Effects of cortisol or corticotropin administration on hepatic 3-hydroxy-3-methylglutaryl-CoA reductase activity and plasma lipids in the pregnant rat and fetuses

Pregnant rats were given pharmacological doses of cortisol or ACTH or no hormone from gestation day 9 to 19 and maternal and fetal hepatic 3-hydroxy-3-methylglutaryl-CoA reductase activity and plasma cholesterol studied on gestation day 20. Reductase activity was also studied in the maternal and fetal adrenal of the rats given cortisol or no hormone. Cortisol administration increased the maternal and fetal plasma cholesterol but had no effect on the hepatic active (phosphorylated) 3-hydroxy-3-methylglutaryl-CoA reductase activity when compared to untreated rats. Total (active + inactive) 3-hydroxy-3-methylglutaryl-CoA reductase activity, however, was reduced in maternal liver but not altered in the fetal liver by cortisol. The maternal cortisol treatment decreased the fetal, but not maternal, adrenal 3-hydroxy-3-methylglutaryl-CoA reductase total enzyme activity. The data support a hypothesis that utilization of plasma cholesterol for adrenal steroidogenesis may be an important determinant of plasma cholesterol homeostasis in the rat fetus. Maternal ACTH administration increased the foetal but not maternal plasma cholesterol, whilst active 3-hydroxy-3-methylglutaryl-CoA reductase activity was increased in the pregnant rat but not her fetuses. This result may suggest coordination of hepatic active reductase activity with adrenal cholesterol utilization in the pregnant rat. The reason for the fetal hypercholesterolaemia caused by ACTH, which is not known to cross the placenta, is uncertain. The studies, however, indicate that fetal cholesterol homeostasis and the rate limiting enzyme of cholesterol synthesis is influenced by maternal glucocorticoid administration.     

Effect on maternal and fetal renal function and plasma renin activity of a high salt intake by the ewe

The effect on renal function of replacing maternal drinking water with a solution containing 0.17 M NaCl was studied in 9 ewes and their chronically catheterised fetuses over a period of 9 days. Maternal sodium intake increased from control values of 2.19 +/- 0.09 mmol/h to 44.3 +/- 7.4 (P less than 0.001) and 46.3 +/- 6.5 mmol/h (P less than 0.001) on the 3rd and 6th days of salt ingestion. Maternal plasma sodium levels were not affected, but the urinary sodium/potassium ratio increased from 0.15 +/- 0.07 to 2.26 +/- 0.34 (P less than 0.001) after 6 days and plasma renin activity fell from 2.87 +/- 0.76 to 1.00 +/- 0.25 ng/ml per h (P less than 0.05). The changes in maternal sodium intake had no effect on fetal plasma sodium levels nor on fetal plasma renin activity. Sodium excretion and fetal urinary sodium/potassium ratio did not change. However, 3 days after the ewes returned to drinking water fetal plasma renin activity was significantly higher than it was prior to maternal ingestion of 0.17 M NaCl. Fetal plasma renin activity was inversely related to fetal plasma sodium levels (P less than 0.01). The results show that changes in maternal sodium intake had no long term effect on fetal plasma sodium levels nor on fetal renal sodium excretion. The fall in maternal plasma renin activity in the absence of any change in the fetal renin activity, indicates that the fetal renin angiotensin system is controlled by factors other than those influencing the maternal renin angiotensin system. Since fetal urinary sodium/potassium ratios remained unchanged it would suggest that fetal sodium excretion is not influenced by maternal levels of aldosterone.     

Do plasma ACTH and cortisol concentrations in the late-gestation fetal sheep vary diurnally?

