Fetal baboon sex-specific outcomes in adipocyte differentiation at 0.9 gestation in response to moderate maternal nutrient reduction.

OBJECTIVE: To investigate in vitro adipocyte differentiation in baboon fetuses in response to reduced maternal nutrition. DESIGN: Cross-sectional comparison of adipocyte differentiation in normally grown fetuses and fetuses of pregnant baboons fed 70% of the control global diet from 30 days of pregnancy to term. SUBJECTS: The subjects comprised control (CTR) fetuses (five female and five male) of mothers fed ad libitum and fetuses of mothers fed 70% of the global diet consumed by CTR (maternal nutrient reduction (MNR), five female and five male fetuses). The expression of genes/proteins involved in adipogenesis (PPARγ, FABP4 and adiponectin) and brown adipose tissue development (UCP1, TBX15 and COXIV) were determined in in vitro-differentiated stromal-vascular cultures from subcutaneous abdominal, subcutaneous femoral and omental adipose tissue depots. Adipocyte number per area (mm(2)) was determined histologically to assist in the evaluation of adipocyte size. RESULTS: Maternal suboptimal nutrition suppressed growth of male but not female fetuses and led to adipocyte hypertrophy accompanied by increased markers of white- and, particularly, brown-type adipogenesis in male but not female fetuses. CONCLUSION: Adipose tissue responses to fetal nonhuman primate undernutrition are sexually dimorphic. While female fetuses adapt adequately, the male ones enhance pathways involved in white and brown adipose tissue development but are unable to compensate for a delayed development of adipose tissue associated with intrauterine growth restriction. These differences need to be considered when assessing developmental programming of adiposity in response to suboptimal maternal nutrition.

Upregulation of growth signaling and nutrient transporters in cotyledons of early to mid-gestational nutrient restricted ewes.

Multiparous ewes received 100% (control, C, n = 13) or 50% (nutrient restricted, NR, n = 14) of NRC dietary requirements from d28-d78 of gestation. On d78, 5 C and 6 NR ewes were necropsied. The remaining 8 C and 8 NR ewes were fed to 100% of NRC from d78-d135 and necropsied. Maternal blood was collected at both necropsies and at weekly intervals for assay of glucose, insulin and leptin. Fetal blood was collected at d78 and d135 necropsies for assay of glucose and lipids. Cotyledonary (COT) tissue was evaluated for protein and mRNA expression [fatty acid transporter (FATP)1, FATP4, CD36, glucose transporter (GLUT)1 and GLUT3], mRNA expression only [placenta fatty acid binding protein (FABPpm) and lipoprotein lipase (LPL)], or expression of phosphorylated and total protein forms [AMP kinase (AMPK)α, acetyl-CoA carboxylase (ACC), extracellular signal-regulated kinase (Erk)1/2, mammalian target of rapamycin (mTOR) and protein kinase B (Akt)]. On d78, but not d135, placental and fetal weights were reduced (P < 0.05) in NR vs. C ewes. Maternal circulating glucose, insulin and leptin levels were decreased in NR vs. C ewes on d78 (P < 0.05) but similar at d135. Fetal blood glucose and triglyceride levels were lower in NR vs. C ewes (P < 0.05) on d78, but similar on d135. On d78, GLUT1, FATP4, CD36 mRNA and protein expression levels, FABPpm mRNA level, and leptin protein level were all increased (P < 0.05) in COT of NR vs. C ewes. AMPK, ACC, and Erk1/2 activities were also increased (P < 0.05) in NR vs. C COT on d78. In contrast, only FATP4 was increased (P < 0.05) at both the mRNA and protein levels in COT of NR realimented vs. C ewes on d135. These data demonstrate placental adaptation to maternal NR through increasing nutrient transporter production and growth signaling activity.

Maternal obesity and increased nutrient intake before and during gestation in the ewe results in altered growth, adiposity, and glucose tolerance in adult offspring.

