Lactation In Rodents Research Articles

OR14-06 Serum Thyroid Hormones Increase At The Onset Of Lactation And Promote Metabolic Pathways Required For Human Milk Synthesis

Abstract Disclosure: H. Rostom: None. X. Meng: None. R. Humphrey: None. A. Fry: None. T. Elajnaf: None. F. Hannan: None. An increase in mammary metabolism is critical for initiating lactation during postpartum days 1-4. We utilised clinical and cellular approaches to investigate whether thyroid hormones, which promote lactation in rodents, are involved in initiating human lactation and regulating mammary metabolism. We recruited n=35 healthy pregnant women intending to breastfeed following informed consent and measured serum thyroid hormones (TSH and free T4) at 36 weeks’ gestation and on postpartum days 1-4 (1). Serum TSH significantly increased at post-partum day 1 compared with 36 weeks’ gestation (2.5±0.3 vs. 1.6±0.1 mU/L, p=0.001). This early postpartum rise in TSH was followed by a significant increase in free T4 on post-partum days 3 and 4 (11.7±0.2 vs. 10.5±0.2pmol/L at 36 weeks’ gestation, p<0.01). We hypothesised that thyroid hormones increase mammary metabolism to promote breast milk production. Reverse transcription-quantitative PCR (RT-qPCR) of genes encoding deiodinase enzymes (DIO1-3), which regulate intracellular thyroid hormone activity, was undertaken in lactating human mammary epithelial cells (HMECs) isolated from breast milk. This showed that DIO1, which converts T4 to T3, was expressed in lactating HMECs, whereas DIO2 and DIO3 were not detected. Moreover, RT-qPCR of the thyroid hormone receptor alpha and beta (THRA and THRB) genes in lactating HMECs showed that THRB has ∼4-fold greater expression than THRA (p<0.05, n=4). THRB phosphorylates Akt, which is required for initiating lactation. Consistent with this, cultured HMECs stimulated with 10nM T3, the most potent thyroid hormone, for 15min showed an ∼2-fold increase in Akt phosphorylation (p<0.0001, n=4). Akt regulates oxidative phosphorylation, which we assessed by measuring oxygen consumption rate (OCR) and ATP synthesis. HMECs treated with 10nM T3 for ≤8hrs showed a 2-fold increase in OCR (p<0.05, n=4) without any change in cellular ATP. These findings suggested that T3 uncoupled mitochondrial respiration from ATP synthesis. In support of this, RT-qPCR of cultured HMECs stimulated with 10nM T3 showed increased expression of genes encoding uncoupling proteins 2 and 3 (≥2-fold, p<0.01, n=4), which also function to divert substrates away from mitochondrial catabolism; and >20-fold increased expression of PPARGC1A (p<0.0001, n=4), which promotes mitochondrial biogenesis. In summary, these findings demonstrate that serum TSH and free T4 are increased at the onset of lactation. Moreover, our cellular studies indicate that thyroid hormones promote mammary mitochondrial biogenesis and may divert substrates used for ATP generation towards the synthesis of breast milk. Reference: (1) Rostom, H. et al. Protocol for an observational study investigating hormones triggering the onset of sustained lactation: the INSIGHT study. BMJ Open 12, e062478. Presentation: Friday, June 16, 2023

Behavioral effects on the offspring of rodent mothers exposed to Tetrahydrocannabinol (THC): A meta-analysis

Pre and perinatal administration of Tetrahydrocannabinol (THC) in rodents and their offspring has many effects that have been studied using different methods that have not been integrated using quantitative methods. The effect of THC administration on behavior can be better understood by meta-analytic techniques. We examined whether there is an overall effect on the behavior of the offspring when THC is administered to mothers. Eligibility criteria included experiments using an experimental design with a control group without THC, in which THC is administered to mothers during pregnancy and lactation in rodents, and in which at least one type of behavioral (locomotor, emotional or cognitive) measurement in the offspring was implemented. Cohen’s d was obtained for each study, then each individual study was weighted, and moderator analysis was performed. Analysis was performed using fixed and random effect models, and the heterogeneity was assessed by calculating Qb, I2 and the prediction interval. Furthermore, 3 sub-meta-analyses were carried out according to the type of behavior. The general analysis determined a low weighted effect size of THC on the behavior of the offspring, moderated by type of rat strain. The sub-meta-analyses showed a medium effect for cognitive effects of THC in the offspring, and a low effect on locomotor activity and emotional behavior. In addition, publication bias was not detected. More research is needed to contribute to the understanding of the effect of THC exposure on offspring.