This study tested the hypothesis that fetal plasma ACTH and cortisol concentrations vary diurnally, and the mean concentration and the amplitude of the rhythm vary as a function of fetal gestational age. Nine chronically-catheterized fetal sheep were studied between 120 and 142 days' gestation. All of the fetuses were born spontaneously and alive. The pregnant ewes were maintained in a room with a regular light cycle (on at 07.30, off at 17.30). Food and water were available ad libitum. Blood samples were drawn at 4-h intervals throughout a 24-h period. There were no significant daily variations in fetal plasma ACTH, cortisol, or progesterone concentrations, except in the last 3 days of fetal life. In these fetuses ACTH and cortisol concentrations were increased in the afternoon and evening. We conclude that there is no diurnal rhythm in ovine fetal hypothalamo-pituitary-adrenal axis activity, and that the increased plasma concentrations of ACTH and cortisol in the afternoon and evening hours of the last few days of fetal life might be a response to increased uterine contraction activity.     

25-Hydroxyvitamin D and 24,25-dihydroxyvitamin D in maternal plasma,fetal plasma and amniotic fluid in the rat

At the end of gestation plasma levels of 25-hydroxyvitamin D and 24,25-dihydroxyvitamin D were lower in pregnant than non pregnant female rats. In fetal plasma, concentrations of both metabolites were higher than in maternal plasma. This materno-fetal gradient led us to compare maternal and fetal plasma binding abilities. Fetal plasma was half as potent in binding 25-hydroxyvitamin D as maternal plasma. In fetal plasma binding was mainly due to the plasma vitamin D binding protein. On the other hand this study clearly showed that amniotic fluid contained 25-hydroxyvitamin D and 24,25-dihydroxyvitamin D. In addition this fluid was found to possess vitamin D-metabolite binding activity. The molecule responsible for this has been identified as the plasma vitamin D binding protein.     

Iron deficiency in the rat: biochemical studies of fetal metabolism

The effects of dietary-induced iron deficiency on fetal and maternal metabolism were studied in the rat. Concentrations of phenylalanine, but not tyrosine, were significantly elevated in plasma from iron-deficient maternal and fetal rats at day 20 of gestation with individual fetal plasma levels of phenylalanine as high as 10 mg per 100 ml. Concentrations of total 5-hydroxyindole compounds were significantly decreased in brain tissue from iron-deficient fetuses (day 20 of gestation), suggesting that synthesis of the compounds may be inhibited by iron deficiency. Mitochondrial NADH oxidase activity was markedly decreased (60%) in homogenates of fetuses at day 14 of gestation and may account for the high fetal resorption rate and small fetal size observed in the rat in iron deficiency.     

Fetal and maternal endocrine changes associated with parturition in the goat

In 4 goats maternal jugular plasma concentrations of progesterone started to decline before labor commenced, but levels remained elevated above basal concentrations throughout parturition. Fetal and maternal plasma concentrations of estrogen rose before and during labor, increasing markedly just prior to fetal delivery. Fetal carotid plasma glucocorticoid concentrations appeared to rise over the 5 days prior to parturition, but a significant increase was only noted during labor. Maternal plasma concentrations of glucocorticoids were significantly lower than fetal plasma concentrations of glucocorticoids. Estradiol-17α was the predominant estrogen in both fetal and maternal plasma, less estrone was detected and apparently little estradiol-17β was present.No good correlation was found between plasma progesterone or estrogen and parturient uterine activity; the latter lasted from 22 to 78 hours, with the most dramatic increase occurring close to delivery. It is probable that other agents are involved in controlling uterine activity during parturition in the goat.     

Nutritional control of respiratory and other muscular activities in relation to plasma prostaglandin E in the fetal sheep

