Prenatal cortisol exposure impairs adrenal function but not glucose metabolism in adult sheep.

Adverse environmental conditions before birth are known to programme adult metabolic and endocrine phenotypes in several species. However, whether increments in fetal cortisol concentrations of the magnitude commonly seen in these conditions can cause developmental programming remains unknown. Thus, this study investigated the outcome of physiological increases in fetal cortisol concentrations on glucose-insulin dynamics and pituitary-adrenal function in adult sheep. Compared with saline treatment, intravenous fetal cortisol infusion for 5 days in late gestation did not affect birthweight but increased lamb body weight at 1-2 weeks after birth. Adult glucose dynamics, insulin sensitivity and insulin secretion were unaffected by prenatal cortisol overexposure, assessed by glucose tolerance tests, hyperinsulinaemic-euglycaemic clamps and acute insulin administration. In contrast, prenatal cortisol infusion induced adrenal hypo-responsiveness in adulthood with significantly reduced cortisol responses to insulin-induced hypoglycaemia and exogenous adrenocorticotropic hormone (ACTH) administration relative to saline treatment. The area of adrenal cortex expressed as a percentage of the total cross-sectional area of the adult adrenal gland was also lower after prenatal cortisol than saline infusion. In adulthood, basal circulating ACTH but not cortisol concentrations were significantly higher in the cortisol than saline-treated group. The results show that cortisol overexposure before birth programmes pituitary-adrenal development with consequences for adult stress responses. Physiological variations in cortisol concentrations before birth may, therefore, have an important role in determining adult phenotypical diversity and adaptability to environmental challenges.

Glucocorticoid maturation of mitochondrial respiratory capacity in skeletal muscle before birth.

In adults, glucocorticoids act to match the supply and demand for energy during physiological challenges, partly through actions on tissue mitochondrial oxidative phosphorylation (OXPHOS) capacity. However, little is known about the role of the natural prepartum rise in fetal glucocorticoid concentrations in preparing tissues for the increased postnatal energy demands. This study examined the effect of manipulating cortisol concentrations in fetal sheep during late gestation on mitochondrial OXPHOS capacity of two skeletal muscles with different postnatal locomotive functions. Mitochondrial content, biogenesis markers, respiratory rates and expression of proteins and genes involved in the electron transfer system (ETS) and OXPHOS efficiency were measured in the biceps femoris (BF) and superficial digital flexor (SDF) of fetuses either infused with cortisol before the prepartum rise or adrenalectomised to prevent this increment. Cortisol infusion increased mitochondrial content, biogenesis markers, substrate-specific respiration rates and abundance of ETS complex I and adenine nucleotide translocator (ANT1) in a muscle-specific manner that was more pronounced in the SDF than BF. Adrenalectomy reduced mitochondrial content and expression of PGC1α and ANT1 in both muscles, and ETS complex IV abundance in the SDF near term. Uncoupling protein gene expression was unaffected by cortisol manipulations in both muscles. Gene expression of the myosin heavy chain isoform, MHCIIx, was increased by cortisol infusion and reduced by adrenalectomy in the BF alone. These findings show that cortisol has a muscle-specific role in prepartum maturation of mitochondrial OXPHOS capacity with important implications for the health of neonates born pre-term or after intrauterine glucocorticoid overexposure.

Effects of Maternal Obesity On Placental Phenotype.

The incidence of obesity is rising rapidly worldwide with the consequence that more women are entering pregnancy overweight or obese. This leads to an increased incidence of clinical complications during pregnancy and of poor obstetric outcomes. The offspring of obese pregnancies are often macrosomic at birth although there is also a subset of the progeny that are growth-restricted at term. Maternal obesity during pregnancy is also associated with cardiovascular, metabolic and endocrine dysfunction in the offspring later in life. As the interface between the mother and fetus, the placenta has a central role in programming intrauterine development and is known to adapt its phenotype in response to environmental conditions such as maternal undernutrition and hypoxia. However, less is known about placental function in the abnormal metabolic and endocrine environment associated with maternal obesity during pregnancy. This review discusses the placental consequences of maternal obesity induced either naturally or experimentally by increasing maternal nutritional intake and/or changing the dietary composition. It takes a comparative, multi-species approach and focusses on placental size, morphology, nutrient transport, metabolism and endocrine function during the later stages of obese pregnancy. It also examines the interventions that have been made during pregnancy in an attempt to alleviate the more adverse impacts of maternal obesity on placental phenotype. The review highlights the potential role of adaptations in placental phenotype as a contributory factor to the pregnancy complications and changes in fetal growth and development that are associated with maternal obesity.

