Abnormal development of motor neurons in perfluorooctane sulphonate exposed zebrafish embryos

Perfluorooctane sulfonate (PFOS) is an environmental organic pollutant, the potential neurotoxicity of which is causing great concern in fish. In the present study, we examined the effects of PFOS on motor neurons, and investigated the potential toxicological mechanisms oxidative stress in zebrafish embryos. Six-hour post-fertilization (hpf) zebrafish embryos were exposed to 1.0 mg/L PFOS, then we examined the expression of alpha-tubulin, proliferating cell nuclear antigen (PCNA), cyclin-dependent kinase 5 (CDK5), and peroxiredoxin 2 (PRX2) after PFOS exposure until 120 hpf. The results showed that PFOS increased alpha-tubulin in the coccygeal spinal cord (CSC) at 96 hpf, whereas decreased alpha-tubulin in the brain and spinal cord at 120 hpf. PCNA expression was highly increased in CSC and abdomen compared with control at 96 and 120 hpf after PFOS exposure. In addition, PFOS exposure caused CDK5 expression to be highly increased in brain region following by down-regulation of PRX2 expression at 96 hpf. These results indicated that, at least in part, the effect on motor neurons induced by PFOS was mediated by dynamically interfering with the expression of alpha-tubulin and PCNA. Furthermore, PFOS-induced toxicity was associated with oxidative stress by deregulating CDK5 and PRX2.     

Apomorphine-induced motor behavior in rats exposed prenatally to alcohol

We psychopharmacologically examined dopamine function in rats exposed to ethanol prenatally. Pregnant rats received liquid diets of 35% or 0% ethanol-derived calories (EDC), or ad lib lab chow (LC). Twenty-eight-day-old offspring received systemic doses of apomorphine chosen to stimulate predominantly presynaptic (0.02 or 0.1 mg/kg) or postsynaptic dopamine receptors (2.0 or 5.0 mg/kg). Behavior was scored automatically for 60 min in an "open field." For males, prenatal ethanol exposure resulted in a dose-response shift to the left for locomotor activity. Females exposed to the liquid diet, with or without ethanol, showed less of an increase in locomotor activity following the 5.0 mg/kg dose of apomorphine than did LC controls. There were no effects of prenatal treatment on repetitious motor behavior in the automated "open field" or on stereotypy scored by direct observation in separate groups of rats. The results are consistent with an hypothesis that prenatal ethanol exposure alters the sensitivity of postsynaptic (perhaps mesolimbic) dopamine systems important to locomotor activity in young male rats.     

Cleavage and development in cultured preimplantation mouse embryos exposed to lidocaine

Two-cell preimplantation mouse embryos were exposed in vitro to lidocaine (0 to 1,000 μg/mL) for 72 h to determine the effects of this anesthetic on subsequent cleavage and development during prolonged exposures. Embryonic development was monitored each 24 h for 3 d. Lidocaine adversely affected the in vitro development of the mouse embryos, altering the distribution of the development stages at the evaluated culture tunes. The percentage of two-cell embryos that cleaved and developed to more advanced stages was decreased by the exposure to lidocaine. After 24 h of culture, two-cell embryos were arrested before completion of cellular division; this occured in 30% of the embryos at concentrations of 10 to 100 μg/mL and in 73.2% of the embryos with 1,000 μg/mL. After 48 h the blastomeres of the arrested embryos began to degenerate, showing lysis or fragmentation. At the lowest concentration, 14.9% of the arrested embryos exhibited the capacity to recover. These embryos continued their cleavage and normal development towards blastocyst formation. The cytotoxic effect and arrest at the two-cell stage were observed in a dose-dependent manner after 72 h of culture. We conclude that sensitivity to lidocaine embryotoxicity occurs during a window at the two-cell stage.     

Predictors of motor development in children prenatally exposed to cocaine

The current study examined the pattern of motor development across the first 18 months of life in infants with in utero exposure to cocaine to determine how prenatal drug effects and level of exposure relates to motor development. Motor development was examined at 1, 4, 12, and 18 months of age (corrected for prematurity). Infants were divided into cocaine exposed (n=392) and comparison (n=776) groups. Exposure status was determined by meconium assay and maternal self-report with alcohol, marijuana, tobacco, and opiates present in both groups. Motor skills were assessed at 1 month using the NICU Network Neurobehavioral Scale (NNNS), at 4 months using the posture and fine motor assessment of infants (PFMAI), at 12 months using the Bayley Scales of Infant Development-Second Edition (BSID-II), and at 18 months using the Peabody Developmental Motor Scales (PDMS). Examiners masked to exposure status performed all assessments. Motor scores were converted to standard (z) scores, and hierarchical linear modeling (HLM) was used to examine the change in motor skills from 1 to 18 months of age. Infants with exposure to cocaine showed low motor skills at their initial status of 1 month but displayed significant increases over time. Both higher and lower levels of tobacco use related to poorer motor performance on average. Heavy cocaine use related to poorer motor performance as compared to no use, but there were no effects of level of cocaine use on change in motor skills.     