We evaluated the effects of preconception and gestational obesity in the ewe on offspring growth, metabolism, and glucose homeostasis. From 60 d before conception through parturition, multiparous ewes were fed 100% (control; n = 8) or 150% (obese, OB; n = 10) of NRC (1985) recommendations. Ewes on the OB diet increased BW by 30% from diet initiation to mating (P = 0.03) and by 52% by d 135 of gestation (P = 0.04), whereas control ewes increased BW by 7% (P = 0.65) from diet initiation to d 135 of gestation. Lambs were weaned at 120 d of age and were maintained as a group. At 19.5 ± 0.5 mo of age, offspring from control and OB ewes were individually penned and subjected to a 12-wk ad libitum feeding challenge. At the beginning and end of the feeding challenge, dual x-ray absorptiometry was used to determine percentage of body fat, and a frequently sampled intravenous glucose tolerance test (FSIGT) with minimal model analysis was used to assess insulin and glucose homeostasis. At the beginning of the feeding challenge, BW and percentage of body fat were similar for control and OB offspring, averaging 69.0 ± 1.5 kg and 5.3 ± 0.5%, respectively. At the initial FSIGT, glucose effectiveness and insulin sensitivity were reduced (P < 0.05) in offspring from OB compared with control ewes. During the feeding challenge, plasma concentrations of leptin were increased (P < 0.05) in offspring from OB compared with control ewes. Fasted plasma glucose before the feeding challenge tended to be greater (P = 0.06) in the OB offspring compared with the control offspring (83.3 ± 1.4 vs. 79.0 ± 1.6 mg/dL, respectively). At the end of the feeding challenge, fasted plasma glucose and insulin were increased (P < 0.05) in the OB offspring compared with the control offspring (84.0 ± 1.4 vs. 79.5 ± 1.5 mg/dL and 30.1 ± 2.1 vs. 23.4 ± 2.2 µIU/mL, respectively). During the feeding challenge, offspring from OB ewes consumed approximately 10% more feed (P < 0.05) and tended to have increased BW gain (approximately 14%; P = 0.08) compared with offspring from control ewes. At the final dual x-ray absorptiometry scan, percentage of body fat was greater (P < 0.05) for offspring from OB ewes than for offspring from control ewes (16.5 ± 1.2 vs. 10.8 ± 1.1%). At the final FSIGT, offspring from OB ewes had a decreased (P ≤ 0.05) acute insulin response to glucose, disposition index, and glucose effectiveness, and tended (P = 0.10) to have a decreased insulin sensitivity compared with offspring from control ewes. Maternal obesity induced before and during gestation leads to alterations in appetite, glucose and insulin regulation, and adiposity of mature offspring.

The effect of early to mid-gestational nutrient restriction on female offspring fertility and hypothalamic-pituitary-adrenal axis response to stress.

Primiparous ewes born as singletons to Rambouillet x Columbia crossbred ewes fed either 100% of NRC recommendations (control, Con; n = 7) or 50% of NRC (nutrient restricted, NR; n = 7) from d 28 through 78 postmating were utilized for this study. At 1 yr of age, a subset of ewes born to Con (n = 4) and NR (n = 4) mothers received jugular catheters and were subjected to a corticotrophin-releasing hormone (CRH)/arginine vasopressin (AVP) challenge, an ACTH challenge, and an isolation stress test, in which ACTH and cortisol responses were determined. A week after these challenges, estrus was monitored twice daily in all ewes from Con (n = 7) and NR mothers (n = 7). Once estrus was observed (d 0), daily blood samples were collected from ewes for progesterone through the subsequent estrus. Estrous detection and daily blood sampling were repeated during an estrous cycle in the next year, ewes were hand mated at the second estrus, and pregnancy was determined by delivery of a live lamb(s). Ewes from NR mothers tended (P = 0.10) to have a greater peak ACTH response after an intravenous CRH/AVP injection than ewes from Con mothers. The cortisol response of ewes to a CRH/AVP or ACTH challenge was not influenced by maternal nutrition. In contrast, ewes from Con mothers tended (P = 0.10) to release more ACTH in response to the isolation stress test and showed a greater (P = 0.04) cortisol release than ewes from NR mothers. Ewes from NR mothers exhibited decreased (P < 0.05) plasma progesterone in both yr 1 and 2 of the study compared with ewes from Con mothers. Furthermore, fewer (P < 0.0001) ewes from NR mothers produced a lamb (1 of 7) than ewes from Con mothers (7 of 7) during yr 2 of the study. These findings indicate that maternal undernutrition during early gestation may affect stress responses by the offspring, but has limited impact on hypothalamo-pituitary-adrenal sensitivity. Furthermore, offspring of NR ewes exhibited reduced progesterone secretion during the luteal phase of their estrous cycles and a markedly reduced fertility compared with offspring from Con ewes.

Down-regulation of growth signaling pathways linked to a reduced cotyledonary vascularity in placentomes of over-nourished, obese pregnant ewes.

