yolk and eye), with a higher prevalence at the highest concentration. However, it was found that the lowest concentration induced a high generation of reactive oxygen species (ROS) and increased activity of SOD, GST, and CarE. In addition, GR and GSSG levels were decreased by the lowest concentration, suggesting an adaptive response to oxidative stress, which is also supported by the increased AChE activity and absence of behavioural changes. These findings advance the knowledge of the azoxystrobin developmental and environmental impacts, which may impose ecotoxicological risks to non-target species.


Assuntos
Embrião não Mamífero/efeitos dos fármacos , Fungicidas Industriais/toxicidade , Pirimidinas/toxicidade , Estrobilurinas/toxicidade , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/embriologia , Animais , Antioxidantes/farmacologia , Catalase/metabolismo , Ecossistema , Embrião não Mamífero/fisiologia , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Glutationa Transferase/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio , Estrobilurinas/farmacologia , Superóxido Dismutase/metabolismo , Peixe-Zebra/metabolismo
10.
Chemosphere ; 263: 127985, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32854011

RESUMO

Although banished in some countries, triclosan (TCS) and triclocarban (TCC) have been detected in surface waters in concentrations ranging from ng L-1 to µg L-1 and have shown to affect non-target organisms posing risk to aquatic ecosystems. However, the majority of the studies consider higher levels of these chemicals and single exposure effects to investigate their potential risks, rather than using environmentally relevant concentrations and their binary mixture. In this study, the toxicity of TCS and TCC, and their binary mixture was assessed in catfish embryos (Rhamdia quelen, a south American native species) exposed to environmental concentrations during 96 h. Organisms were evaluated through the endpoints of developmental abnormalities (spine, fin, facial/cranial and thorax), biochemical biomarkers related to oxidative stress responses: catalase (CAT), superoxide dismutase (SOD), glutathione-S-transferase (GST) activities, protein carbonylation (PCO) and neurotoxicity by acetylcholinesterase activity (AChE). The data showed that TCS caused fin abnormalities, decrease of SOD activity and increase of AChE activity in the catfish embryos of 96hpf. On the other hand, TCC and the binary mixture showed a higher abnormality index for the 96hpf embryos, and an induction of CAT and GST activities for the mixture treatment. The results obtained were able to show potential, but not severe, toxicity of TCS and TCC even in low concentrations and a short period of exposure. The relevance of studies approaching real scenarios of exposure should be reinforced, considering environmental concentrations of chemicals, interactions of contaminants in complex mixtures and the use of a native species such as R. quelen exposed during initial stages of development.


Assuntos
Carbanilidas/toxicidade , Embrião não Mamífero/fisiologia , Triclosan/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Biomarcadores/metabolismo , Catalase/metabolismo , Peixes-Gato/embriologia , Peixes-Gato/metabolismo , Ecossistema , Estresse Oxidativo/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Testes de Toxicidade Subaguda
11.
Chemosphere ; 261: 127753, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32745739

RESUMO

Selective serotonin reuptake inhibitors (SSRIs) have been shown to interfere with various physiological functions of aquatic organisms, yet the neuroactive potential of low concentrations of SSRIs in the aquatic environment is unclear. The current study investigated the effects of fluoxetine and citalopram on the visual motor response (VMR) of 107 h old zebrafish (Danio rerio) embryos. Results document a reduction in stress-related swimming activity of zebrafish embryos at environmentally relevant concentration levels, with fluoxetine being more effective than citalopram. Further experiments were designed to elucidate (1) if the lower neuroactive potential of citalopram is due to differences in uptake kinetics, (2) if the metabolite of fluoxetine, norfluoxetine, contributes to the neuroactive potential of fluoxetine, (3) and how SSRIs and their metabolites interact in equimolar mixtures. At the stage of 120 h, zebrafish embryos accumulate citalopram at significantly lower rates (up to 127 times) than fluoxetine. Moreover, it was demonstrated that norfluoxetine reduces the embryonic VMR similarly to fluoxetine resulting in additive effects of these substances on stress-related behavior in zebrafish embryos. In contrast, the interaction of fluoxetine, norfluoxetine and citalopram varied with test concentrations of the equimolar mixtures. Findings provide evidence that environmentally relevant concentrations of fluoxetine reduce stress-related behavior of zebrafish embryos, while these effects may be enhanced by the interaction of multiple SSRIs and their metabolites in environmental exposure scenarios.