Two types of abnormal automaticity in canine ventricular muscle fibers

Summary When ventricular muscle fibers from dog hearts were exposed to different [Ca²⁺]₀ in K⁺-free solutions, two types of abnormal automaticity were observed. In the K⁺-free, Ca²⁺-free, solution, 9 out of 55 preparations developed spontaneous activity at a reduced membrane potential. The maximum diastolic potential was –47.4±7.68 mV (n=9) with slow (less than 20 V/s) upstroke velocity of action potentials. The automaticity was based on membrane oscillations with increasing amplitude to become full-sized responses. It was suppressed after reduction of [Na⁺]₀, by an increase in [Ca²⁺]₀ above 3.6 mmol/l and by application of verapamil. In the K⁺-free, high-Ca²⁺, solutions (Ca²⁺ =3.6–7.2 mmol/l), oscillatory afterpotentials (OAPs) were observed following the driven action potentials at normal membrane potentials in all 55 preparations. Once the OAPs became large enough to attain threshold, there was the appearance of triggered-automaticity. The automaticity was induced by applying premature stimuli or manipulating [Ca²⁺]₀. It was abolished by application of either verapamil or tetrodotoxin by decreasing the amplitude of the OAPs or the fast Na⁺ current, respectively, or both in combination. These results indicate that the ventricular muscle fibers can develop abnormal automaticity of two different mechanisms in some conditions and can be a focus of ectopic impulse formation.This work was supported in part by the Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan     

The motivation of alcohol dependents to undergo treatment.

The purpose of this study was to investigate the determinants and factors influencing the motivation of alcohol dependents to undergo treatment. As well as exploring the influence of socio-demographic and illness-related variables, the study concentrated in particular on the influence of the clients' attitudes towards their current life situation, their alcohol problem and the treatment, also the influence of specific interventions aimed at increasing their motivation for treatment. On the basis of the results obtained in the study from 239 alcohol-dependent clients, it is shown to what extent these variables are of value as a means of predicting and modifying the motivation for treatment. In the motivation programme a variety of procedures was applied using a 2 x 2 design: motivation on an individual basis and motivation on a group basis were combined with two different approaches, one client-centred and one in accordance with the principles of cognitive-behavioural therapy. A group placed on a waiting list for 3 weeks was used as a control group. The clients were allocated to the various groups on a random basis. Specific attitudes on the part of the clients and aspects of the motivation programme provided significant indicators of the clients' likelihood to undergo treatment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Gene expression analysis on the early development of pig embryos exposed to malathion

Malathion is a widely used pesticide and there is evidence that it could alter mammal's germ and somatic cells, as well as cell lines. There are not enough studies showing how the nonacute malathion doses affect gene expression. This study analyzes gene expression alterations in pig morular embryos exposed in vitro, for 96 h, to several malathion concentrations after in vitro fertilization. cDNA libraries of isolated morular embryos were created and differential screenings performed to identify target genes. Seven clones were certainly identified. Genes related to mitochondrial metabolism as cytochrome c subunits I and III, nuclear genes such as major histocompatibility complex I (MHC I), and a hypothetical protein related with a splicing factor were the target of malathion's deregulation effect. The widespread use of malathion as a pesticide should be regarded with reproductive implications and more detailed analysis would yield more about molecular mechanisms of malathion injury on embryo cells.     

Quercetin and naringenin reduce abnormal development of mouse embryos produced by hydroxyurea

Objectives There is limited evidence about the impact of quercetin and naringenin on embryonic development. The purpose of this work was to evaluate in vitro their direct teratogenic potential as well as their protective activity against teratogenesis mediated by oxidative damage on mouse embryos. Methods Quercetin and naringenin toxicity on whole mouse cultured embryos, as well as their ability to protect embryos against hydroxyurea‐induced insult were evaluated. Key findings Quercetin 100 µm and naringenin 300 µm produced significant reduction of developmental and growth parameters, in comparison with those of the control group. Embryos exposed to the concurrent administration of quercetin or naringenin with hydroxyurea (2 µm , 2 h) were significantly protected from growth and developmental retardation, and abnormalities induced by hydroxyurea. Interestingly, embryos exposed to hydroxyurea and dimethyl sulfoxide 0.1%, the vehicle employed to dissolve flavonoids, also showed significant damage amelioration. Conclusions These results indicate that quercetin and naringenin have not only a minor toxic effect on development, but also a protective effect against hydroxyurea‐induced embryonic damage.     