Both protein kinase B (Akt) and extracellular signal-regulated kinase 1/2 (ERK1/2) are down-stream components of the insulin/insulin like growth factor-1 (IGF-1) signaling pathway. AMP-activated protein kinase (AMPK) is known to sensitize cells to insulin/IGF-1 signaling. The objective of this study was to assess the activity of AMPK and its role in the observed down-regulation of insulin/IGF-1 signaling in cotyledonary (COT) arteries supplying the placental component of the ewe placentome. Nonpregnant ewes were randomly assigned to a control (C, 100% of NRC recommendations) or obesogenic (OB, 150% of NRC) diet from 60 days before conception until necropsy on day 75 of gestation (n=5/group) or until lambing (n=5/group). At necropsy on day 75 of gestation, the smallest terminal arteries that entered the COT tissues (0.5-1.0 mm in diameter) were collected for analyses. Fetal weights were approximately 20% greater (P<0.05) on OB than C ewes, but birth weights of lambs were similar across dietary groups. Fetal plasma concentrations of glucose, insulin and IGF-1 were higher (P<0.05) in the blood of fetuses from OB than C ewes. Total AMPK and phosphorylated AMPK at Thr 172 (the active form) were reduced (P<0.05) by 19.7+/-8.4% and 25.9+/-7.7%, respectively in the COT arterial tissues of OB ewes. Total acetyl-CoA carboxylase (ACC), a down-stream target of AMPK, and its phosphorylated form were also reduced (P<0.05) by 32.9+/-9.2% and 45.4+/-14.6%, respectively. The phosphorylation of IRS-1 at Ser 789, a site phosphorylated by AMPK, was 24.5+/-9.0% lower (P<0.05) in COT arteries of OB than C ewes. No alteration in total insulin receptor, total IGF-1 receptor or their phosphorylated forms was observed, down-stream insulin signaling was down-regulated in COT arteries of OB ewes, which may have resulted in the observed decrease in COT vascular development in OB ewes.

Maternal undernutrition during early to mid-gestation in the ewe results in altered growth, adiposity, and glucose tolerance in male offspring.

This study utilized maternal undernutrition from early to midgestation in the ewe to determine the impact(s) of intrauterine growth restriction on postpartum growth of male offspring and the potential mechanisms involved. Multiparous ewes were fed 50% (nutrient-restricted) or 100% (control-fed) of their nutrient requirements (NRC, 1985) between d 28 and 78 of gestation, and then all ewes were fed 100% of the NRC requirements from d 79 through lambing. Male lambs born to nutrient-restricted (n = 9) and control-fed (n = 9) ewes exhibited similar BW (5.8 vs. 6.0 +/- 0.3 kg) and crown-rump lengths (53.8 vs. 55.4 +/- 1.0 cm) at birth. At 63 and 250 d of postnatal age, wether lambs were subjected to a glucose tolerance test, in which a bolus of glucose was administered i.v. to evaluate changes in glucose and insulin concentrations. After i.v. glucose administration at 63 d of age, lambs from nutrient-restricted ewes exhibited a greater area under the curve for glucose (AUCg; 6,281 vs. 5,242 +/- 429; P < 0.05) and insulin (AUCi; 21.0 vs. 8.6 +/- 1.9; P < 0.001) than lambs from control-fed ewes. After glucose administration at 250 d of age, lambs from nutrient-restricted ewes had greater AUCg (7,147 vs. 5,823 +/- 361; P < 0.01) but a lower AUCi (6.4 vs. 10.2 +/- 1.9; P = 0.05) than lambs from control-fed ewes. Lambs from nutrient-restricted ewes were heavier (26.6 vs. 21.8 +/- 2.3 kg; P < 0.05) and had more backfat (0.30 vs. 0.21 +/- 0.03 cm, P < 0.05) by 4 mo of age than the lambs from control-fed ewes. At slaughter at 280 d of age, lambs from nutrient-restricted ewes remained heavier than lambs from control-fed ewes, had greater (P < 0.05) amounts of kidney and pelvic-area adipose tissue, and tended (P < 0.10) to have reduced LM and semitendinosus muscle weights as a percentage of HCW. These data demonstrate that a bout of maternal undernutrition during early to midgestation in sheep increased BW and fat deposition during adolescence and dysregulated glucose uptake in the absence of any change in birth weight.

Placentomal differentiation may compensate for maternal nutrient restriction in ewes adapted to harsh range conditions.