Development and thyroid hormone dependence of skeletal muscle mitochondrial function towards birth.

KEY POINTS: Skeletal muscle energy requirements increase at birth but little is known regarding the development of mitochondria that provide most of the cellular energy as ATP. Thyroid hormones are known regulators of adult metabolism and are important in driving several aspects of fetal development, including muscle fibre differentiation. Mitochondrial density and the abundance of mitochondrial membrane proteins in skeletal muscle increased during late gestation. However, mitochondrial functional capacity, measured as oxygen consumption rate, increased primarily after birth. Fetal hypothyroidism resulted in significant reductions in mitochondrial function and density in skeletal muscle before birth. Mitochondrial function matures towards birth and is dependent on the presence of thyroid hormones, with potential implications for the health of pre-term and hypothyroid infants. ABSTRACT: Birth is a significant metabolic challenge with exposure to a pro-oxidant environment and the increased energy demands for neonatal survival. This study investigated the development of mitochondrial density and activity in ovine biceps femoris skeletal muscle during the perinatal period and examined the role of thyroid hormones in these processes. Muscle capacity for oxidative phosphorylation increased primarily after birth but was accompanied by prepartum increases in mitochondrial density and the abundance of electron transfer system (ETS) complexes I-IV and ATP-synthase as well as by neonatal upregulation of uncoupling proteins. This temporal disparity between prepartum maturation and neonatal upregulation of mitochondrial oxidative capacity may protect against oxidative stress associated with birth while ensuring energy availability to the neonate. Fetal thyroid hormone deficiency reduced oxidative phosphorylation and prevented the prepartum upregulation of mitochondrial density and ETS proteins in fetal skeletal muscle. Overall, the data show that mitochondrial function matures over the perinatal period and is dependent on thyroid hormones, with potential consequences for neonatal viability and adult metabolic health.

Ascorbate prevents placental oxidative stress and enhances birth weight in hypoxic pregnancy in rats.

This study isolated the effects of maternal hypoxia independent of changes in maternal nutrition on maternal circulatory and placental molecular indices of oxidative stress and determined whether maternal antioxidant treatment conferred protection. Pregnant rats were subjected to normoxic pregnancy or 13% O2 chronic hypoxia for most of gestation with and without maternal treatment with vitamin C in the drinking water. Maternal hypoxia with and without vitamin C did not affect maternal food or water intake and led to a significant increase in maternal and fetal haematocrit. At gestational day 20, maternal plasma urate and L-cysteine concentrations, and placental levels of 4-hydroxynonenal and heat shock protein 70 were increased while placental heat shock protein 90 levels were decreased in hypoxic pregnancy. The induction of maternal circulatory and placental molecular indices of oxidative stress in hypoxic pregnancies was prevented by maternal treatment with vitamin C. Maternal hypoxia during pregnancy with or without vitamin C increased placental weight, but not total or compartmental volumes. Maternal treatment with vitamin C increased birth weight in both hypoxic and normoxic pregnancies. The data show that maternal hypoxia independent of maternal undernutrition promotes maternal and placental indices of oxidative stress, effects that can be prevented by maternal treatment with vitamin C in hypoxic pregnancy. While vitamin C may not be the ideal candidate of choice for therapy in pregnant women, and taking into consideration differences in ascorbic acid metabolism between rats and humans, the data do underlie that antioxidant treatment may provide a useful intervention to improve placental function and protect fetal growth in pregnancy complicated by fetal hypoxia.

Prenatal hypoxia independent of undernutrition promotes molecular markers of insulin resistance in adult offspring.

Molecular mechanisms predisposing people to insulin resistance are starting to emerge. Altered insulin signaling for hepatic gluconeogenesis and muscle glucose uptake is thought to play a central role. Development under suboptimal conditions is also known to increase the risk of insulin resistance in adulthood. However, the partial contributions of reduced oxygen vs. nutrient delivery to the fetus, two common adverse conditions in utero, to developmental programming of insulin resistance remain unknown. The aim of this study was to determine the effects of developmental hypoxia or undernutrition on the expression of insulin-signaling proteins in liver and skeletal muscle in adult rat offspring. We show that the expression of hepatic phospho-Akt and muscle Akt2 were significantly reduced in offspring of hypoxic, relative to offspring from normoxic or undernourished, pregnancies. Hepatic Akt-1, Akt-2, and PKCζ protein expression was reduced in offspring from both hypoxic and undernourished pregnancies. Muscle GLUT4 expression was decreased in undernourished, and further decreased in hypoxic, offspring. These findings link prenatal hypoxia to down-regulation of components of hepatic and muscle Akt expression in adult offspring. Akt may represent a pharmaceutical target for clinical intervention against the developmental programming of metabolic disease resulting from prenatal hypoxia.

