Anxiogenic Effects of Developmental Bisphenol A Exposure Are Associated with Gene Expression Changes in the Juvenile Rat Amygdala and Mitigated by Soy

<div><p>Early life exposure to Bisphenol A (BPA), a component of polycarbonate plastics and epoxy resins, alters sociosexual behavior in numerous species including humans. The present study focused on the ontogeny of these behavioral effects beginning in adolescence and assessed the underlying molecular changes in the amygdala. We also explored the mitigating potential of a soy-rich diet on these endpoints. Wistar rats were exposed to BPA via drinking water (1 mg/L) from gestation through puberty, and reared on a soy-based or soy-free diet. A group exposed to ethinyl estradiol (50 µg/L) and a soy-free diet was used as a positive estrogenic control. Animals were tested as juveniles or adults for anxiety-like and exploratory behavior. Assessment of serum BPA and genistein (GEN), a soy phytoestrogen, confirmed that internal dose was within a human-relevant range. BPA induced anxiogenic behavior in juveniles and loss of sexual dimorphisms in adult exploratory behavior, but only in the animals reared on the soy-free diet. Expression analysis revealed a suite of genes, including a subset known to mediate sociosexual behavior, associated with BPA-induced juvenile anxiety. Notably, expression of estrogen receptor beta (<em>Esr2</em>) and two melanocortin receptors (<em>Mc3r</em>, <em>Mc4r</em>) were downregulated. Collectively, these results show that behavioral impacts of BPA can manifest during adolescence, but wane in adulthood, and may be mitigated by diet. These data also reveal that, because ERβ and melanocortin receptors are crucial to their function, oxytocin/vasopressin signaling pathways, which have previously been linked to human affective disorders, may underlie these behavioral outcomes.</p> </div

BPA, EE and dietary effects on adult Elevated Plus Maze (EPM) activity.

<p>As expected, a significant gender effect was found for all measurements except latency to enter an open arm. In general, BPA exposure eliminated these sex differences. (A) A higher percent of females entered the open arms than males regardless of exposure group. (B) Soy-free males took significantly longer to enter an open arm than females maintained on the same diet. BPA exposure eliminated this sex difference, with a tendency for reversal. Soy diet also eliminated this behavioral sex difference. (C) Females made significantly more open arm entries than males in all groups except those on the soy-free diet and exposed to BPA (BPA). There was no overall effect of BPA or diet on this behavior. (D) Similarly, in all groups except those on the soy-free diet and exposed to BPA, females spent significantly more time on the open arms than males. Graphs depict mean ± SEM, *<i>P</i>≤0.05, ** <i>P</i>≤0.001.</p

BPA, EE and dietary effects on juvenile Elevated Plus Maze (EPM) activity.

<p>(A) BPA exposure significantly decreased the percent of males that entered an open arm. No significant effect of gender was found for any subsequent measure, therefore the data were collapsed across sexes. (B) There were no significant group differences in latency to enter the open arms. (C) A main effect of diet on mean number of open arm entries was identified (not depicted), with soy-fed animals making significantly more entries. BPA exposed animals on the soy-free diet (BPA) made fewer entries compared to the diet matched controls (Soy-free) and also the BPA exposed animals on the soy diet (BPA + Soy). (D) Soy-based diet significantly increased time spent on the open arms, regardless of BPA exposure. EE had no significant effect on any EPM endpoint examined. Graphs depict mean ± SEM, *<i>P</i>≤0.05, **<i>P</i>≤0.001, # main effect of diet; <i>P</i>≤0.05.</p

Mean Pup Intake (ml) and Exposure Levels (µg) of BPA or EE.

<p>Average daily water consumption was quantified from the pups during postnatal days 21–40, from which average daily BPA or EE was calculated. Soy exposure occurred via diet only and therefore is not included.</p

Mean Dam Fluid Intake (ml) and Exposure Levels (µg) of BPA or EE.

<p>Average daily water consumption was quantified from the dams during mid-gestation and mid-lactation, from which average daily BPA or EE was calculated. Soy exposure occurred via diet only and therefore is not included.</p

Fold change in juvenile amygdalar gene expression levels.

<p>Of the 48 genes examined, a significant main effect of, or interaction with, BPA was found for the 8 genes shown here. The genes depicted met the threshold criteria established for biological significance. For each gene, the raw <i>P</i>-values obtained for the main and interaction effects of BPA are indicated. Significant group differences within each sex, identified by post-hoc t-tests, are also shown. <i>Esr2</i> (ERβ) and <i>Gad2</i> expression were significantly down-regulated by BPA in both sexes compared to Soy-free controls. <i>Tac2</i> and <i>Mc4r</i> were significantly down-regulated by BPA exposure in females. Among the soy-fed animals, no significant effect of BPA was identified for any gene, indicating a protective influence of soy. Graphs depict mean fold change in expression levels ± SEM. *<i>P</i>≤0.05, †<i>P</i>≤0.1 compared to soy-free controls.</p