Maternal nutrient restriction from early to midgestation can lead to fetal growth retardation, with long-term impacts on offspring growth, physiology, and metabolism. We hypothesized that ewes from flocks managed under markedly different environmental conditions and levels of nutrition might differ in their ability to protect their own fetus from a bout of maternal nutrient restriction. We utilized multiparous ewes of similar breeding, age, and parity from 2 flocks managed as 1) ewes adapted to a nomadic existence and year-long, limited nutrition near Baggs, WY (Baggs ewes), and 2) University of Wyoming ewes with a sedentary lifestyle and continuous provision of more than adequate nutrition (UW ewes). Groups of Baggs ewes and UW ewes were fed 50 (nutrient restricted) or 100% (control fed) of National Research Council recommendations from d 28 to 78 of gestation, then necropsied, and fetal and placental data were obtained. Although there was a marked decrease (P < 0.05) in fetal weight and blood glucose concentrations in nutrient-restricted vs. control fed UW ewes, there was no difference in these fetal measurements between nutrient-restricted and control-fed Baggs ewes. Nutrient-restricted and control-fed UW ewes exhibited predominantly type A placentomes on d 78, but there were fewer (P c0.05) type A and greater (P < 0.05) numbers of type B, C, and D placentomes in nutrient-restricted than control-fed Baggs ewes. Placental efficiency (fetal weight/placentomal weight) was reduced (P = 0.04) in d 78 nutrient-restricted UW ewes when compared with control-fed UW ewes. In contrast, nutrient-restricted and control-fed Baggs ewes exhibited similar placental efficiencies on d 78. This is the first report of different placental responses to a nutritional challenge during pregnancy when ewes were selected under different management systems. These data are consistent with the concept that Baggs ewes or their conceptuses, which were adapted to both harsh environments and limited nutrition, initiated conversion of type A placentomes to other placentomal types when subjected to an early to mid-gestational nutrient restriction, whereas this conversion failed to occur in UW ewes. This early placentomal conversion in the Baggs ewes may function to maintain normal nutrient delivery to their developing fetuses during maternal nutrient restriction.

Fetal fornix transection and gestation length in sheep.

Experiments in several species indicate that the hippocampus influences hypothalamo-pituitary-adrenal (HPA) axis function. In fetal sheep, simultaneous ACTH and cortisol rises over the last 30 days of gestation peak at term and are necessary for birth. We hypothesized that if the fetal hippocampal formation is functional in late gestation, loss of hippocampal input to the HPA axis following fetal fornix transection would change gestation length in comparison to controls. At 118-121 days of gestation (dG), stereotaxic technique was used in fetal sheep to sham transect (SHAM; n = 8) or transect (FXTX; n = 6) the dorsal fornix at the level of the hippocampal commissure. No differences were found between SHAM and FXTX fetuses in daily hormone profiles over the last week of gestation or in gestation length (148.0 +/- 1.2 vs. 149.0 +/- 0.4 dG, respectively). We conclude that the fetal hippocampus is immature in late gestation and we speculate that an immature hippocampus is necessary for the loss of negative feedback control that gives rise to the long term, simultaneous increases in ACTH and cortisol that are indispensable for labor and delivery at term in sheep.

Effect of 30 per cent maternal nutrient restriction from 0.16 to 0.5 gestation on fetal baboon kidney gene expression.

Previous studies in rodents and sheep show that maternal nutrient restriction during pregnancy alters fetal renal development. To date, no studies using fetal baboon RNA with human Affymetrix gene chips have been published. In the present study we have (1) evaluated the specificity of the Affymetrix human gene array 'Laboratory on a Chip' system for use with fetal baboon mRNA and (2) investigated the effects of moderate maternal global nutrient restriction (NR; 70% of ad libitum animals) from early (30 days gestation (dG)) to mid-gestation (90 dG; term = 184 dG) on the fetal baboon kidney. Morphometric and blood measurements were made on 12 non-pregnant baboons before they were bred. All baboons were fed ad libitum until 30 days pregnant, at which time six control baboons continued to feed ad libitum (control - C) while six received 70% of the C diet on a weight adjusted basis. Fetal kidneys were collected following caesarean section at 90 dG, with samples flash frozen and fixed for histological assessment. Fetal hip circumference was decreased in the NR group (68 +/- 2 versus 75 +/- 2 mm), while fetal body weight and all other measurements of fetal size were not different between C and NR at 90 dG. Maternal body weight was decreased in the NR group (12.16 +/- 0.34 versus 13.73 +/- 0.55 kg). Having established the specificity of the Affymetrix system for fetal baboon mRNA, gene expression profiling of fetal kidneys in the context of our maternal nutrient restriction protocol shows that NR resulted in a down-regulation of genes in pathways related to RNA, DNA and protein biosynthesis, metabolism and catabolism. In contrast, genes in cell signal transduction, communication and transport pathways were up-regulated in the NR group. These changes indicate that even a moderate level of maternal global NR impacts fetal renal gene pathways. Our histological assessment of renal structure indicates decreased tubule density within the cortex of NR kidneys compared with controls. The number of glomerular cross-sections per unit area were unaffected by NR, suggesting that tubule tortuosity and/or tubule length was decreased in the NR kidney. Taken together the changes indicate that NR results in accelerated fetal renal differentiation. The negative impact of poor maternal nutrition on the fetal kidney may therefore be in part due to shortening of critical phases of renal growth resulting in decreased functional capacity in later life. These findings may have important implications for postnatal renal function, thereby contributing to the observed increased predisposition to hypertension and renal disease in the offspring of nutrient restricted mothers.