Zebrafish embryos as models for embryotoxic and teratological effects of chemicals

The experimental virtues of the zebrafish embryo such as small size, development outside of the mother, cheap maintenance of the adult made the zebrafish an excellent model for phenotypic genetic and more recently also chemical screens. The availability of a genome sequence and several thousand mutants and transgenic lines together with gene arrays and a broad spectrum of techniques to manipulate gene functions add further to the experimental strength of this model. Pioneering studies suggest that chemicals can have in many cases very similar toxicological and teratological effects in zebrafish embryos and humans. In certain areas such as cardiotoxicity, the zebrafish appears to outplay the traditional rodent models of toxicity testing. Several pilot projects used zebrafish embryos to identify new chemical entities with specific biological functions. In combination with the establishment of transgenic sensor lines and the further development of existing and new automated imaging systems, the zebrafish embryos could therefore be used as cost-effective and ethically acceptable animal models for drug screening as well as toxicity testing.     

Nose-poking and head-dipping behaviors in rats prenatally exposed to alcohol

In three experiments pregnant female rats consumed liquid diets containing various amounts of the total calories in the form of ethanol. In the first study, offspring of these females were tested in a nose-poking paradigm. The frequency of this response was found to be a direct function of the level of alcohol exposure in utero. In a second study when nose poking produced the onset of a dim light, animals prenatally exposed to alcohol were again found to poke more often, and this effect was not attenuated by preweanling handling. Finally, the generality of these findings became evident when head dipping rather than nose poking was examined; head-dip frequency was higher in alcohol-exposed offspring, and this effect was independent of stimulus complexity. Additionally, offspring body weights and survival rates following this prenatal alcohol exposure are presented.     

Sensorimotor maturation and alcohol responsiveness in rats prenatally exposed to alcohol during gestational day 8

On gestational day 8, pregnant Sprague-Dawley derived rats received two intraperitoneal injections of 0.015 ml/g body weight of a 24% v/v ethanol solution representing an absolute alcohol dose of 2.82 g/kg per administration (ETOH Group). Control females were injected with similar volumes of saline (SAL Group) or did not receive any type of intraperitoneal administration of drugs (AC Group). In comparison with the control treatments, the ethanol treatment did not affect long-term maternal body weight gain during pregnancy, total length of gestation, probability of delivery, number of pups born per litter or the offsprings' preference of alcohol odor. Nevertheless, this ethanol insult was sufficient to significantly reduce body weights at birth and to retard the ontogeny of some sensorimotor patterns among offspring, as assessed through the Righting Reflex, Horizontal Screen Test and opening of the external auditory canals. The maturation of other reflexes (Cliff Aversion and Negative Geotaxis) appeared to be retarded indirectly as an outcome of the maternal injection procedure, with ETOH and SAL pups exhibiting similar developmental delays when compared with AC animals. During adulthood, rats prenatally exposed to alcohol exhibited a decreased hypothermic response to an intoxicating dose of alcohol as well as significant increases in voluntary alcohol consumption in comparison with both control conditions. Collectively, these results suggest that relatively acute alcohol exposure early in gestation may not only affect normal patterns of development but also later responsiveness to this pharmacological agent.     

Notochordal anomaly in frog embryos exposed to tetraethylthiuram monosulphide and tetraethylthiuram disulphide

Gastrulating embryos of the frog Microhyla ornata were treated with tetraethylthiuram monosulphide (TETM) and tetraethylthiuram disulphide (TETD) for 48-96 h. It was observed that both chemicals induce severe notochordal abnormalities at very low concentration. The notochord in abnormal embryos was highly swollen and entirely wavy. The notochordal cells were swollen and disarrayed. The embryos were also highly oedematous and there was considerable disarray of other tissues due to the abnormal notochord.     

Developmental toxicity and alteration of gene expression in zebrafish embryos exposed to PFOS