Assuntos
Comportamento Animal/efeitos dos fármacos , Embrião não Mamífero/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Inibidores de Captação de Serotonina/toxicidade , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/fisiologia , Animais , Bioacumulação/efeitos dos fármacos , Citalopram/metabolismo , Citalopram/toxicidade , Embrião não Mamífero/metabolismo , Embrião não Mamífero/fisiologia , Exposição Ambiental , Fluoxetina/metabolismo , Fluoxetina/toxicidade , Inibidores de Captação de Serotonina/metabolismo , Natação , Poluentes Químicos da Água/metabolismo
12.
Sci Rep ; 10(1): 8521, 2020 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-32444613

RESUMO

LSD1/KDM1A is a widely conserved lysine-specific demethylase that removes methyl groups from methylated proteins, mainly histone H3. We previously isolated the zebrafish LSD1 gene and demonstrated that it is required for primitive hematopoiesis. Recently, a neuron-specific splicing variant of LSD1 was found in mammals and its specific functions and substrate specificities were reported. To our surprise, zebrafish LSD1 cDNA, which we previously analyzed, was corresponded to the neuron-specific variant in mammals. In this study, we investigated the structures and expression of LSD1 splicing variants in zebrafish and found all 4 types of LSD1 isoforms: LSD1, LSD1+2al, LSD1+8al and LSD1+2al8al. Interestingly, LSD1+8al/LSD1+2al8al, which correspond to mammalian neuron-specific variants, expressed ubiquitously in zebrafish. We also performed phenotypic rescue experiments of a zebrafish LSD1 mutant (kdm1ait627) using human and zebrafish LSD1 variants to identify which variant is involved in primitive hematopoiesis. Unexpectedly, the overexpression of all types of human and zebrafish variants was able to rescue the hematopoietic phenotypes in LSD1 mutants. Furthermore, enzymatic-deficient LSD1K661A (human) and K638A (zebrafish) were also able to rescue the mutant phenotypes. These results suggest that the LSD1 functions in zebrafish primitive hematopoiesis are free from any splicing-dependent regulation or demethylation reaction.


Assuntos
Embrião não Mamífero/fisiologia , Hematopoese , Histona Desmetilases/metabolismo , Lisina/genética , Splicing de RNA , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Sequência de Aminoácidos , Animais , Células Cultivadas , Embrião não Mamífero/citologia , Histona Desmetilases/genética , Humanos , Lisina/metabolismo , Metilação , Mutação , Isoformas de Proteínas , Homologia de Sequência , Peixe-Zebra/embriologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética
13.
Chemosphere ; 253: 126762, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32302915

RESUMO

17ß-trenbolone (17ß-TBOH) is one of the dominant metabolites of trenbolone acetate, which is widely applied in beef cattle operations around the globe. The effects of environmental concentrations of 17ß-trenbolone on the early development of zebrafish embryos have received very little attention. Melatonin could regulate sleep-wake cycle and plays a protective role in various adverse conditions. Here, environmentally realistic concentrations of 17ß-trenbolone (1 ng/L, 10 ng/L, 50 ng/L) has been exposure to zebrafish embryos at 2 h postfertilization (hpf). The results showed that 10 ng/L and 50 ng/L 17ß-trenbolone disturbed the distribution of caudal primary motoneurons and downregulated expression of motoneuron development related genes along with locomotion decreasing. While melatonin could recover the detrimental effects caused by 17ß-trenbolone. Interestingly, 17ß-trenbolone exposure increased waking activity and decreased rest even in a low dose (1 ng/L). Moreover, it upregulated hypocretin/orexin (Hcrt) signaling which promotes wakefulness. Melatonin restored the insomnia-like alternation induced by 17ß-trenbolone exposure. Collectively, we conclude that 17ß-trenbolone disturbed motoneuron development and altered sleep/wake behavior, while melatonin could alleviate the deleterious influence on motoneuron development and recover the circadian rhythm.