Betamethasone in the last week of pregnancy causes fetal growth retardation but not adult hypertension in rats.

We tested the hypotheses that (1) maternal betamethasone (betaM) treatment over the range of clinical doses for prevention of prematurity-related pathologies from day 15 to 21 of rat gestation would produce growth retardation, and (2) the lowest betaM dose to produce growth retardation would result in hypertension in adult offspring. In experiment 1, pregnant Sprague-Dawley rats were administered betaM (0, 50, 100, 200, 400, or 600 microg/kg per day, subcutaneously) on days 15-21 of pregnancy and necropsied on day 21.5, with fetal lung and placental weights recorded. In experiment 2, two more groups of rats (0 or 100 microg/kg per day, subcutaneously) were allowed to deliver, and offspring were instrumented at 100 +/- 4 days of postnatal life with indwelling left carotid arterial catheters. After 48 hours of recovery, blood pressure was recorded continuously for 24 hours. In experiment 1, all newborn rats treated with betaM, and their placentas, except those receiving 50 microg/kg per day, were growth retarded in comparison with controls (P <.05). All treated lungs were smaller than those of controls (P <.05). In experiment 2, no differences were found in the mean arterial blood pressure of adult offspring given the lowest effective dose of betaM (100 microg/kg per day) compared with controls (114.2 +/- 5.3 mmHg versus 114.6 +/- 3.4 mmHg, respectively). These data suggest that glucocorticoids given in the last week of rat pregnancy in the lowest human clinical dose do not cause hypertension and somatic growth retardation. However, the presence of lung growth restriction at this dose argues for more studies on the efficacy of even lower concentrations for their ability to improve lung and other organ and tissue function while avoiding unwanted side effects.

Life before birth: effects of cortisol on future cardiovascular and metabolic function.

UNLABELLED: The concept of fetal programming is an area that is now under rigorous investigation in many laboratories throughout the world. We need to engender a fascination in all segments of society, not just pregnant women, about life in the womb. CONCLUSION: Everyone needs to understand that improving the condition of the fetus will have personal, social and economic benefits. The time has come to realize that, in a sense, it is not just women who are pregnant but it is the family and the whole of society.

Simple roost nests confer large energetic savings for sparrow-weavers.

White-browed sparrow-weavers (Plocepasser mahali, body mass 40 g) are group-living passerines adapted to the semi-arid environment of north-eastern and south-western Africa. During winter, the nocturnal ambient temperature of these regions often falls below 0 degrees C. imposing conditions demanding an increase in thermoregulatory heat production. Individuals roost throughout the year in inverted U-shaped roost nests. We investigated the energetic advantages of roosting by measuring nest and ambient temperatures in the field, as well as the resting metabolic rate (RMR) of the birds. The sparrow-weavers exhibited a wide thermoneutral zone (13 degrees C - 32 degrees C). Although RMR at thermoneutrality (40.2 J g.h(-1)) conforms with those of other passerines. the value at 0 degrees C (74.8 J g.h(-1)) is significantly lower than expected. The slope of the line below the lower critical temperature is unexpectedly steep, however, and appears to cause the physiological requirement for nest roosting due to a high cost of thermoregulation at low temperatures, perhaps due to shivering or non-shivering thermogenesis. The nest temperature at 0 degrees C ambient is 5 degrees C. resulting in a saving of some 7% in the energy spent during winter nights when food resources are in short supply compared with the rest of the year.

Intact osmoregulatory centers in the preterm ovine fetus: Fos induction after an osmotic challenge.