Perfluorooctanesulfonate (PFOS) is a persistent organic pollutant, the potential toxicity of which is causing great concern. In the present study, we employed zebrafish embryos to investigate the developmental toxicity of this compound. Four-hour post-fertilization (hpf) zebrafish embryos were exposed to 0.1, 0.5, 1, 3 and 5 mg/L PFOS. Hatching was delayed and hatching rates as well as larval survivorship were significantly reduced after the embryos were exposed to 1, 3 and 5 mg/L PFOS until 132 hpf. The fry displayed gross developmental malformations, including epiboly deformities, hypopigmentation, yolk sac edema, tail and heart malformations and spinal curvature upon exposure to PFOS concentrations of 1 mg/L or greater. Growth (body length) was significantly reduced in the 3 and 5 mg/L PFOS-treated groups. To test whether developmental malformation was mediated via apoptosis, flow cytometry analysis of DNA content, acridine orange staining and TUNEL assay was used. These techniques indicated that more apoptotic cells were present in the PFOS-treated embryos than in the control embryos. Certain genes related to cell apoptosis, p53 and Bax, were both significantly up-regulated upon exposure to all the concentrations tested. In addition, we investigated the effects of PFOS on marker genes related to early thyroid development (hhex and pax8) and genes regulating the balance of androgens and estrogens (cyp19a and cyp19b). For thyroid development, the expression of hhex was significantly up-regulated at all concentrations tested, whereas pax8 expression was significantly up-regulated only upon exposure to lower concentrations of PFOS (0.1, 0.5, 1 mg/L). The expression of cyp19a and of cyp19b was significantly down-regulated at all exposure concentrations. The overall results indicated that zebrafish embryos constitute a reliable model for testing the developmental toxicity of PFOS, and the gene expression patterns in the embryos were able to reveal some potential mechanisms of developmental toxicity.     

Lack of response inhibition in rats prenatally exposed to alcohol

Offspring of mothers who consumed either 32, 19, 8, or 0% of their daily caloric intake in the form of ethanol during pregnancy were tested for passive avoidance. At 18 days of age, the number of trials to criterion and the within-group variability were direct functions of the amount of ethanol consumed by the mother during pregnancy. At 41–53 days of age, alcohol-treated pups still required more trials to criterion than controls and had faster speeds into the shock compartment on the first trial. When the progeny of mothers consuming either 35, 17, or 0% ethanolderived calories during pregnancy were compared for conditioned taste aversion to a lithium chloride solution, a linear dose-response function was again evident. Animals in the alcohol-treated groups showed less suppression of drinking than controls. These investigations indicated that the effects of alcohol exposure in utero were manifested in behavioral out-comes involving response inhibition that were not correlated with the more familiar physical symptoms.     

Cell viability and proteomic analysis in cultured neurons exposed to methylmercury

Methylmercury is an environmental contaminant with special selectivity for cerebellar granule cells. The aim of this study was to determine the effect of long-term methylmercury exposure on cell viability and cellular proteome in cultured cerebellar granule cells. Primary cultures of mice cerebellar granule cells were treated with 0-300 nM methylmercury at 2 days in vitro (div) and afterwards the cells were harvested at 12 div. 100 nM methylmercury produced loss of cell viability, reduced intracellular glutamate content and increased lipid peroxidation. Glutamate transport was not modified by methylmercury treatment. Cell death induced by 300 nM methylmercury at 8 div was apoptotic without producing activation of caspase 3. Extracts of total protein were separated by 2D electrophoresis. Around 800 protein spots were visualized by silver staining in SDS-polyacrylamide gels. Gel images were digitized and protein patterns were analysed by image analysis. Several spots were identified through a combination of peptide mass fingerprinting and matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS). The mitochondrial protein 3-ketoacid-coenzyme A transferase I was decreased up to 39% of controls at concentrations of methylmercury that did not produce cytotoxic effects, whereas the cytoplasmic proteins lactate dehydrogenase chain B and actin did not change.     

Cognitive processes and motor skills differ in sensitivity to alcohol impairment

OBJECTIVE: Research reviews have suggested that cognitive and motor skills are not equally sensitive to a moderate dose of alcohol; they disagree, however, on which type of task is more sensitive to impairment. This issue is addressed in two experiments testing a cognitive and a motor skill task performed by the same person at comparable blood alcohol concentrations (BACs) during a moderate dose of alcohol. METHOD: A motor skill task (PR) and a rapid information processing (RIP) task requiring no learned motor skill were performed in counterbalanced order, and tests on the pair of tasks occurred at intervals as BAC rose and declined. In the first experiment, two groups of male social drinkers (each n = 10) received either a moderate dose of alcohol (0.62 g/kg) or placebo and performed the tasks with no consequence for performance. The second experiment comprised four groups (each n = 14) to verify and extend the results to a situation in which unimpaired performance under alcohol was rewarded on one or both tasks. RESULTS: In both experiments, impairment on the PR task tended to increase and diminish with rising and declining BACs, whereas impairment on the RIP task showed no such pattern. Reinforcement reduced the degree of impairment displayed on each task, but the different task profiles of impairment were still evident. CONCLUSIONS: The results indicate how disagreement over the sensitivity of cognitive and motor skills to a moderate dose of alcohol may occur when impairment is only assessed at some particular BACs. Theoretical and practical implications of the findings are discussed.