Maternal programming of sexual behavior and hypothalamic-pituitary-gonadal function in the female rat

Abstract

Variations in parental care predict the age of puberty, sexual activity in adolescence and the age at first pregnancy in humans. These findings parallel descriptions of maternal effects on phenotypic variation in reproductive function in other species. Despite the prevalence of such reports, little is known about potential biological mechanisms and this especially true for effects on female reproductive development. We examined the hypothesis that parental care might alter hypothalamic-pituitary-ovarian function and thus reproductive function in the female offspring of rat mothers that vary pup licking/grooming (LG) over the first week postpartum. As adults, the female offspring of Low LG mothers showed 1) increased sexual receptivity; 2) increased plasma levels of luteinizing hormone (LH) and progesterone at proestrus; 3) an increased positive-feedback effect of estradiol on both plasma LH levels and gonadotropin releasing-hormone (GnRH) expression in the medial preoptic region; and 4) increased estrogen receptor alpha (ERalpha) expression in the anterioventral paraventricular nucleus, a system that regulates GnRH. The results of a cross-fostering study provide evidence for a direct effect of postnatal maternal care as well as a possible prenatal influence. Indeed, we found evidence for increased fetal testosterone levels at embryonic day 20 in the female fetuses of High compared to Low LG mothers. Finally, the female offspring of Low LG mothers showed accelerated puberty compared to those of High LG mothers. These data suggest maternal effects in the rat on the development of neuroendocrine systems that regulate female sexual behaviour. Together with studies revealing a maternal effect on the maternal behavior of the female offspring, these findings suggest that maternal care can program alternative reproductive phenotypes in the rat through regionally-specific effects on ERalpha expression.

Figure 1. Mating behavior of the female offspring and the effect of cross-fostering.

(a) Tested with a stud male in a small traditional testing arena, the mean (±SEM) lordosis rating in the adult female offspring of High LG mothers was lower compared to Low and Mid offspring (n = 10–12/group).* p<0.05. (b) Tested in a pacing chamber, the mean (±SEM) lordosis rating of adult female offspring of High LG mothers was lower compared to Low female offspring. * p<0.05. The inter-intromission interval showed no significant differences (n = 12/group). (c) Mean±SEM lordosis rating or inter-intromission interval in the adult female offspring of Low or High LG mothers fostered to and reared by Low (Low-Low; High-Low) or High (Low-High; High-High) tested with a stud male in the pacing chamber (n = 7–13/group). Groups lying underneath the line differ significantly (p <0.05) from the Low-Low group. (d) Mean±SEM testosterone level in amniotic fluid of embryonic day 20 female offspring of High or Low LG mothers (n = 20–25 pups from 12–14 litters/group; *p<.01).

Figure 2. Hormonal levels during proestrus.

Mean±SEM plasma levels of lutenizing hormone (LH), estradiol and progesterone in the adult offspring of High or Low LG mothers (n = 8–10/group) over the day of proestrus (lights off at 12:00).

Figure 3. Estrogen effects on plasma LH and GnRH immunoreactive cells.

(a) Mean±SEM plasma level of lutenizing hormone (LH) in ovariectomized adult female offspring of High or Low (n = 5–7/group) provided with low-level E2B replacement and then treated acutely with 0, 3 or 10 µg of E2B 24h prior to sampling. Adult female offspring of Low LG mothers treated with 10 µg of E2B showed a greater level of LH than all other groups (* p<0.05). (b) Photomicrograph of representative GnRH immunolabeling in the medial preoptic area of High or Low provided with low-level E2B replacement and then treated acutely with oil or 10 µg of E2B 24h prior to tissue sampling. The right panel reveals that the mean±SEM total number of GnRH-labeled cells located in the preoptic region (anteroventral preoptic nucleus, the median and lateral preoptic nucleus, the medial preoptic area) as well as the diagonal band nucleus in the Low offspring treated with E2B was greater than all other groups (n = 5/group;* p<0.05 ).

Figure 4. PhosphoERα mRNA and immunoreactivity in intact and estrogen-treated females.

(a) Left panel shows representative autoradiograms of ERα mRNA expression in the anteroventral paraventricular nucleus of the hypothalamus (AVPVn; outlined region) in the adult female offspring of High or Low LG mothers. The right panel shows the mean±SEM for the quantification (relative optical density) of ERα mRNA in the AVPVn from the adult offspring of High or Low LG mothers (n = 5/group). * p<0.05. (b) Representative western blot depicting phosphoERα immunoreactivity at the expected 66kDa band in ovariectomized (O) treated with oil or 10 µg E2B, 90 min prior to tissue sampling. The Western blots revealed a limited banding pattern and the expected increase in phosphoERα−labelling. The right panels depict representative phosphoERα−labelling at the level of the AVPVn in ovariectomized adult female offspring of High or Low (n = 5–6/group) provided with low-level E2B replacement and then treated acutely with 10 µg of E2B or oil, 90 min prior to tissue sampling. (c) Mean±SEM number of phosphoERα−labeled cells in the AVPVn of the animals described above. Statistical analysis revealed a significant (p<0.05) overall effect of maternal care such that the number of phosphoERα−labeled cells was increased in the offspring of Low LG mothers in both treatment conditions.

Figure 5. Effect of ovariectomy and steroid treatment in the sexual behavior of the female.

Mean±SEM lordosis rating or inter-intromission interval in the adult female offspring of Low or High LG mothers tested with a stud male in the pacing chamber arena (n = 7–10/group) either intact (at proestrus) or following ovariectomy with E2B+progesterone replacement. For the lordosis rating data, groups lying underneath the line differ significantly (p <0.05) from the High LG/intact group such that the maternal effect is apparent among intact animals, but not ovariectomized animals with a common level of steroid replacement. For the inter-intromission interval data, the maternal effect is apparent (* p <0.05) among both intact and ovariectomy/steroid-primed conditions.

Figure 6. Onset of puberty.

Mean±SEM day of vaginal opening in the female offspring of High (n = 16) and Low (n = 12) LG mothers revealing a maternal effect on the timing of pubertal development. * p<0.05