We previously demonstrated a functional systemic dipsogenic response in the near-term fetal sheep (128-130 days; 145 days = full-term) with swallowing activity stimulated in response to central and systemic hypertonic saline. Preterm fetal sheep (110-115 days) do not consistently demonstrate swallowing in response to hypertonic stimuli, and it is unclear whether this is due to immaturity of osmoreceptor mechanisms or neuronal pathways activating swallowing motor neurons. To determine whether osmoreceptive regions in the preterm fetus are activated by changes in plasma tonicity, we examined Fos expression with immunostaining in these neurons in response to an osmotic challenge. Nine preterm fetal sheep [five hypertonic saline-treated fetuses (Hyp) and four isotonic saline-treated fetuses (Iso)] were prepared with vascular and intraperitoneal catheters. Seventy-five minutes before tissue collection, hypertonic (1.5 M) or isotonic saline was infused (12 ml/kg) via an intraperitoneal catheter to fetuses. Brains were examined for patterns of neuronal activation (demonstrated by Fos protein expression). Hyp demonstrated increases in plasma osmolality (~10 mosmol/kg H(2)O) and Na concentrations (5 meq/l). Increased Fos expression was detected in Hyp in the organum vasculosum of the lamina terminalis (OVLT), subfornical organ (SFO), median preoptic nucleus (MnPO), supraoptic (SON), and paraventricular nuclei (PVN) compared with Iso animals. Neuronal activation within the OVLT, SFO, and MnPO indicates intact osmoregulatory mechanisms, whereas activation of the SON and PVN suggests intact fetal neural pathways to arginine vasopressin neurons. These results suggest that preterm fetal swallowing insensitivity to osmotic stimuli may be due to immaturity of integrated motor neuron pathways.

Ovine fetal swallowing: expression of preterm neurobehavioral rhythms.

OBJECTIVE: Fetal swallowing contributes importantly to amniotic fluid volume regulation and fetal gastrointestinal maturation. Near-term ovine fetal swallowing occurs in discrete bouts of activity (at approximately 30-min intervals) in association with fetal electrocortical voltage changes. Thus, swallowing rhythms have been hypothesized to be entrained to fetal neurobehavioral states. In the preterm ovine fetus, electrocortical activity does not demonstrate differentiation into high- and low-voltage periods until 120-130 days' gestation. We sought to quantify patterns of preterm (114 days, 0.75 gestation) ovine fetal swallowing activity and volume, and, in view of the lack of electrocortical pattern changes, to explore whether swallowing activity was regulated by an independent central pacemaker. METHODS: Six singleton ovine pregnancies were chronically prepared with fetal and maternal femoral artery and vein catheters. Biparietal electrocortical electrodes were placed on the fetal skull. Following a minimum 5-day recovery period, fetuses were studied at 114 +/- 1 days. Patterns of fetal swallowing behavior were quantified by computer analysis of laryngeal-esophageal electromyography (EMG) and thoracic esophageal fluid flow during a 12-h period. RESULTS: Esophageal fluid flow was bidirectional, although antegrade flow predominated, leading to an average fluid acquisition rate of 13 +/- 3 ml/h (7.3 +/- 1.8 ml/h per kg) during the 12-h study (302 +/- 87 ml/day). Propagated esophageal EMG activity, representing coordinated 'swallows', averaged 56 +/- 6 swallows/h and correlated well with net esophageal fluid flow. 'Bouts' of swallowing activity (> or = 3 swallows/min) averaged 9 +/- 1 swallows/bout, lasted 1.8 +/- 1.4 min and accounted for 31 +/- 4% of the swallowed volume. Despite the absence of fetal electrocortical high-voltage/low-voltage transitions, there was a 26.1 +/- 3.9-min interval between periods of swallowing bout activity. CONCLUSIONS: Preterm (0.75 gestation) ovine fetal volume swallowed (302 ml/day) and volume swallowed for body weight (175 ml/day per kg) was significantly less than that previously noted at 0.85 gestation (831 ml/day, 274 ml/day per kg, respectively; p < 0.05) although the rates of swallowing activity were similar. The presence of swallowing bout activity at periodic intervals, in the absence of electrocortical differentiation, suggests an intrinsic central pacemaker regulating preterm fetal neurobehavior.

Effect of gestational age, corticosteroids, and birth on expression of prostanoid EP receptor genes in lamb and baboon ductus arteriosus.

The aim of this study was to determine the effect of corticosteroids, gestational age, and birth on the expression of genes encoding prostanoid receptors in the lamb and baboon ductus arteriosus. The ductus arteriosus was obtained from 34 lambs and eight baboons, including chronically instrumented fetuses of both species exposed to either corticosteroid or vehicle. Expression of prostanoid receptor genes was quantified using Northern blot analysis relative to each of two housekeeping genes. Expression of both the EP3 and EP4 receptor genes was detected in lamb ductus and the level of expression of both genes was unaffected by corticosteroids. Expression of the EP4 receptor gene was lower in the ductus obtained from term lambs compared with preterm lambs and was lower still in neonatal animals, whereas no variation was observed in EP3 receptor gene expression. Expression of the EP4 receptor gene was also confirmed in fetal baboon ductus arteriosus, and maternal administration of corticosteroid did not reduce EP4 receptor gene expression in the baboon. We conclude that advancing gestational age and birth may inhibit prostaglandin E2-mediated relaxation of the ductus through a corticosteroid-independent reduction in EP4 receptor gene expression.

