Effects Of Long-Term BPA Exposure On Anxiety And Depression Behaviors In Adult Mice And Underlying Mechanisms

Bisphenol-A (BPA) is one of the typical environmental endocrine disruptors which had both weak-estrogenic, anti-estrogenic action and anti-androgens. BPA have a certain affinity with estrogen receptors, which can enter the body through a variety of ways such as food, beverages, skin contact and so on to endanger our health. There is increasing concern about the potential negative impact on brain and behavioral development of BPA. Recently, the effects of low dose exposure to BPA during fetal/neonatal stages have been extensively investigated. For example, fetal or neonatal exposure to BPA inhibits sexual differentiation of nonreproductive behaviors of adult animals, including learning-memory behavior, anxiety-and depression-like behavior. The sex hormone involved in regulating nerve behavior of adult animals. Prolonged exposure of adult female rats to a low dose of BPA during pregnancy and lactation altered maternal behavior. Our previous study indicated that exposure to BPA during adulthood sex-specifically affected the behaviors including spatial learning and memory, and the hippocampal synaptic plasticity processes, such as remodeling of spinal synapses and the expressions of synaptic proteins (e.g. synapsin I and PSD-95) and NMDA and AMPA receptors. Adult animals exposed to BPA in the environment for longer, suggesting that long-term exposure to BPA may affect the behavior of anxiety and depression in adult mice.ERβ and GABAA-2areceptors are known to be related to anxiety-and depression-related behaviors. The ERβ play an important role in the central nervous system (CNS). The ERβ selective agonists can decrease anxiety levels in mice, and ERβ knockout female mice exhibited increased levels of anxiety. The ERβ has an important role in the antidepressant effect of cognitive decline caused by middle-aged and elderly disease, and concentrated in the hippocampus region. Others’ research results also show that the ER is beta rather than alpha agonist reduce the immobility time of mice in the forced swimming test, and BPA can inhibit the expression of ERβin the hippocampus. Depression is related to the low of Gamma-aminobutyric acid (GABA). GABAA-2asubunit is associated with the pathophysiology of depression and can regulate anxiety behavior which is mainly located in the hippocampus and amygdala. GABAA-2α knockout mice showed the severity of depression in males. Therefore, Whether the effects of BPA on anxiety-and depression-like behaviors are related to the levels of ER β and GABAA-2αR.We hypothesized that long-term exposure to BPA in adulthood would sex-specifically induce excessive levels of anxiety-and depression-like behaviors. To verify this hypothesis, we use a variety of behavioral models to verify. In addition, we also assayed the ERP and GABAA-2αR activity in hippocampus brain region witch associated with anxiety and depression behaviors.The research content:To investigate the adverse impacts BPA on anxiety-and depression-like behaviors, explore the potential mechanism; provide the new idea of BPA on anxiety-and depression-like behaviors and its mechanism. Exposed to bisphenol A in the adult mice, investigated the effects of BPA on anxiety-and depression-like behaviors. To observed the effect of the long-term BPA exposure in mice of testosterone and estradiol levels in serum and brain, the ERβ (estrogen receptor-β) and GABAA-2αR (gamma-aminobutyric acid receptor-2a) levels in hippocampus.The research methods:Nine-week-old ICR mice, males and females were randomly divided into5groups by weight. BPA (0.04,0.4,4and40mg/kg body weight) by oral gavage once a day for12weeks. The control group given the same volume.Measuring changes in brain and blood levels of testosterone and estradiol levels by radioimmunoassay. The mice from the five groups received a battery of behavioral tests in the following order: the open field, Mirrored maze, elevated-plus maze and forced swim to test the anxiety and depression behaviors of male and female mice. Western blot analysis the GABAA-2αR ERβ protein expression in the hippocampus to explore the possible molecular mechanismsThe results of the study:1. No significant gender difference of body weight (BW) was found in mice at the age of10-week-old. After exposed to BPA for12weeks, a significant gender difference of BW was found (F(1,118)=3.51, p=0.01, males> females). Exposure to BPA had no significant effects on BW and the ratio of uterus or bilateral testis weight.2. Exposure to BPA for12weeks did not affect the levels of estradiol in the brain and serum, but significantly reduced the levels of testosterone in the brain and serum. Compared with the control males, the level of serum testosterone was significantly reduced by BPA at0.4mg/kg/day (p<0.05).3. Behavior test results:1) The open field test found that no significant interaction of sex x treatment in the number of grid crossings, the frequency rearing, the frequency of the grooming, and the time spent in the central area. A significant main effect of BPA exposure on the number of grid crossings was found, suggesting the stronger desire to explore and the lower anxiety state in males than females.2) In the Light-dark box task, BPA inhibits the activity in males and eliminate the gender difference.3) In the mirrored maze task, the interactions of sex x BPA treatment had no significant effects on both the total time staying in the mirrored chamber (F(4,110)=0.817, p=0.517) and the mirrored chamber entries (F(4,110)=0.399, p=0.809). The main factor of gender significantly affect the total time staying in the mirrored chamber (F(1,118)=3.965,p=0.049) but not the mirrored chamber entries in mice. When compared with the controls of the same sex, the mirrored chamber entries was significantly increased in females by BPA at0.04mg/kg/day, suggesting that BPA improved the locomotion of the females.4) The Elevated plus maze found that the influence of long-term exposure to BPA in adulthood can not be ignored. Consistent with previous reports, sex differences are found in the EPM task. The males had more frequent open arm entries and more time spent in open arms than the females (p<0.05), suggesting the higher anxiety-like status in females than in males. However, after exposure to BPA at0.4,4, and40mg/kg/day for12weeks, the number of the open arm entries and the time spent in open arms were decreased in males but were increased in females when compared with the vehicle controls of the same sex (p<0.01or p<0.05). These results suggest that long-term BPA exposure had an anxiogenic effect in males but an anxiolytic effect in female mice, and thus eliminated or reversed gender differences of anxiety status. In the control, a larger number of total entries in the open and closed arms was found in males than in females (p<0.05), suggesting a higher activity in males. Exposure to BPA, especially at0.4and40mg/kg/day, significantly decreased the number of total entries in the open and closed arms in males (p<0.05), suggesting that long-term BPA exposure inhibited the activity of the male mice, and thus eliminated the gender differences of activity.5) The Forced swim task found that the interaction of BPA treatment X sex and the main factor of BPA exposure were significantly influence the duration of immobility in the forced swim task (F(4,110)=5.014,p=0.001; F(4,115)=3.028,p=0.021), but no main effect of sex was found. An obvious sex differences in the duration of immobility was shown in the control mice (females>males,p<0.05), suggesting a higher level of depression-like in females than in males. BPA exposure at0.4-40mg/kg/day significantly prolonged the immobility time in the male mice (p<0.05and p<0.01), and thus eliminated the gender differences. The results suggest that BPA exposure significantly increased depression-like status of the male mice.4. Western blot analysis further indicated that BPA significantly down-regulated the expression of estrogen receptorβ (ERβ) in the hippocampus of the adult males but not females, and inhibited the expression of GABAA receptor submit2a (GABAA-2aR) in hippocampus of males but promoted that of females.Conclusion:Long-term BPA exposure can sex-specific affect the anxiety and depression of adult mice. This effect may be through changing the brain and serum sex hormone levels in mice. The changes of ERβ and GABAA-2aR protein in the hippocampus may be due in part to the change of the anxiety-and depression-like behaviors in adult mice.