Toxicity evaluation of bisphenol A administered by gavage to Sprague Dawley rats from gestation day 6 through postnatal day 90

Abstract

Bisphenol A (BPA) is a high production volume industrial chemical to which there is widespread human oral exposure. Guideline studies used to set regulatory limits detected adverse effects only at doses well above human exposures and established a no-observed-adverse-effect level (NOAEL) of 5 mg/kg body weight (bw)/day. However, many reported animal studies link BPA to potentially adverse effects on multiple organ systems at doses below the NOAEL. The primary goals of the subchronic study reported here were to identify adverse effects induced by orally (gavage) administered BPA below the NOAEL, to characterize the dose response for such effects and to determine doses for a subsequent chronic study. Sprague Dawley rat dams were dosed daily from gestation day 6 until the start of labor, and their pups were directly dosed from day 1 after birth to termination. The primary focus was on seven equally spaced BPA doses (2.5-2700 μg/kg bw/day). Also included were a naïve control, two doses of ethinyl estradiol (EE2) to demonstrate the estrogen responsiveness of the animal model, and two high BPA doses (100,000 and 300,000 μg/kg bw/day) expected from guideline studies to produce adverse effects. Clear adverse effects of BPA, including depressed gestational and postnatal body weight gain, effects on the ovary (increased cystic follicles, depleted corpora lutea, and antral follicles), and serum hormones (increased serum estradiol and prolactin and decreased progesterone), were observed only at the two high doses of BPA. BPA-induced effects partially overlapped those induced by EE2, consistent with the known weak estrogenic activity of BPA.

Keywords: 90-day study; Bisphenol A; Sprague Dawley rat; ethinyl estradiol.

FIG. 1.

Schematic representation of the study experimental design. A detailed description of the protocol is found in the Materials and Methods section. Blood draws and terminal necropsies were conducted in PND 80 ± 3 and PND 90 ± 5 windows, respectively. The indicated measurements of serum BPA levels are reported (Churchwell et al., in addition to the animals indicated here, male and female pups from each litter were maintained until termination at PND 90 ± 5 and tissues were transferred to other studies (results not reported here). Data obtained from those animals were included in the analyses of AGD and pubertal landmarks.

FIG. 2.

F₁ female body weights. Mean weekly body weights (g) are shown for the study-defined subgroups (low BPA, high BPA, EE₂, and naïve) versus vehicle.

FIG. 3.

F₁ male body weights. Mean weekly body weights (g) are shown for the study-defined subgroups (low BPA, high BPA, EE₂, and naïve) versus vehicle.

FIG. 4.

Ovary histology of animals dosed with the vehicle [(A) and (B)] or 300,000 μg BPA/kg bw/day [(C) and (D)]; tissue sections stained with hematoxylin and eosin. (A) Ovary from a normal cycling rat. A minimum of three generations (sets) of corpora lutea (*) from current and previous consecutive estrous cycles are present. The sets were determined morphologically by the relative size, cellularity, and lipochrome pigment content typically present in various stages of corpora luteal growth, degeneration, and regression. (B) Normal appearing antral follicles (arrows) in various stages of maturation from early to late. (C) Numerous cystic follicles (CF) and depletion of corpora lutea [*, compare with panel (A)]. Follicular cysts and fewer sets of corpora lutea than normal imply that either the estrous cycle has been arrested or prolonged. (D) Lack of normal appearing antral follicles [compare with panel (B)]. Depletion of antral follicles was defined as less than two to three normal appearing antral follicles and/or a relative increase in atretic ones. Severity for cystic follicles was subjectively based on the relative number of cysts present in sections; severity for corpora lutea depletion was based on the number of sets present (three sets only = grade 1; two sets = grade 2; one set = grade 3; and no sets = grade 4); depletion of antral follicles coded as present or absent. Bar, 50 μm.