Satoshi Yokota

Effect of prenatal exposure to diesel exhaust on dopaminergic system in mice.

Diesel exhaust (DE) is composed of particles and gaseous compounds. It has been reported that DE causes pulmonary and cardiovascular disease. We have previously reported that fetal exposure to DE had deleterious effects to the reproductive system of mice offspring. However, there is still little known about the effects of prenatal exposure to DE to the central nervous system (CNS). In the present study, we found that prenatal exposure to DE induced reduction of locomotion, furthermore, dopamine (DA) turnover was significantly decreased in the striatum and nucleus accumbens. These results suggest that prenatal exposure to DE has an effect on the CNS. Hypolocomotion could be due to a decrease in DA turnover associated with DA nervous system abnormality. The present study provides the possibility that maternally inhaled DE might influence the development of central dopaminergic system and result in behavior disorder.

Gene Expression Changes in the Olfactory Bulb of Mice Induced by Exposure to Diesel Exhaust Are Dependent on Animal Rearing Environment

There is an emerging concern that particulate air pollution increases the risk of cranial nerve disease onset. Small nanoparticles, mainly derived from diesel exhaust particles reach the olfactory bulb by their nasal depositions. It has been reported that diesel exhaust inhalation causes inflammation of the olfactory bulb and other brain regions. However, these toxicological studies have not evaluated animal rearing environment. We hypothesized that rearing environment can change mice phenotypes and thus might alter toxicological study results. In this study, we exposed mice to diesel exhaust inhalation at 90 µg/m3, 8 hours/day, for 28 consecutive days after rearing in a standard cage or environmental enrichment conditions. Microarray analysis found that expression levels of 112 genes were changed by diesel exhaust inhalation. Functional analysis using Gene Ontology revealed that the dysregulated genes were involved in inflammation and immune response. This result was supported by pathway analysis. Quantitative RT-PCR analysis confirmed 10 genes. Interestingly, background gene expression of the olfactory bulb of mice reared in a standard cage environment was changed by diesel exhaust inhalation, whereas there was no significant effect of diesel exhaust exposure on gene expression levels of mice reared with environmental enrichment. The results indicate for the first time that the effect of diesel exhaust exposure on gene expression of the olfactory bulb was influenced by rearing environment. Rearing environment, such as environmental enrichment, may be an important contributive factor to causation in evaluating still undefined toxic environmental substances such as diesel exhaust.

Social Isolation-Induced Territorial Aggression in Male Offspring Is Enhanced by Exposure to Diesel Exhaust during Pregnancy

Diesel exhaust particles are a major component of ambient particulate matter, and concern about the health effects of exposure to ambient particulate matter is growing. Previously, we found that in utero exposure to diesel exhaust affected locomotor activity and motor coordination, but there are also indications that such exposure may contribute to increased aggression in offspring. Therefore, the aim of the present study was to test the effects of prenatal diesel exhaust exposure on social isolation-induced territorial aggression. Pregnant mice were exposed to low concentrations of diesel exhaust (DE; mass concentration of 90 μg/m3: DE group: n = 15) or clean air (control group: n = 15) for 8 h/day during gestation. Basal locomotion of male offspring was measured at 10 weeks of age. Thereafter, male offspring were individually housed for 2 weeks and subsequently assessed for aggression using the resident−intruder test at 12 weeks of age, and blood and brain tissue were collected from the male offspring on the following day for measuring serum testosterone levels and neurochemical analysis. There were no significant differences in locomotion between control and DE-exposed mice. However, DE-exposed mice showed significantly greater social isolation-induced territorial aggressive behavior than control mice. Additionally, socially-isolated DE-exposed mice expressed significantly higher concentrations of serum testosterone levels than control mice. Neurochemical analysis revealed that dopamine levels in the prefrontal cortex and nucleus accumbens were higher in socially isolated DE-exposed mice. Serotonin levels in the nucleus accumbens, amygdala, and hypothalamus were also lower in the socially isolated DE-exposed mice than in control mice. Thus, even at low doses, prenatal exposure to DE increased aggression and serum testosterone levels, and caused neurochemical changes in male socially isolated mice. These results may have serious implications for pregnant women living in regions with high levels of traffic-related air pollution.

In utero exposure of mice to diesel exhaust particles affects spatial learning and memory with reduced N-methyl-D-aspartate receptor expression in the hippocampus of male offspring.

