Prolactin Secretion During Postnatal Development in Artificial Rearing Rats (Rattus norvegicus)

Abstract

During the lactation period, rat pups are fed by their mother and are with their siblings. In the pituitary, lactotrope and somatolactotrope cells increase in number during this period and are associated with milk-borne factors. In the artificial rearing (AR) paradigm, pups are deprived of mother- and sibling-pup interactions and are fed artificial milk. AR rats present more hypothalamic neurons associated with less apoptosis during postnatal development. Moreover, AR infantile rats show growth hormone (GH), insulin-like growth factor-1 (IGF-1), and ghrelin alterations resulting from suckling behavior and the absence of meal transition at the second week of life. In the present study, the variation in prolactin (PRL) throughout the lactation period was analyzed in AR pups and compared with that in mother-reared (MR) pups. At postnatal Day 7 (pnd7), AR pups have less serum PRL than MR pups do, and a further decrease was observed at pnd14. However, only at pnd14, AR pituitary cells secrete less PRL, which was correlated with a smaller number of somatolactotrope cells unlike lactotrope cells. Analysis of the hypothalamic dopamine and DOPAC concentrations in both groups revealed no differences at 7, 14, and 21pnd. Nevertheless, the pituitary showed higher concentrations in AR pups than in MR pups at pnd14. However, the number of dopaminergic neurons in the arcuate nucleus was similar in both groups, but they were less spread in the AR pup hypothalamus. Our results revealed the importance of mothers' and siblings' interactions and mothers' milk in the maturation of the PRL hypothalamic‒pituitary axis during the lactating period.

Keywords: artificial rearing; dopamine/DOPAC; infantile period; pituitary cells; prolactin.

FIGURE 1

Effect of the artificial rearing procedure on the serum PRL concentration of rat pups at pnd7, pnd14 and pnd21. PRL serum concentrations of MR (open bars) and AR (dark bars) pups after a 4 h fasting period at pnd7, pnd14, and pnd21. The numbers per group and age of pups were pnd7, MR 11, and AR 11; pnd14, MR 11, and AR 10; pnd21, MR 15, and AR 9. Statistical analysis was performed via two‐way ANOVA, followed by Bonferroni's posttest for differences between ages in the MR or AR groups, ^# p < 0.05, ^## p < 0.01, and Student's t test and Welch corrected for differences between the MR and AR groups at each age, ^* p < 0.05, ^**** p < 0.0001.

FIGURE 2

Effect of the artificial rearing procedure on basal and bromocriptine‐stimulated PRL secretion in primary pituitary cell cultures from rat pups at pnd7, pnd14, and pnd21. Primary cell cultures from pituitaries from MR (open bars) and AR (dark bars) pups at (A) pnd7, (B) pnd14, and (C) pnd21 were performed, and after 24 h of culture, basal secretion followed by bromocriptine‐stimulated secretion was evaluated, “+” and “−” correspond to stimulated and basal secretion, respectively. Statistical analyses were performed with two‐way ANOVA followed by Bonferroni's posttest for differences between the MR and AR groups, ^# p < 0.05, ^## p < 0.01, and a paired t‐test for differences between the basal and bromocriptine‐stimulated groups, ^* p < 0.05, ^** p < 0.01, ^*** p < 0.001.

FIGURE 3

Percent bromocriptine inhibition of PRL secretion in pituitary cultures from maternal and artificial rearing pups at different postnatal ages. The percentage of PRL secretion inhibition by bromocriptine in pituitary culture cells from MR (open bars) and AR (drak bars) pups in the experiments shown in Figure 2. Statistical analysis was performed via two‐way ANOVA, followed by Bonferroni's posttest for differences between ages in the MR or AR groups, ^# p < 0.05, ^## p < 0.01, ^### p < 0.001.

FIGURE 4

Effects of the artificial rearing procedure on somatotrope and somatolactotype cell populations of pituitaries from rat pups at pnd7, pnd14, and pnd21. Pituitary cell cultures from MR (open bars) and AR (drak bars) pups at pnd7, pnd14, and pnd21 were performed, and after 24 culture periods, the cells were immunoassayed (A), and the percentages of positive cells for (B) GH, (C) PRL, and (D) both hormones were determined. Cells stained for GH in red (thin arrows), for PRL in green (thick arrows), GH and PRL in red and green or yellow (arrow heads) and nucleus in blue. Statistical analyses were performed via two‐way ANOVA, followed by Bonferroni's test for differences between ages in the MR or AR groups, ^### p < 0.001, and Student's t test and Welch corrected for differences between the MR and AR groups, ^* p < 0.05, ^** p < 0.01. Barr = 50 µm.

FIGURE 5

Effect of the artificial rearing procedure on the population of tyrosine hydroxylase (TH⁺)‐positive cells in the arcuate nucleus. The hypothalami from MR and AR rat pups at pnd14 were dissected, and immunohistochemistry for TH was performed. Representative images showing TH⁺ cells of the arcuate nucleus of MR (left) and AR (right) rats (A). The number of TH⁺ cells in the arcuate nucleus area (B) of MR (empty dots) and AR (filled dots) pups was counted. An example of the distribution of TH⁺ cells along the arcuate nucleus of MR (empty bars) and AR (filled bars) is shown in (C).