PMC

(A) The white box outlines the area of the LH examined for NT peptide expression. Image modified from the Allen Mouse Brain Atlas. (B) A one-way ANOVA revealed a significant elevation in the density of NT peptide labeling in the postpartum LH compared to virgin females. Sample size: 7 virgins and 8 postpartum females. (C) Few cell bodies that contain NT mRNA are found in the LH (left panel). Image modified from the Allen Brain Atlas. NT peptide IHC revealed dense staining of fibers in the LH (right panel). * p<0.05. f = fornix, opt = optic tract.

Relative expression (y-axis) represents a ratio of gene expression in postpartum versus virgin females, with ratios under one indicating genes that have lower expression in lactating females. The box whisker plots demarcate the range (whiskers), interquartile range (box), and median (solid white line) for each gene tested. (A) In the LS, NT mRNA expression was significantly down-regulated in lactating females, and there was a nonsignificant decrease in Sort1 expression in postpartum females compared to virgin females. Sample size: 10 virgins and 11 postpartum females. (B) In the BnSTd, NT was significantly up-regulated in postpartum compared to virgin females. Sample size: 7 virgin and 7 postpartum females pooled from a total of 14 individuals from each group. (C) In the VTA, NT was nonsignificantly down-regulated in lactating females versus virgins. Sample size: 7 virgin and 7 postpartum females. * p<0.05; ** p<0.01; *** p<0.005.

(A) Bands were visualized at approximately 19 kDa, which corresponds with the molecular weight of the neurotensin/neuromedin N precursor protein, and at approximately 45 kDa. The identity of the protein weighing 45 kDa is unknown. (B) Example of the normalization method used to control for differences in background labeling. Background staining was relatively low across all sections analyzed (black square and top right panel), and was subtracted from the densitometric mean of regions of interest, such as the vLSM (white square and bottom right panel) in order to normalize brain sections analyzed. (C–D) NT staining was found in fibers and cell bodies (D, black arrows), which was expected. Regions labeled for NT were consistent with past immunohistochemical studies, suggesting that the antibody is specific to NT.

(A) Relative expression (y-axis) represents a ratio of gene expression in postpartum versus virgin females, with ratios over one indicating genes that have higher expression in lactating females. The box whisker plots demarcate the range (whiskers), interquartile range (box), and median (solid white line) for each gene tested. qPCR revealed significant up-regulation of NT and Sort1 in the postpartum MPOA, with a nonsignificant increase in Ntsr1. Sample size: 6 virgin and 8 postpartum females. (B) The white box outlines the area of the MPOA examined for NT peptide expression. Image modified from the Allen Mouse Brain Atlas. (C) A one-way ANOVA revealed a significant elevation in the density of NT peptide labeling in the MPOA of postpartum versus virgin females. Sample size: 7 virgins and 9 postpartum females. (D) Many cell bodies in the ventral and medial portion of the MPOA contain NT mRNA (left panel). Image modified from the Allen Mouse Brain Atlas. Dense NT peptide labeling is found throughout the MPOA (right panel), suggesting that NT peptide may be transported into the MPOA from other regions. * p<0.05; *** p<0.005. BST = bed nucleus of stria terminals.

(A) Relative expression (y-axis) represents a ratio of gene expression in postpartum versus virgin females, with ratios under one indicating genes that have lower expression in lactating females. The box whisker plots demarcate the range (whiskers), interquartile range (box), and median (solid white line) for each gene tested. qPCR revealed significant down-regulation of Ntsr1 in postpartum versus virgin females. Sample size: 7 virgin and 7 postpartum females pooled from a total of 14 individuals for each group. (B) The white box outlines the areas of the PVN examined for NT peptide expression. Images modified from the Allen Mouse Brain Atlas. (C) A one-way ANOVA revealed a significant elevation in the density of NT peptide labeling in the lPVN and cPVN of postpartum versus virgin females. Sample size: 7 virgins and 8 postpartum females for lPVN, and 7 virgins and 7 postpartum females for cPVN. (D–E) Very few cell bodies that contain NT mRNA are found in certain subregions of the PVN (D and E, left panels). Images modified from the Allen Brain Atlas. NT peptide IHC revealed dense staining of fibers in the mPVN and cPVN (D and E, right panels), and moderate staining in the lPVN (D, right panel). This suggests that NT peptide is transported into the PVN for release. * p<0.05; ** p<0.01; *** p<0.005. AHN = anterior hypothalamic nucleus, opt = optic tract, lPVN = lateral PVN, mPVN = medial PVN, cPVN = caudal PVN.