Central leptin stimulates ingestive behavior and urine flow in the near term ovine fetus.

Leptin inhibits ingestive behavior and induces diuresis and natriuresis. To examine whether leptin influences fetal physiologic functions, we investigated the effect of central leptin on ovine fetal swallowing activity and urine flow. Six pregnant ewes with singleton fetuses (130 +/- 2 d gestation) were prepared with maternal and fetal arterial and venous catheters, fetal lateral intra-ventricle cannula, fetal bladder and amniotic fluid catheters. Electromyogram wires were placed in the fetal thyrohyoid muscle and upper and lower nuchal esophagus and electrodes were implanted on the parietal dura. Five days after surgery, recombinant human leptin was infused into the lateral ventricle and the fetus monitored for 8 h. Central leptin increased fetal swallowing activity during low-voltage electrocortical activity from basal values (0.96 +/- 0.08 swallows/min) at 2 h (1.41 +/- 0.24 swallows/min), 4 h (2.81 +/- 0.57 swallows/min), 6 h (2.53 +/- 0.59 swallows/min) and 8 h (2.08 +/- 0.39 swallows/min, p < 0.05). In comparison to basal values, low voltage electrocortical activity decreased (57 +/- 5% to 42 +/- 4%) and high voltage electrocortical increased (43 +/- 5% to 61 +/- 4%). In response to leptin, fetal urine flow initially decreased from basal values at 2 h (0.12 +/- 0.03 to 0.08 +/- 0.02 ml/kg/min, p < 0.05) then subsequently increased at 4 h and 6 h (0.20 +/- 0.04; 0.21 +/- 0.04 ml/kg/min, respectively, p < 0.05). Central leptin significantly increases near term ovine fetal swallowing activity and urine output, suggesting that leptin contributes to in utero development of ingestive behavior.

Plasma osmolality dipsogenic thresholds and c-fos expression in the near-term ovine fetus.

In ovine and human pregnancy, fetal swallowing contributes importantly to amniotic fluid homeostasis. Fetal dipsogenic responsiveness to short-term plasma hyperosmolality develops in late gestation, although fetal swallowing is not stimulated in response to long-term plasma osmolality increases (2 to 3%), which typically stimulate adult drinking behavior. To explore the near-term fetal plasma osmolality threshold for swallowing stimulation, we examined the effects of i.v. hypertonic saline-induced subacute increases in plasma hypertonicity on fetal swallowing behavior. Central sites of activation were examined by c-fos expression in putative dipsogenic nuclei. The results demonstrate that subacute 2 to 3% plasma osmolality increases do not stimulate near-term ovine fetal swallowing. However, fetal swallowing activity significantly increased (3 times) after plasma osmolality increased >6% above basal values. Consistent with a specific dipsogenic response, i.v. hypertonic saline induced c-fos expression in the anterior third ventricle region, a putative dipsogenic center, as well as in the fetal hindbrain. The stimulation of fetal swallowing under conditions of higher osmotic stimulation and the correlation with forebrain c-fos expression indicates that near-term fetal osmoregulation mechanisms are functional, although not completely mature. Reduced fetal dipsogenic responsiveness may result from altered osmoreceptor sensitivity, downstream neuronal or synaptic immaturity, or potentially inhibitory actions of stimulated hindbrain nuclei.

Assessment of fetal scalp oxygen saturation determination in the sheep by transmission pulse oximetry.