Assuntos
Comportamento Animal/efeitos dos fármacos , Poluentes Ambientais/toxicidade , Melatonina/farmacologia , Atividade Motora/efeitos dos fármacos , Distúrbios do Início e da Manutenção do Sono/prevenção & controle , Acetato de Trembolona/toxicidade , Peixe-Zebra , Animais , Bovinos , Ritmo Circadiano/efeitos dos fármacos , Ritmo Circadiano/genética , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Neurônios Motores/efeitos dos fármacos , Orexinas/genética , Fenótipo , Distúrbios do Início e da Manutenção do Sono/induzido quimicamente
14.
Integr Zool ; 15(4): 338-348, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32297704

RESUMO

Low-elevation species can migrate toward higher elevations to survive in a warming world. However, animals' responses to hypoxia when migrating to high elevations have rarely been addressed. To identify the response of low-elevation lizards to high-elevation hypoxia, we collected field body temperatures (Tfb ) and operative temperatures (Te ) of lizards (Eremias argus) from a low-elevation population (1036 m) and a high-elevation population (2036 m), and then determined adult thermal physiology, embryonic development, and hatchling phenotypes after acclimating low-elevation lizards and incubating their eggs in conditions mimicking the low-elevation oxygen condition (18.5% O2 ) and high-elevation oxygen (hypoxic) condition (16.5% O2 ). Our study revealed that Tfb and Te were higher for the low-elevation population compared to the high-elevation population. We also found adults from low elevation acclimated to hypoxia preferred lower body temperatures, but did not show changes in locomotor performance or growth. In addition, hypoxia did not affect embryonic development (hatching time and success) or hatchling phenotypes (body size and locomotor performance). These results suggest that adult lizards from low elevations can respond to hypoxia-induced stress when migrating to high elevations by behaviorally thermoregulating to lower body temperatures in order to sustain normal functions. Similarly, low-elevation embryos can develop normally (with unchanged hatching success and offspring phenotypes) under the high-elevation hypoxic condition. This study highlights that low-elevation populations of a species that inhabits a range of elevations can buffer the impact of high-elevation hypoxic conditions to some degree and thus attain similar fitness to the source population.


Assuntos
Regulação da Temperatura Corporal , Desenvolvimento Embrionário/fisiologia , Lagartos/fisiologia , Oxigênio/metabolismo , Fenótipo , Aclimatação , Altitude , Anaerobiose , Animais , China , Embrião não Mamífero/fisiologia , Feminino , Lagartos/crescimento & desenvolvimento , Masculino
15.
J Therm Biol ; 88: 102528, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32126003

RESUMO

As global temperatures continue to rise, so too will the nest temperatures of many species of turtles. Yet for most turtle species, including the estuarine diamondback terrapin (Malaclemys terrapin), there is limited information on embryonic sensitivity to elevated temperature. We incubated eggs of M. terrapin at three, mean temperatures (31, 34, 37 °C) under two thermal exposure regimes (constant or semi-naturally fluctuating temperature) and measured hatching success, developmental rate, and hatchling size. Hatching success was 100% at 31 °C and 67% at 34 °C, respectively; at 37 °C, all eggs failed early in the incubation period. These values were unaffected by exposure regime. The modeled LT50 (temperature that was lethal to 50% of the test population) was 34.0 °C in the constant and 34.2 °C in the fluctuating thermal regime, reflecting a steep decline in survival between 33 and 35 °C. Hatchlings having been incubated at a constant 34 °C hatched sooner than those incubated at 31 °C under either constant or fluctuating temperature. Hatchlings were smaller in straight carapace length (CL) and width after having been incubated at 34 °C compared to 31 °C. Larger (CL) hatchlings resulted from fluctuating temperature conditions relative to constant temperature conditions, regardless of mean temperature. Based upon recent temperatures in natural nests, the M. terrapin population studied here appears to possess resiliency to several degrees of elevated mean nest temperatures, beyond which, embryonic mortality will likely sharply increase. When considered within the mosaic of challenges that Maryland's M. terrapin face as the climate warms, including ongoing habitat losses due to sea level rise and impending thermal impacts on bioenergetics and offspring sex ratios, a future increase in embryonic mortality could be a critical factor for a population already experiencing ecological and physiological challenges due to climate change.