Cardiac and vascular disease prior to hatching in chick embryos incubated at high altitude.

The partial contributions of reductions in fetal nutrition and oxygenation to slow fetal growth and a developmental origin of cardiovascular disease remain unclear. By combining high altitude with the chick embryo model, we have previously isolated the direct effects of high-altitude hypoxia on growth. This study isolated the direct effects of high-altitude hypoxia on cardiovascular development. Fertilized eggs from sea-level or high-altitude hens were incubated at sea level or high altitude. Fertilized eggs from sea-level hens were also incubated at high altitude with oxygen supplementation. High altitude promoted embryonic growth restriction, cardiomegaly and aortic wall thickening, effects which could be prevented by incubating eggs from high-altitude hens at sea level or by incubating eggs from sea-level hens at high altitude with oxygen supplementation. Embryos from high-altitude hens showed reduced effects of altitude incubation on growth restriction but not on cardiovascular remodeling. The data show that: (1) high-altitude hypoxia promotes embryonic cardiac and vascular disease already evident prior to hatching and that this is associated with growth restriction; (2) the effects can be prevented by increased oxygenation; and (3) the effects are different in embryos from sea-level or high-altitude hens.

The role of catecholamines in memory impairment in chicks following reduced gas exchange in ovo.

We have shown previously that reducing gas exchange to chick embryos by half wrapping eggs with an impermeable membrane from either days 14-18 (W14-18) or days 10-18 (W10-18) of the 21 day incubation results in post-hatch memory deficits. In the W10-18 chicks, short-term memory following training is impaired, whereas in the W14-18 chicks, memory is intact for 30 min but does not consolidate into long-term storage. The reduction in gas exchange caused by half wrapping eggs resulted in alterations in hematocrit, O2 and CO2 tensions suggesting that the embryos are hypoxic and hypercapnic. Our aim was to test the hypothesis that increases in circulating levels of catecholamines in ovo, as a result of hypoxia, lead to a disturbance of the central noradrenergic pathways resulting in cognitive impairment. Noradrenaline is critical for memory consolidation and a disturbance during development could compromise cognitive ability. In the present study, plasma noradrenaline levels were significantly elevated compared with control levels 2 days after hatch in W14-18 chicks. There was also a decrease in tissue noradrenaline concentration in the anterior forebrain in both W14-18 and W10-18 chicks. The differential ability of centrally administered beta2- and beta3-adrenoceptor agonists to overcome the memory deficit post-training, suggests altered responsiveness of central beta2-adrenoceptors to noradrenaline in W14-18 chicks. By comparing the W10-18 and W14-18 chicks with those from eggs wrapped from W10-14 we show that it is the timing of the prenatal hypoxia, rather than its duration, that determines the nature of cognitive dysfunction. We conclude that prenatal hypoxia induced by restriction of gas exchange can disrupt or alter central noradrenergic transmission causing cognitive impairment.

Restriction of prenatal gas exchange impairs memory consolidation in the chick.

Our aim was to assess the effects of restricting gas exchange during incubation on postnatal memory formation and growth in the chick. Gas exchange across the eggshell was restricted by covering 50% of the eggshell with an impermeable membrane for 4 or 8 days, commencing at days 14 and 10, respectively, of a 21-day incubation. Memory formation was examined postnatally at 1-2 days using a one-trial discriminated bead task, and at 5-6 days using a discriminated wheat task. For both tasks, chicks from eggs wrapped from days 14 to 18 had impaired memory retention at 60 min after training, although learning and labile memory were not impaired. Chicks from eggs wrapped from days 10-18 appeared to be poorer in their ability to form memories, and did not discriminate as well as controls in any of the tasks. Body weights of chicks from wrapped eggs were reduced from 2 days after hatching; chicks from eggs wrapped from day 10 had lower body weights at hatching. We conclude that a period of altered prenatal gas exchange can impair memory consolidation in the chick soon after hatching. The ability to form memories may be permanently altered, as this impairment is still apparent at 5-6 days after hatching. Pre- and postnatal growth was also impaired in the chicks from wrapped eggs. Our results suggest that the extent to which postnatal neurological function and growth is impaired depends on the timing and possibly the duration of the prenatal insult.