The effects of nutrient availability on fetal plasma prostaglandin E (PGE) concentrations, on fetal breathing movements and electromyographic (EMG) activities of fetal nuchal and forelimb muscles were investigated in pregnant ewes by varying dietary intake and by manipulation of fetal plasma glucose concentration. The incidence of fetal breathing movements (06.00-10.00 h) decreased with increasing gestational age while fetal arterial concentrations of plasma PGE increased significantly over the same period of gestation. Maternal fasting for 48 h reduced the incidence of fetal breathing movements and the amount of nuchal EMG activity (06.00-10.00 h) in animals older than 130 days but had no effect earlier in gestation. No changes in forelimb EMG activity were observed during fasting at any gestational age. Plasma PGE levels increased significantly during fasts begun both before and after 130 days of gestation. When data from fed and fasted states were combined for all fetuses, irrespective of gestational age, there was a significant inverse correlation between fetal breathing movements incidence and plasma PGE concentration in utero. This relationship was even more pronounced when the fetuses were considered individually. Insulin infusions induced hypoglycaemia, an increase in fetal plasma PGE concentration and a significant reduction in the incidence of fetal breathing movements at all ages. Glucose infusions of fetal breathing movements only after 130 days and had no effect on plasma PGE levels in utero at any gestational age. Neither insulin nor glucose infusions altered the EMG activities of the nuchal and forelimb muscles. The results show that glucose availability is an important factor in determining the incidence of fetal breathing movements in utero and indicate that nutritionally induced changes in fetal breathing movements are mediated in part by PGE. They also suggest that PGE is a physiological regulator of fetal breathing movements in the sheep during late gestation.     

Studies on the growth of the fetal guinea pig. The effects of nutritional manipulation on prenatal growth and plasma somatomedin activity and insulin-like growth factor concentrations

In guinea pigs between days 41-46 of pregnancy prenatal growth has been manipulated by alteration of nutritional state. Three methods were used. Uterine artery ligation at day 30 of pregnancy depressed fetal growth rate by greater than 50% and was associated with falls in plasma insulin, IGF-1, cortisol, thyroid hormone, glucose, acetate and free fatty acid concentrations and rises in that of IGF-2, glucagon and amino acids. Fetal plasma was inhibitory to sulphate incorporation into pig costal cartilage. Complete food withdrawal from pregnant guinea pigs for 2 days at days 43-44 of pregnancy caused mild fetal growth retardation and similar changes in plasma constituents, except in that plasma IGF-2 concentrations were now depressed and plasma was not inhibitory to sulphate incorporation into pig costal cartilage. Production of hypoglycaemia by 4-times-daily maternal injections of glucose between days 41-46 of pregnancy accelerated fetal growth rate. It also elevated fetal plasma concentrations of insulin, IGF-1, IGF-2, sulphation-promoting activity, thyroid hormones, glucose and free fatty acids and depressed that of glucagon and amino acids. Fetal growth rate during the experimental period showed a good correlation with plasma glucose, insulin and IGF-1 and, to a certain extent, with sulphation-promoting activity. It did not correlate closely with fetal plasma IGF-2 concentration. Hepatic glycogen concentrations showed a good correlation with plasma IGF-2 levels.     

Androgen level in the sheep fetus during gestation.

The androgen content of amniotic fluid, plasma, and gonads from 107 fetal lambs was determined by radioimmunoassay in an attempt to understand the ontogeny of gonadal function. Testosterone (T) was too low to be reliably measured in the amniotic fluid from fetuses of either sex. Ovaries were without T activity at any of the stages of gestation studied. Testicular T-5alpha-dihydrotestosterone (T-DHT) concentration steadily decreased from 1.4/ml near term. It is suggested that nongonadal testosterone production increases during fetal life and that T secretion by the fetal testis may contribute steadily less to the plasma pool of T as gestation proceeds.     

The effects of pancreatectomy on the plasma concentrations of insulin-like growth factors 1 and 2 in the sheep fetus