Developmental origins of health and disease: experimental and human evidence of fetal programming for metabolic syndrome

The concept of developmental origins of health and disease has been defined as the process through which the environment encountered before birth, or in infancy, shapes the long-term control of tissue physiology and homeostasis. The evidence for programming derives from a large number of experimental and epidemiological observations. Several nutritional interventions during diverse phases of pregnancy and lactation in rodents are associated with fetal and neonatal programming for metabolic syndrome. In this paper, recent experimental models and human epidemiological studies providing evidence for the fetal programming associated with the development of metabolic syndrome and related diseases are revisited.

Bacterial Imprinting of the Neonatal Immune System: Lessons From Maternal Cells?

We examined the presence of a natural bacterial inoculum in breast milk and its intracellular transport from the maternal intestine to the breast through the circulation. Breast milk and peripheral blood were collected aseptically from healthy donors at various times after delivery, and the presence of viable bacteria was determined through plating. Temporal temperature gradient gel electrophoresis was used to examine the bacterial ribosomal DNA content in milk cells, maternal peripheral blood mononuclear cells, and feces and in corresponding infant feces. Blood from nongravid nonlactating women served as control samples. Bacterial translocation to extraintestinal tissues was also evaluated in virgin, pregnant, and lactating mice. Breast milk contained a low total concentration of microbes of <10(3) colony-forming units per mL. Temporal temperature gradient gel electrophoresis revealed that maternal blood and milk cells contained the genetic material of a greater biodiversity of enteric bacteria. Some bacterial signatures were common to infant feces and to samples of maternal origin. Bacterial translocation from the gut to mesenteric lymph nodes and mammary gland occurred during late pregnancy and lactation in mice. Bacterial translocation is a unique physiologic event, which is increased during pregnancy and lactation in rodents. Human breast milk cells contain a limited number of viable bacteria but a range of bacterial DNA signatures, as also found in maternal peripheral blood mononuclear cells. Those peripheral blood mononuclear cells showed greater biodiversity than did peripheral blood mononuclear cells from control women. Taken together, our results suggest that intestinally derived bacterial components are transported to the lactating breast within mononuclear cells. We speculate that this programs the neonatal immune system to recognize specific bacterial molecular patterns and to respond appropriately to pathogens and commensal organisms.

Passive immunization of lactating mice with stanniocalcin-1 antiserum reduces mammary gland development, milk fat content, and postnatal pup growth

During pregnancy and lactation in rodents, stanniocalcin-1 (STC-1) production by the ovaries is upregulated markedly and released into the circulation. The mammary glands are one target of this systemically delivered hormone. The purpose of this study was to lower serum levels of STC-1 in lactating mice through passive immunization so as to monitor the effects on mammary gland function and postnatal pup growth. Passive immunization significantly reduced circulating hormone levels, and pup growth was significantly compromised (30%), even though control and experimental litters had ingested equal amounts of milk. When mammary glands were analyzed, the alveolar area was significantly reduced in antibody-treated mothers. An analysis of milk composition revealed no changes in lactose, protein, or electrolyte levels but an approximately 40% reduction in triglyceride levels. The latter was due to a significant reduction in mammary gland lipoprotein lipase activity and led to a significant buildup of triglycerides in the serum. Body fat content was also significantly reduced in pups from antibody-treated mothers, whereas pup fecal fat content was increased. In mothers, passive immunization also caused significant behavioral effects, in particular, increased locomotor and hindleg rearing activities. Collectively, the results suggest that systemically derived STC-1 has important effects on mammary gland development and the transfer of serum-based triglycerides into milk. Locomotor effects suggest that STC-1 also has a role in maternal behavior.

Leptin and the Adaptations of Lactation in Rodents and Ruminants

Lactation markedly increases nutrient requirements in both rodents and ruminants. This is met mostly by increased food intake, but there are also adaptations to increase metabolic efficiency. Despite such changes, lactating animals usually experience periods of negative energy balance. This is not due to a physical constraint on food intake, at least in the rat. Leptin, a hormone secreted by adipocytes, plays an important role in the regulation of appetite and energy balance. During lactation, serum leptin concentration is decreased in both rodents and ruminants, and the nocturnal rise in concentration is lost in rats. Hypoleptinaemia in lactation is primarily a result of negative energy balance. There is also increased clearance of serum leptin, and the attenuation of the nocturnal rise in leptin in rats is at least partly due to the suckling stimulus. Hypoleptinaemia is not the major factor driving hyperphagia in lactating rats, but it probably facilitates the increased food intake. Leptin may play a more important role in this respect in lactating ruminants. Leptin is probably involved in other adaptations that increase metabolic efficiency during lactation. The ability of hypothalamic neuropeptides to respond to leptin does not appear to be altered by lactation in either rodents or ruminants. The reason why lactating animals do not respond to hypoleptinaemia with a further increase in appetite, thereby achieving energy balance, appears to be due to a failure to respond to changes in neuropeptides which mediate the effects of leptin.