Assuntos
Mudança Climática , Embrião não Mamífero/fisiologia , Temperatura , Tartarugas/fisiologia , Animais , Feminino , Zigoto
16.
Chemosphere ; 252: 126433, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32182507

RESUMO

Different studies have demonstrated effects of pesticides during embryo development in vertebrates and stage-dependent effects, but there is no information concerning this for Salvator merianae. We evaluated the effects of the herbicides Glyphosate Roundup (GLY-RU) and Glyphosate Panzer Gold (GLY-PZ); and the insecticides Chlorpyrifos (CPF) and Cypermethrin (CYP), and their complex mixtures, at different concentrations in hematological parameters of S. merianae embryos at two different development stages. The analyzed parameters were Total and Differential White Blood Cells Count, Heterophils/Lymphocytes index (H/L), Lobularity index, and Natural Antibodies (Nabs titres), as well as growth, embryo mortality and birth delay. Heterophils decreased in the intermediate concentrations tested of CYP and GLY-RU, in animals exposed at 33-days development. Lymphocytes increased in the intermediate concentration tested of GLY-RU, and the H/L index decreased in the maximum concentration tested of GLY-RU. NAbs titres increased in those animals exposed to the maximum CYP concentration tested. However, animals exposed at 3/5-days development showed no differences among treatments in most of the analyzed parameters, suggesting a stage-dependent response. Nevertheless, those animals exposed to GLY-PZ showed lower Nabs titres in relation to negative control. These results suggest effects on different hematological parameters related to the immune system of S. merianae, according to the used pesticide (herbicide or insecticide), its concentration and commercial formulation (GLY-RU or GLY-PZ), and the stages of development of the exposed animals. Our results reveal the importance of carrying out studies that evaluate the effects of permanent exposure of living beings and their environments to these toxics.


Assuntos
Lagartos/fisiologia , Praguicidas/toxicidade , Animais , Clorpirifos/toxicidade , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário , Glicina/análogos & derivados , Glicina/toxicidade , Herbicidas/toxicidade , Sistema Imunitário , Inseticidas/toxicidade , Lagartos/sangue , Lagartos/embriologia , Piretrinas/toxicidade
17.
J Fish Biol ; 97(1): 113-120, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32222964

RESUMO

A laboratory flume was constructed to examine substrate effects on aquatic development. The flume was designed as a once-through system with a submerged cobble-filled corebox. Lake whitefish (Coregonus clupeaformis) embryos and temperature probes were deployed at multiple sites within the cobble and in the open water channel. Embryos were incubated in the flume for two different experimental periods: one to examine substrate impacts during natural lake cooling (37 days: 5 December 2016 to 10 January 2017) and the second to investigate substrate effects while administering a twice weekly 1 h heat shock (51 days: 11 January to 2 March 2017). During incubation, no significant difference was found in the average temperature between locations; however, temperatures were more stable within the cobble. Following both incubation periods, embryos retrieved from the cobble were significantly smaller in both dry mass and body length by up to 20%. These results demonstrate differences between embryos submerged in a cobble substrate and in the open water column, highlighting the need to consider the physical influences from the incubation environment when assessing development effects as part of any scientific study or environmental assessment.


Assuntos
Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário/fisiologia , Salmonidae/embriologia , Animais , Meio Ambiente , Salmonidae/fisiologia , Temperatura
18.
Nat Commun ; 11(1): 1269, 2020 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-32152267

RESUMO

Multicellular rosettes are transient epithelial structures that serve as intermediates during diverse organ formation. We have identified a unique contributor to rosette formation in zebrafish Kupffer's vesicle (KV) that requires cell division, specifically the final stage of mitosis termed abscission. KV utilizes a rosette as a prerequisite before forming a lumen surrounded by ciliated epithelial cells. Our studies identify that KV-destined cells remain interconnected by cytokinetic bridges that position at the rosette's center. These bridges act as a landmark for directed Rab11 vesicle motility to deliver an essential cargo for lumen formation, CFTR (cystic fibrosis transmembrane conductance regulator). Here we report that premature bridge cleavage through laser ablation or inhibiting abscission using optogenetic clustering of Rab11 result in disrupted lumen formation. We present a model in which KV mitotic cells strategically place their cytokinetic bridges at the rosette center, where Rab11-associated vesicles transport CFTR to aid in lumen establishment.