The effects of hypoinsulinaemia and altered metabolite concentrations on the fetal plasma concentrations of insulin-like growth factors (IGF) have been investigated in chronically catheterized fetal sheep made insulin deficient by pancreatic ablation. Fetal pancreatectomy reduced significantly the plasma IGF-1 concentration and increased plasma IGF-2 activity in comparison with the values observed in sham operated fetuses. Mean plasma IGF-1 concentrations in the sham operated and pancreatectomized fetuses were 18.6 +/- 3.1 ng/ml (n = 7) and 13.4 +/- 1.4 ng/ml (n = 13) respectively. When all the data were combined, there was a significant positive correlation between the plasma concentrations of IGF-1 and insulin in utero. The mean IGF-2 activity was 2349 +/- 83 ng/ml (n = 7) in the sham operated fetuses and 3800 +/- 532 ng/ml in the pancreatectomized animals (n = 13). Plasma IGF-2 activity was correlated positively with plasma glucose, fructose and alpha-amino nitrogen levels and inversely related to the plasma insulin concentration in utero. These observations demonstrate that the fetal pancreas is essential for normal IGF production in the fetus and suggest that insulin, substrate availability and the IGFs may interact in the regulation of fetal growth.     

Insulin-like growth factors and binding proteins in the fetal rat: Alterations during maternal starvation and effects in fetal brain cell culture

Maternal malnutrition adversely affects fetal body and brain growth during late gestation. We utilized a fetal brain cell culture model to examine whether alternations in circulating factors may contribute to reduce brain growth during maternal starvation; we then used specific immunoassay and western blotting techniques, and purified peptides to investigate the potential role that altered levels of insulin-like growth factors (IGFs) and IGF binding proteins (IGFBPs) may play in impaired growth during maternal nutritional restriction.Fetal, body, liver, and brain weight were reduced after 72 hr maternal starvation, and plasma from starved fetuses were less potent than fed fetal plasma in stimulating brain cell growth. Circulating levels of IGF-I were reduced in starved compared to fed fetuses, while levels of IGF-II were similar in both groups. In contrast, [¹²⁵I]-IGF-I binding assay demonstrated an increase in the availability of plasma IGFBPs following starvation. Western ligand blotting and densitometry indicated that levels of 32 Kd IGFBPs were 2-fold higher in starved compared to fed fetal plasma. Immunoblotting and immunoprecipitation with antiserum against rat IGFBP-1 confirmed that heightened levels of immunoreactive IGFBP-1 accounted for the increase in 32 Kd IGFBPs in starved plasma. Levels of 34 Kd BPs, representing IGFBP-2, were unaffected by starvation. Reconstitution experiments in cell culture showed that IGF-I promoted fetal brain cell growth, and that when they were supplemented with IGF-I, the growth promoting activity of starved fetal plasma was restored to fed levels. These changes were measured using MTT to assess mitochondrial reductase activity. Conversely, addition of physiological amounts of rat IGFBP-1 inhibited the effects of fed fetal plasma on brain cell growth, and bioactivity was reduced even further with higher concentrations of IGFBP-1. Based on these results, we conclude that reciprocal changes in circulating levels of IGFBP-1 (increased) and IGF-I (decreased) may combine to reduce the availability of IGF-I to this tissue and limit fetal brain cell growth when maternal nutrition is impaired.     

Fetal sympathetic activity, transcutaneous PO2, and skin blood flow during repeated asphyxia in sheep

To improve detection of fetal distress, we examined whether increased fetal sympathetic activity during repeated episodes of asphyxia decreases skin blood flow, which can be monitored by recording transcutaneous PO2. Sympathetic activity was assessed by relating catecholamine concentrations in the fetal plasma to blood gas, acid-base, and heart rate variables which are commonly used to determine fetal distress. Fifteen experiments were conducted on 8 anaesthetised fetal sheep in utero between 125 and 145 days of gestation (term is at 147 days). They were subjected to 11 consecutive episodes of asphyxia of 30 (n = 3), 60 (n = 9), or 90 (n = 3) s over 33 min, achieved by arrest of uterine blood flow. Blood samples were drawn at 0, 33, and 60 min to determine arterial blood gases, acid base-balance, and concentrations of lactate, glucose, norepinephrine, and epinephrine. Fetal transcutaneous PO2, relative local skin blood flow, heart rate, arterial blood pressure, and arterial O2 saturation were recorded continuously. Fetal plasma concentrations of norepinephrine and epinephrine increased logarithmically as the duration of repeated asphyxia, anaerobic metabolism, and glucose concentrations increased, and as the mean O2 saturation, transcutaneous PO2, and local skin blood flow decreased. We conclude that during repeated episodes of asphyxia in fetal sheep near term, a significant increase in sympathetic activity can be detected indirectly by transcutaneous PO2 monitoring, because sympathetic activation reduces skin blood flow.     

