Location: Children's Nutrition Research Center
2022 Annual Report
Objectives
Objective 1. In wild-type animals and transgenic animals lacking a functional Connexin43 gap junction within the adipocyte, characterize the effects of Connexin43 deletion on (1) the quantity of the milk a dam produces; (2) the composition of the milk, especially the lipid composition; and (3) other non-nutritional components in the milk.
Objective 2. Characterize the effects of a dam's adipocyte Connexin43 gap junction ablation on her offspring's body weight, adipose tissue mass, and late-life responses to metabolic challenges such as overnutrition.
Objective 3: Determine whether central supplement of E2 will attenuates lactational hyperphagia and hyperprolactinemia.
Objective 4: Determine whether activation of ERalphavlVMH neurons attenuates lactational hyperphagia and hyperprolactinemia.
Objective 5: Determine whether inhibition of ERalphavlVMH neurons rescues the reduced lactational hyperphagia and hyperprolactinemia by central supplement of E2.
Objective 6: Determine which ERalphavlVMH circuit is inhibited during lactation.
Approach
Adipocytes, the primary cell type in the non-lactating breast, display a drastic morphological change during the lactation in rodent studies: lipid-filled cells undergo lipolysis to provide fatty acids for triglyceride synthesis and also as an energy source to support milk production; they come back as lipid-laden cells once the animal is done lactating. Our objective is to assess whether breast-milk composition can be regulated by maternal adipose tissue physiology – more specifically, adipose tissue gap junction. The results of this research can be leveraged to improve milk quantity and quality to benefit infant growth and future metabolic status. The central hypothesis is that adipose tissue responds to metabolic and hormonal cues to support milk production and that the Connexin43 gap junction is required to facilitate this process by coupling a group of adipocytes together. Genetic manipulation of adipose tissue Connexin43 will be used to determine the effects of Connexin43 deletion on milk production and milk composition as well as on the offspring's metabolic health.
Additionally, researchers will develop mouse models to specifically activate or inhibit ERavlVMH neurons during lactation. We will measure serum PRL levels in lactating dams and monitor the metabolic status of lactating dams and their pups, including daily body weight and food intake of the dams, as well as the survival and body weight of pups. Further, we will also map the ERavlVMH originated neuron circuits and compare their neuron activities between virgin and lactating female mice. Accomplishment of the studies will identify a novel neuroendocrine mechanism underlying hyperprolactinemia and hyperphagia during lactation, which should have significant impact on both the postpartum metabolic health of breastfeeding moms and the nutrition supply for infants.
Progress Report
Connexin43 is a component of the gap junction, a specialized cellular structure that allows for intercellular communication between cells. Connexin43 plays an important role in adipocyte (fat cell) remodeling during lactation. In Objective 2, we proposed in mice to describe the effects of a dam's adipocyte Connexin43 gap junction ablation on her offspring's body weight, adipose tissue mass, and late-life responses to metabolic challenges such as overnutrition.
During the previous reporting period, we found that adipocyte Connexin43 knockout (Cx43 KO) dams appeared normal and did not develop obesity or impairment in handling exogenous glucose or lipids. However, when these animals produced offspring and lactated, the lactose levels in their breast milk were significantly lower than breast milk extracted from the control dams. Pups fostered by Cx43 KO dams weighed lower on Day 9, suggesting that breast milk from Cx43 KO dams was less efficient in supporting neonatal growth. This data identifies the vital role of adipocyte Cx43 in breast milk production, but a dispensable role of adipocyte Cx43 during basal state.
This year we followed and compared the body weight of pups fostered by control or Cx43 KO dams. Our results showed that neonates fostered by Cx43 KO dams continue to have a lower body weight at weaning than neonates fostered by control dams; however, their body weight caught up at 8 weeks of age to that of the control offspring. Their fat and lean mass were not different from their counterparts fostered by control dams. They continued to gain weight at a similar pace as their counterparts fostered by control dams; however, when challenged by a high-fat diet starting at 8 weeks old, they gained slightly more body weight, which was predominantly contributed by the fat mass. They also showed higher blood glucose levels after a glucose injection or pyruvate solution injection, but they maintained a similar level of insulin sensitivity and capability to eliminate orally administered lipids. These results suggest undernutrition in neonates fostered by Cx43 KO dams increased the risks of metabolic disorders, especially glucose intolerance, in offspring after metabolic challenge. Future research directions will call for investigating epigenetic modifications, including DNA methylation, miRNAs, and histone modification in key metabolic organs in this process.
Additionally, a new project was established due to the hiring of a new research scientist. The progress associated with this new project is as follows. During breastfeeding, female mammals eat a lot more food, which is associated with higher levels of the hormone prolactin and lower effects of estrogen when compared to a non-lactating status. We propose to determine whether this low estrogen effect causes a higher food intake and prolactin levels during lactation.
For Objective 3, this year we conducted two experiments to distinguish the effect of estrogen between peripheral organs and the brain. It has been well documented in previous studies that estrogen can act in the pituitary to increase prolactin levels. To find out whether estrogen inhibits prolactin levels through actions in the brain, we supplemented estrogen to lactating mice through either circulation or directly into the brain, respectively. From our pilot study, we found that only the brain estrogen supplement, but not systemic estrogen supplement, can inhibit prolactin levels. This result supports our hypothesis that estrogen acts on the brain to inhibit prolactin levels, which is opposite to the well-known estrogen effect in the pituitary to increase prolactin levels. We will continue to determine if the brain estrogen supplement can also reduce feeding of lactating dams and/or the survival of pups.
For Objective 5, we will supplement estrogen to the brain while artificially inhibiting this population of neurons in lactating mice. We expect that the inhibition of these neurons will block the effect of brain estrogen supplement. This year we performed a pilot study on a small cohort of female mice to test the surgery conditions, and we will continue to set up more mice to test the hypothesis.
Accomplishments