Diesel exhaust consists of diesel exhaust particles (DEPs) and gaseous compounds. Previous studies reported that in utero exposure to diesel exhaust affects the central nervous system. However, there was no clear evidence that these effects were caused by diesel exhaust particles themselves, gaseous compounds, or both. Here, we explored the effects of in utero exposure to DEPs on learning and memory in male ICR mice. DEP solutions were administered subcutaneously to pregnant ICR mice at a dose of 0 or 200 μg/kg body weight on gestation days 6, 9, 12, 15, and 18. We examined learning and memory in 9-to-10-week-old male offspring using the Morris water maze test and passive avoidance test. Immediately after the behavioral tests, hippocampi were isolated. Hippocampal N-methyl-D-aspartate receptor (NR) expression was also measured by quantitative RT-PCR analysis. Mice exposed to DEPs in utero showed deficits in the Morris water maze test, but their performance was not significantly different from that of control mice in the passive avoidance test. In addition, DEP-exposed mice exhibited decreased hippocampal NR2A expression. The present results indicate that maternal DEP exposure disrupts learning and memory in male offspring, which is associated with reduced hippocampal NR2A expression.

In utero exposure to diesel exhaust particles induces anxiogenic effects on male offspring via chronic activation of serotonergic neuron in dorsal raphe nucleus.

Diesel exhaust consists of diesel exhaust particles (DEPs) and gaseous compounds. Because previous research suggested that in utero exposure to DEPs affected spatial learning and memory in male offspring, while epidemiological evidence suggested disturbances in affect after prenatal exposure to particulates, we hypothesized that DEP exposure during pregnancy might also disturb affect. Here, we explored the effects of in utero exposure to DEPs on anxiety in male ICR mice. DEP solutions were administered subcutaneously to pregnant ICR mice at a dose of 0 or 200 μg/kg body weight on gestation days 6, 9, 12, 15, and 18. We assessed anxiety in 6 week-old male offspring using the hole board test and elevated plus maze test. After the behavioral tests, animals were sacrificed and serotonin (5-HT) levels in the dorsal raphe nucleus (DRN) were measured using HPLC. Mice exposed to DEPs in utero demonstrated increased anxiety in both behavioral tests. HPLC analysis revealed a significant increase in 5-HT levels in the DRN. Double immunolabeling of the DRN using anti-5-HT and anti-FosB (a chronic neuronal activation marker) antibodies indicated chronic activation of the DRN might underlie the increased anxiety after prenatal DEP exposure.

Prenatal exposure to diesel exhaust particles affected cholinergic systems in senile mice

balance ability was measured by a stabilometer, G6100 (Anima Ltd.) The training prolonged the balance holding time while standing on one foot with both eyes closed or eyes opend, and reduced the locus length per unit area both while standing on one foot with and without eyes closed, shortened the performance time of Kazufusen, but did not show a significant effect on Kana Pick Up Test. The leg muscle strength and body weight ratio were not changed before and after the training. These results indicate the existence of interaction between the cognitive and vestibular function.

Analysis of neuronal lipid metabolism in mouse by phospholipids-modified nanoparticles

TrkB, a member of the receptor tyrosine kinase family, is established as a crucial molecule regulating cell functions in a nervous system. However, physiological function of TrkB within local region in cells largely remains unclear. To clarify the function in restricted region in cells, here we developed a photo-activation system which can activate TrkB signaling with high spatiotemporal resolution in living cells. To control the activity of TrkB, we designed chimeric receptor constructs by replacing the extracellular domain of TrkB with an anti-fluorescein single-chain antibody. This chimeric receptor was expected to be activated upon closslinking by multiple fluorescein-labeled molecule (OligoFL), and then triggers TrkB specific signal transduction. To test whether chimeric receptor can work as expected, intracellular phosphorylation of chimeric receptor and calcium response were examined in NIH3T3 cells expressing the chimeric receptor. On this test, phosphorylation of chimeric receptor on the intracellular domains and increase in intracellular Ca2+ concentrations were observed upon OligoFL application. These results suggest that the chimeric receptor can transmit downstream signaling pathways. Furthermore, aiming at controlling of TrkB signaling with high spatiotemporal resolution, we developed a light driven ligand activation system. We generated caged-fluorescein labeled macromolecule (caged-OligoFL), and then expected that irradiation triggers uncaging of caged-OligoFL and generate OligoFL. Irradiation to NIH3T3 cells expressing chimeric receptor in the presence of cagedOligoFL elicited increase in intracellular Ca2+ concentrations. This result suggests that our strategy enables spatially-controlled activation of TrkB signaling by irradiation within a restricted region. The combination of chimeric receptor and its specific caged-ligand and irradiation constitutes a new research platform for studying neuronal circuits, motility and polarity.

Impairment of spatial learning and memory induced by maternal exposure to diesel exhaust particles

nonanone or 1-octanol to the nematode C. elegans causes enhancement of avoidance behavior, rather than adaptation, to these odors. We have also found that (1) the memory of preexposure is maintained for at least 1 h and that (2) the enhancement requires dopamine biosynthesis in the animals. Here, we report that a D2-like dopamine receptor is required for the enhancement of 2-nonanone avoidance. We found that D2 dopamine receptor antagonists specifically suppressed the enhancement of preexposed animals. Further, we found that deletion mutations in a D2-like receptor significantly affected the enhancement. These results suggest that dopamine signaling pathway regulates a novel behavioral plasticity in the animals.