Assuntos
Divisão Celular/fisiologia , Polaridade Celular/fisiologia , Embrião não Mamífero/fisiologia , Desenvolvimento Embrionário/fisiologia , Macrófagos do Fígado/fisiologia , Organogênese/fisiologia , Peixe-Zebra/embriologia , Animais , Linhagem Celular , Movimento Celular , Cílios/fisiologia , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Embrião não Mamífero/metabolismo , Desenvolvimento Embrionário/genética , Células Epiteliais/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Macrófagos do Fígado/citologia , Mitose , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
19.
Environ Toxicol Chem ; 39(4): 904-912, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32072671

RESUMO

Estrogen toxicity has been an area of priority in aquatic toxicology over the last 20 yr. Currently, estrogen toxicity is primarily linked to classical estrogen signaling, the interaction of estrogen receptors alpha and beta (ERα and ERß). Recent evidence has indicated that a rapid, nongenomic, nonclassical estrogen signaling pathway exists via the G protein-coupled estrogen receptor (GPER), which is expressed in many biological systems, with roles in the cardiovascular system. The objective of the present study was to investigate the effect of 17α-ethinylestradiol (EE2) on the heart rate of embryonic Japanese medaka (Oryzias latipes). A significant decrease (bradycardia) in embryonic heart rate was observed at all treatment concentrations (0.1, 1, 10, 100, and 1000 ng/L EE2) at 144, 168, and 192 h postfertilization (hpf; p ≤ 0.05), whereas 120 and 216 hpf embryos experienced a significant decrease from the control at 10, 100, and 1000 ng/L EE2 and 0.1, 100, and 1000 ng/L EE2, respectively (p ≤ 0.05). In addition, using select estrogen receptor modulators, it was demonstrated that estrogen-induced bradycardia appears to be linked to GPER and not ERα and ERß. The present study highlights GPER as a novel and alternative mode of action for EE2 toxicity at environmentally relevant concentrations. Environ Toxicol Chem 2020;39:904-912. © 2020 SETAC.


Assuntos
Embrião não Mamífero/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Etinilestradiol/toxicidade , Frequência Cardíaca/efeitos dos fármacos , Oryzias/fisiologia , Poluentes Químicos da Água/toxicidade , Animais , Relação Dose-Resposta a Droga , Embrião não Mamífero/fisiologia , Receptor alfa de Estrogênio/metabolismo , Receptor beta de Estrogênio/metabolismo , Estrogênios/metabolismo , Feminino , Masculino , Transdução de Sinais
20.
Chemosphere ; 249: 126148, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32062212

RESUMO

Cypermethrin is one of the widely used type-II pyrethroid and the indiscriminate use of this pesticide leads to life threatening effects and in particular showed developmental effects in sensitive populations such as children and pregnant woman. However, the molecular mechanisms underlying cypermethrin-induced development toxicity is not well defined. To address this gap, the present study was designed to investigate the phenotypic and transcriptomic (next generation RNA-Seq method) impact of cypermethrin in zebrafish embryos as a model system. Zebrafish embryos at two time points, 24 h postfertilization (hpf) and 48 hpf were exposed to cypermethrin at a concentration of 10 µg/L. Respective control groups were maintained. Cypermethrin induced both phenotypic and transcriptomic changes in zebrafish embryos at 48 hpf. The phenotypic anomalies such as delayed hatching rate, increased heartbeat rate and deformed axial spinal curvature in cypermethrin exposed zebrafish embryos at 48 hpf as compared to its respective controls. Transcriptomic analysis indicated that cypermethrin exposure altered genes associated with visual/eye development and gene functional profiling also revealed that cypermethrin stress over a period of 48 h disrupts phototransduction pathway in zebrafish embryos. Interestingly, cypermethrin exposure resulted in up regulation of only one gene, tnnt3b, fast muscle troponin isoform 3T in 24 hpf embryos as compared to its respective controls. The present model system, cypermethrin exposed zebrafish embryos elaborates the toxic consequences of cypermethrin exposure during developmental stages, especially in fishes. The present findings paves a way to understand the visual impairment in sensitive populations such as children exposed to cypermethrin during their embryonic period and further research is warranted.


Assuntos
Piretrinas/toxicidade , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/embriologia , Animais , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/fisiologia , Perfilação da Expressão Gênica , Larva/efeitos dos fármacos , Praguicidas/metabolismo , Transcriptoma , Peixe-Zebra/metabolismo
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