Postnatal outcomes in term and preterm lambs following fetal growth restriction.

1. Epidemiological evidence indicates that low birthweight increases the risk of a number of adult-onset diseases. It is now apparent that many babies with a low birthweight may have been subjected to a combination of reduced growth rates in utero as well as preterm birth. However, the long-term effects of preterm birth following intra-uterine growth restriction (IUGR) are unknown. Thus, our objectives were: (i) to identify prenatal factors associated with preterm birth in IUGR fetuses; and (ii) to characterize postnatal effects of preterm birth following IUGR. 2. We studied pregnant sheep and their offspring, in which fetal growth was restricted by umbilico-placental embolization during late gestation. Some of these animals were born at term (146 +/- 1 days) and some were born prematurely (139 +/- 1 days). In both groups, we have conducted longitudinal studies of postnatal respiratory function, cardiovascular function and learning ability up to 6-8 weeks of age. 3. Before birth, IUGR fetuses born prematurely (P-IUGR) were more hypoxaemic and acidaemic and had higher haemoglobin concentrations than both control fetuses and IUGR fetuses born at term (T-IUGR). In P-IUGR fetuses, plasma cortisol concentrations increased earlier than in the two other groups. The P-IUGR lambs had lower birthweights than T-IUGR lambs and both groups of IUGR lambs remained lighter than controls for 8 weeks. 4. After birth, P-IUGR lambs were hypoxaemic compared with T-IUGR and control lambs. Pulmonary diffusing capacity (adjusted for lung volume) was significantly lower in both groups of IUGR lambs than in controls, with P-IUGR lambs having lower values than T-IUGR lambs. Lung compliance (adjusted for lung volume), was not different between P-IUGR and control lambs, but values were higher in T-IUGR lambs than in control and P-IUGR lambs. Chest wall compliance (adjusted for lung volume) was higher in both groups of IUGR lambs than in controls. 5. During the 8 week postnatal study period, both groups of IUGR lambs had lower mean arterial pressures than control lambs; this relative hypotension was greatest in P-IUGR lambs. 6. In tests of learning ability, P-IUGR lambs took longer to complete a simple maze task at all ages and, in the second postnatal week, made a greater number of errors compared with controls. In an obstacle course, P-IUGR lambs recorded longer trial durations; they also made more errors than control lambs. 7. We conclude that preterm birth in the presence of late- gestational placental insufficiency and IUGR can result in specific effects on respiratory and cardiovascular development after birth, in addition to the effects of IUGR alone.

Assessment of learning ability and behaviour in low birthweight lambs following intrauterine growth restriction.

The present study used behavioural tasks to assess learning ability and behaviour in postnatal lambs, and to examine the effects of low birthweight (LBW) and age on subsequent performance. It was hypothesized that intrauterine growth restriction (IUGR) and LBW lead to learning and behavioural deficits in the early postnatal period. IUGR and LBW were induced by umbilico-placental embolization from 120 days of gestational age (g.a.) to the onset of labour. Behavioural studies were performed on 6 LBW and 6 control lambs between 2 and 6 weeks after birth. LBW lambs were born at 139+/-1 days g.a. (2.4+/-0.2 kg) and control lambs were born at 149+/-1 days g.a. (4.5+/-0.4 kg). Three tasks were used to assess the learning ability and behaviour of the lambs: a simple maze, an obstacle course, and a T-maze. LBW lambs took longer to complete the simple maze at all ages, and made a greater number of errors at Week 1 of testing compared to control lambs; the total trial duration and number of errors decreased with age for both groups. In the obstacle course, the times taken to complete the first and third trials were used for analysis; a decrease in trial time and the number of errors from Trial 1 to Trial 3 were indications of the lamb's ability to learn how to negotiate the objects within the course. LBW lambs recorded longer trial durations for the first trial at Week 5 of testing, and for the third trial at Week 4. LBW lambs made more errors for the first trial at Week 5 of testing than control lambs. In the T-maze, there was no significant effect of treatment or age. It was concluded that differences between the groups may have been the result of LBW lambs being prematurely born. The value of these tasks in the assessment of learning ability and behaviour in young lambs is discussed.