N6-Methyladenine in Eukaryotic DNA: Tissue Distribution, Early Embryo Development, and Neuronal Toxicity.

DNA methylation is one of the most important epigenetic modifications and is closely related with several biological processes such as regulation of gene transcription and the development of non-malignant diseases. The prevailing dogma states that DNA methylation in eukaryotes occurs essentially through 5-methylcytosine (5mC) but recently adenine methylation was also found to be present in eukaryotes. In mouse embryonic stem cells, 6-methyladenine (6mA) was associated with the repression and silencing of genes, particularly in the X-chromosome, known to play an important role in cell fate determination. Here, we have demonstrated that 6mA is a ubiquitous eukaryotic epigenetic modification that is put in place during epigenetically sensitive periods such as embryogenesis and fetal development. In somatic cells there are clear tissue specificity in 6mA levels, with the highest 6mA levels being observed in the brain. In zebrafish, during the first 120 h of embryo development, from a single pluripotent cell to an almost fully formed individual, 6mA levels steadily increase. An identical pattern was observed over embryonic days 7-21 in the mouse. Furthermore, exposure to a neurotoxic environmental pollutant during the same early life period may led to a decrease in the levels of this modification in female rats. The identification of the periods during which 6mA epigenetic marks are put in place increases our understanding of this mammalian epigenetic modification, and raises the possibility that it may be associated with developmental processes.

Short-term effects of a perinatal exposure to a 16 polycyclic aromatic hydrocarbon mixture in rats: assessment of early motor and sensorial development and cerebral cytochrome oxidase activity in pups.

Humans are exposed to polycyclic aromatic hydrocarbons (PAHs), a family of ubiquitous neurotoxic pollutants, mainly through ingestion of contaminated food. Developing organisms can be exposed also to PAHs due to the ability of these compounds to pass through the placental barrier as well as through the breast milk. Previous animal studies have reported that the exposure of rats to a 16 PAH mixture at environmental doses strictly limited to gestation did not induce any long-lasting consequences, whereas gestational and lactational PAH exposure induced long-term behavioral and cerebral metabolic effects. In the present study, short-term effects of exposures to the same PAH mixture during gestation, or during gestation and lactation, were assessed by evaluating motor and sensory development of rat pups, and by measuring cerebral cytochrome oxidase activity (a marker of energetic metabolism) in different brain areas. Brain levels of PAHs and some monohydroxylated metabolites were also evaluated in pups at birth and at 21 days of postnatal life. No significant short-term modifications of behavioral development and of cerebral metabolism were observed following an early PAH exposure whatever the dose and the period of exposure. Surprisingly, the same brain levels of concentration of PAHs and metabolites were observed in control and exposed pups in both studies. These analytical results raise the difficulty in overcoming environmental contamination of control animals and the choice of such controls in experimental studies which focus on neurotoxicity of exposure to low levels of pollutants.

Exclusive prenatal exposure to a 16 PAH mixture does not impact anxiety-related behaviours and regional brain metabolism in adult male rats: a role for the period of exposure in the modulation of PAH neurotoxicity.

Polycyclic aromatic hydrocarbons (PAHs) are persistent organic pollutants originating from incomplete combustion processes. Humans are mainly exposed through contaminated food ingestion. PAHs are neurotoxic compounds both for human and rodents, and may be found in placenta, umbilical cord blood and breast milk, suggesting that early exposure may impact developing central nervous system. In a previous study we showed that PAH exposure during both gestation and lactation periods in rats increased anxiety-related behaviours and decreased cerebral metabolism in several key structures linked to the limbic system on male pups at the adult stage. The aim of the present study was to assess the effects of an exclusive gestational PAH exposure on the same aspects of brain functionality. Female rats were exposed through diet to a 16 PAH mixture at doses of 2 µg/kg/day or 200 µg/kg/day during gestation. Late neurotoxic effects were evaluated by carrying out behavioural and cognitive tests and histochemical analyses using cytochrome oxidase activity as a cerebral metabolism marker in different brain areas. The results of this study revealed that behaviour and cerebral metabolism on prenatally PAH exposed adult rats was not significantly affected by the exposure to these pollutants. Finally this work highlights that the exposure period to pollutants such as PAHs at very early stages of development play a key role on the neurological impairment induced.

Late effects of a perinatal exposure to a 16 PAH mixture: Increase of anxiety-related behaviours and decrease of regional brain metabolism in adult male rats.

Polycyclic Aromatic Hydrocarbons (PAHs) are ubiquitous pollutants originated from incomplete combustion processes. Ingestion of contaminated food is the main route of exposure for humans. These molecules are able to cross the placental barrier and are also found in breast milk. Since PAHs are neurotoxic agents, the potential adverse effects of a perinatal exposure of the developing brain is a key issue for public health especially concerning PAH mixture. In this study, female rats were exposed trough diet to a mixture of 16 PAHs, at doses of 2 µg/kg/day or 200 µg/kg/day during gestation and 1.5 µg/kg/day or 150 µg/kg/day during breast-feeding period. To assess late neurotoxic effects in male offsprings, behavioural and cognitive tests were carried out and histochemical analyses using cytochrome oxidase as a cerebral metabolism marker were performed on adult animals. Results showed that anxiety-related behaviours significantly increased in exposed animals, but there was no significant alteration of motor activity and learning and memory abilities. Several brain areas of the limbic system showed a neuronal hypometabolism in exposed animals. This work highlights that exposure to PAHs at early stages of brain development can cause later troubles on behaviour and that PAHs are able to partly alter the central nervous system metabolism on adulthood.