Prenatal EDCs Impair Mate and Odor Preference and Activation of the VMN in Male and Female Rats

Abstract

Environmental endocrine-disrupting chemicals (EDCs) disrupt hormone-dependent biological processes. We examined how prenatal exposure to EDCs act in a sex-specific manner to disrupt social and olfactory behaviors in adulthood and underlying neurobiological mechanisms. Pregnant rat dams were injected daily from embryonic day 8 to 18 with 1 mg/kg Aroclor 1221 (A1221), 1 mg/kg vinclozolin, or the vehicle (6% DMSO in sesame oil). A1221 is a mixture of polychlorinated biphenyls (weakly estrogenic) while vinclozolin is a fungicide (anti-androgenic). Adult male offspring exposed to A1221 or vinclozolin, and females exposed to A1221, had impaired mate preference behavior when given a choice between 2 opposite-sex rats that differed by hormone status. A similar pattern of impairment was observed in an odor preference test for urine-soaked filter paper from the same rat groups. A habituation/dishabituation test revealed that all rats had normal odor discrimination ability. Because of the importance of the ventrolateral portion of the ventromedial nucleus (VMNvl) in mate choice, expression of the immediate early gene product Fos was measured, along with its co-expression in estrogen receptor alpha (ERα) cells. A1221 females with impaired mate and odor preference behavior also had increased neuronal activation in the VMNvl, although not specific to ERα-expressing neurons. Interestingly, males exposed to EDCs had normal Fos expression in this region, suggesting that other neurons and/or brain regions mediate these effects. The high conservation of hormonal, olfactory, and behavioral traits necessary for reproductive success means that EDC contamination and its ability to alter these traits has widespread effects on wildlife and humans.

Keywords: endocrine-disrupting chemicals (EDCs); mate preference; olfactory discrimination; polychlorinated biphenyls (PCBs); ventromedial nucleus; vinclozolin.

Figure 1.

Time spent near the stimulus rats during 10 minutes of exploration. (A) Females exposed to DMSO and VIN spent more time near GDX + T males over GDX males, whereas PCB females did not show a preference. (B) Males exposed to DMSO showed a preference for OVX + E2 + P4 females over OVX females; this was abolished in PCB and VIN males. *P < 0.05; **P < 0.01; ****P < 0.0001; ns, not significant.

Figure 2.

Time spent near the odor stimuli during 10 minutes of exploration. (A) Females exposed to DMSO and VIN spent more time near urinary odors from GDX + T males over GDX males, whereas PCB females did not show a preference. (B) Males exposed to DMSO showed a preference for odors from OVX + E2 + P4 females over OVX females; this was abolished in PCB and VIN males. *P < 0.05; **P < 0.01; ****P < 0.0001; ns, not significant.

Figure 3.

Habituation-dishabituation results. The type of odor stimulus is indicated in the box below the y-axis for trials 1-4 (presentation of first odor) and trial 5 (switch). Females of all treatments were able to discriminate between neutral odors (A) and sexually relevant odors (C, E). Males of all treatments were able to discriminate between neutral odors (B) and sexually relevant odors (D, F). Asterisks refer to significant differences between trial 4 and 5 (the latter when the stimulus was switched) to evaluate dishabituation; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001. Abbreviations: GDX, castrated; GDX + T, castrated with testosterone replacement; OVX, ovariectomized; OVX + E2 + P4, ovariectomized with estradiol and progesterone replacement.

Figure 4.

The percent of Fos+ cells in the VMNvl (A, B) of females and males perfused 90 minutes after the odor preference test. (A) Prenatal exposure to PCB increased the percent of Fos+ cells in the VMNvl of females. (B) Fos+ cells did not differ by treatment in the VMNvl of males. Bars with different letters are significantly different within a sex. For females, n = 5 DMSO, 8 PCB, 8 VIN; for males: 7 DMSO, 6 PCB, 7 VIN.

Figure 5.

Representative images of Fos and ERα immunofluorescence in the VMNvl of females euthanized 90 minutes after odor preference behavior. Scale bar = 25 µm.

Figure 6.

The number of Fos+ cells in the VMNvl of females (A) and males (B). Females exposed to PCBs had significantly more Fos+ cells than DMSO and VIN females. Males of all treatments had similar numbers of Fos+ cells. The number of ERα+/Fos+ double-labeled cells in the VMNvl of females (C) and males (D). Females and males of all 3 treatments had no significant differences in double-labeled cells in the VMNvl, and overall numbers were very low. Bars with different letters are significantly different within a sex. For females, n = 6 DMSO, 6 PCB, 6 VIN. For males, n = 6 DMSO, 5 PCB, 5 VIN.