Abstracts of Communications

Neuropeptide Y (NPY), a thirty-six amino acid peptide, is a potent centrally-acting appetite-stimulating agent which is believed to regulate eating behaviour and body weight (Williams et al. 1991). Lactation in rodents is associated with increased nutrient requirements which are met by extraordinary increases in food intake (Vernon, 1989). The aim of the present study was to investigate whether hyperphagia in lactating rats is associated with increased hypothalamic NPY levels, as in the case of other hyperphagic states such as starvation and diabetes. Twenty female Wistar rats, initially matched for age and weight, were studied. The experiment began when the lactating group (n 10) was on average at day 16 of lactation (range, 13-17). The lactating rats’ food intake was over 330% that of the controls (P 0.05). Four of the eight hypothalamic areas showed significantly increased NPY levels in the lactating rats compared with controls, namely: the arcuate nucleus-median eminence (+41%; P<0.001); the paraventricular nucleus (+35%; P<0.00l); the ventromedial nucleus (+66% ; P=0.003); the dorsomedial nucleus (+78%; P<0.00l). Other hypothalamic regions showed no differences between the two groups. Increased NPY levels in the arcuate nucleus (its principal hypothalamic site of synthesis) and in the paraventricular and dorsomedial nuclei (NPY-sensitive sites to which the arcuate nucleus projects) suggest increased activity of the NPY-ergic system in lactation. NPY may therefore mediate the markedly increased food intake in lactation.

Expression of Neuropeptide Y Precursor-Immunoreactivity in the Hypothalamic Dopaminergic Tubero-Infundibular System during Lactation in Rodents*

Evidence from physiological studies in rats shows that neuropeptide Y (NPY) has marked neuroendocrine effects on anterior pituitary function, and especially on LHRH and LH secretions. However, previous immunohistochemical studies in rats have revealed only scarce NPY-axons of medullary origin in the external zone of the hypothalamic median eminence, the common termination site of neuroendocrine adenohypophysiotropic systems. In view of this apparent contradiction, we used light microscopic immunohistochemistry to reassess the distribution of NPY in the hypothalamus of rodents of both sexes under physiological (estrous cycle in rats, pregnancy in rats, and lactation in both rats and mice) and experimental (gonadectomy in rats and adrenalectomy in both rats and mice) conditions with alterations of reproductive functions. We reasoned that such manipulations could induce changes in immunoreactivity in the NPY system involved in neuroendocrine regulation and would thus make it apparent to us. We show here that immunoreactivity for NPY and its carboxyterminal precursor-associated peptide are dramatically increased in the external median eminence of lactating female animals when compared to the other animal groups. This NYP-precursor-immunoreactivity is present, throughout lactation, in the tyrosine hydroxylase-immunoreactive (and therefore possibly dopaminergic) tubero-infundibular system. This immunoreactivity disappears rapidly from the median eminence after pup-removal. These observations suggest a role for NPY-precursor-derived peptides in the control of the suckling-induced PRL secretion and also demonstrate the chemical plasticity of the median eminence during a normal physiological event. Since in nonlactating animals and especially in normal cycling females NPY-precursor-immunoreactivity was detected in the system of medullary origin only, we conclude that, by exclusion, this system might be the one responsible for modulating gonadotropic secretion at the median eminence and/or pituitary levels.

Reproductive Efficiency and the Timing of Gestation and Lactation in Rodents

Reproductive Efficiency and the Timing of Gestation and Lactation in Rodents

Brown adipose tissue thermogenesis and the energetics of pregnancy and lactation in rodents.

There has been considerable recent interest in the concept that thermogenesis in brown adipose tissue (BAT) may play an important role in the regulation of energy balance, in addition t o its traditional role in the generation of heat for thermoregulatory purposes. This interest has stemmed primarily from studies on the metabolic responses t o voluntary overfeeding in rats given a ‘cafeteria’ diet, and from investigations on the efficiency of energy utilization of various types of obese animal, particularly genetically obese rodents (for reviews see Rothwell & Stock, 1983; Trayhurn, 1984). The main focus has been on providing an explanation for tlie causes of obesity, but consideration of the importance of BAT in whole-body energy metabolism has now extended to other situations. In particular, the activity of BAT during pregnancy and lactation in rodents is currently being investigated in relation t o the overall energetics of reproduction.