OBJECTIVE: Electronic fetal heart rate monitoring has an unacceptable false-positive nonreassuring rate, which results in an excess of operative interventions. As a more objective measure of fetal oxygenation, fetal scalp pulse oximetry has been used to assess fetal blood oxygen saturation (SO (2)). The current devices use reflectance oximetry, which has inherent limitations. These include varying depths of signal penetration, variation with position, and potential for optical interference. In this study we evaluated a newly developed transmission pulse oximetry device consisting of transmitter and receiver diodes mounted within the coil of a standard scalp electrode (Spiral O(2)CTG; Respironics Inc, Marietta, Ga). STUDY DESIGN: Six pregnant ewes at 127 to 135 days' gestation (term, 145 days' gestation) were anesthetized, intubated, and prepared with a femoral artery catheter. Fetuses were prepared with brachial artery and jugular vein catheters. Maternal inspired oxygen fraction was titrated from 21% to 3%. Oximetry O(2)CTG devices were positioned on the fetal scalp, and recordings were compared with directly determined fetal arterial pH, PO (2), and SO (2) values. RESULTS: Maternal SaO (2) and PaO (2) ranged from 102% to 16% and 110 to 18 mm Hg, respectively. Fetal SaO (2) and PaO (2) ranged from 76% to 12% and 28 to 8 mm Hg, respectively. There was excellent correlation between direct fetal SaO (2) and scalp SO (2) (r (2) = 0.90; scalp SO (2) = 0.79 SaO (2) + 6.89). With an SaO (2) of <30% as the cutoff point for assessment of fetal compromise, scalp SO (2) measurements had a 94% +/- 10% specificity and a 94% +/- 10% positive predictive value. CONCLUSION: (1) Preliminary studies of the Spiral O(2)CTG sensor demonstrated high correlation of scalp SO (2) with fetal SaO (2). (2) Although potential inaccuracies remain, transmission oximetry may offer potential advantages in consistency, ease of application, and technology with respect to the current reflection oximeter devices.

Central neuropeptide Y stimulates ingestive behavior and increases urine output in the ovine fetus.

We hypothesized that central neuropeptide Y (NPY) increases swallowing activity and alters renal function in the near-term ovine fetus. Six ewes with singleton fetuses (130 +/- 2 days of gestation; 148 days = term) were chronically prepared with arterial and venous catheters, a fetal lateral cerebroventricular cannula, and fetal bladder and amniotic fluid catheters. For determination of fetal swallowing, electromyogram wires were placed in the fetal thyrohyoid muscle and the upper and lower nuchal esophagus. Electrodes were implanted on the parietal dura for determination of fetal electrocorticogram (ECoG). After 5 days of recovery, fetal swallowing, ECoG, blood pressure, and heart rate were monitored during a 3-h basal period. At t = 3 h, ovine NPY (0.05 mg/kg) was administered into the lateral ventricle, and fetuses were monitored for an additional 8 h. A control study of central administration of artificial cerebral spinal fluid was performed on an alternate day. Central NPY significantly increased swallowing activity during low-voltage ECoG from basal activity (1.26 +/- 0.15 swallows/min) at 4 h (1.93 +/- 0.37 swallows/min), 6 h (1.69 +/- 0.27 swallows/min), and 8 h (2.38 +/- 0.31 swallows/min). NPY significantly increased fetal urine flow (basal: 0.13 +/- 0.02; 4 h: 0.21 +/- 0.04; 6 h: 0. 19 +/- 0.03 ml.kg(-1).min(-1)). These results demonstrate that central NPY stimulates fetal swallowing activity and increases urine output, which may contribute to the in utero development of ingestive behavior.

Ovine hourly fetal urine production: relation to fetal electrocortical activity.

OBJECTIVE: Ultrasound studies of hourly urine production rate in human fetuses have suggested that a fall in urine production occurs in state 2F (fetal quiet sleep) secondary to a state-dependent decrease in renal blood flow. We sought to ascertain the relationship between fetal hourly urine production rate and behavioral state in the near-term ovine fetus, a model in which urine production and fetal brain activity can be directly measured. METHODS: Six ewes with singleton pregnancies were prepared with vascular and amniotic fluid catheters. Fetuses were prepared with hindlimb vascular catheters, a bladder catheter, and biparietal ECoG electrodes. After at least 5 days of recovery (ga 130 +/- 2 days; term = 145-150 days), each animal was monitored for a 6-h period. Urine production was measured by draining the bladder catheter through a drop counter and fetal ECoG was continuously recorded (sampling rate of 50 Hz). ECoG activity was analyzed using power spectral analysis and periods of active and quiet sleep identified using both signal amplitude and corresponding 85% spectral edge frequency. RESULTS: Basal fetal arterial pH (7.36 +/- 0.01), pO2 (22.0 +/- 1.2 mmHg) and pCO2 (47.0 +/- 1.6 mmHg) and plasma (295 +/- 2 mOsm/kg) and urine (179 +/- 3 mOsm/kg) osmolalities were within normal ranges. Active and quiet sleep comprised 50 +/- 2 and 43 +/- 1% time, respectively. There was no difference in hourly urine production rate in active sleep (21.4 +/- 9.7 ml/h) and quiet sleep (18.8 +/- 7.7 ml/h). CONCLUSIONS: 1) Hourly fetal urine production rate is independent of ECoG activity state in the near-term ovine fetus. 2) Assuming only minor species differences, ultrasound measurement of human fetal hourly urine production rate can be performed without concern for fetal neurobehavioral state changes.