Studies on the growth of the fetal guinea pig. Effects of reduction in uterine blood flow on the plasma sulphation-promoting activity and on the concentration of insulin-like growth factors-I and -II

The effects of prenatal growth restriction caused by uterine artery ligation at midgestation has been studied in pregnant guinea pigs. Ligation of a uterine artery at day 30 of pregnancy commonly caused a reduction in fetal growth of greater than 45% by days 40-65 of gestation. This was associated with substantial delays in the development of a number of fetal tissues and in particular that of the skeleton which remained cartilagenous for longer than normal. Hence normally by day 50 of pregnancy clear evidence of epiphyseal ossification in the long bones of the fore- and hindlimbs was present, but in growth retarded fetuses of less than 50% of normal size such evidence was sparce. Delayed skeletal development and the slowing of fetal growth rate correlated well with marked depression of plasma sulphation-promoting activity. Indeed plasma from fetuses that were less than 40% of normal size inhibited sulphate incorporation into pig costal cartilage. This indicated the presence of inhibitory factors in the plasma of such fetuses, an interpretation that was re-inforced by the observation that plasma IGF-II concentrations were 2-4 times above normal. In contrast plasma IGF-I concentration was depressed upto 50% by growth retardation in line with the fall in fetal plasma insulin concentration. The changes in plasma sulphation-promoting activity and of IGF-I are consistent with slowing of DNA, RNA and protein synthesis and of gene expression in tissues of the growth-retarded fetus. The elevated fetal plasma IGF-II concentration provided further evidence that in the fetal guinea pig this hormone has a potentially glyconeogenic action and maintains essential glycogen stores.     

High acyl-CoA cholesterol acyl transferase activity in fetal rabbit aorta: evidence for the presence of stimulating factor(s) in amniotic fluid

The activity of Acyl CoA-cholesterol acyl transferase was markedly high in fetal aortas when compared to maternal and adult male rabbits. This activity dropped by 50% at 1 week of age. This high activity in fetal aorta a) did not appear to be due to changes in plasma cholesterol levels or to the later development of endogenous inhibitor in the aorta, but rather b) due to stimulatory factor(s) present in amniotic fluid.     

Developmental changes in plasma erythroid colony-stimulating activity in mice: cyclic erythropoiesis associated with rapid growth.

To establish a role of erythropoietin (Epo) in regulation of fetal and neonatal erythropoiesis, plasma erythroid colony-stimulating activity (ECSA) in developing mice was measured by an erythroid colony-forming assay using fetal mouse liver cells. The ECSA in fetal and neonatal plasmas showed dose-response curves parallel to standard Epo curve and additive effects with standard Epo on the colony formation. Most of the plasma ECSA was neutralized by an anti-Epo monoclonal antibody. These results suggest that the plasma ECSA detected by the present bioassay is predominantly due to Epo. On day 12-14 of gestation, the plasma ECSA levels were at the highest values; thereafter the levels oscillated up to the age of 4 weeks. The packed cell volume (PCV) also oscillated, but with the reverse phase. Oscillation in PCV was associated with the growth. There was an inverse relationship between plasma ECSA and PCV levels throughout the prenatal and early postnatal periods. The results indicate that erythropoiesis in fetal and neonatal mice is regulated mainly on the basis of PCV-ECSA feedback control mechanism.     

Prevention of the normal expansion of maternal plasma volume: a model for chronic fetal hypoxaemia

The effects of inadequate expansion of maternal blood volume on uterine blood flow, fetal oxygen levels and vasoactive mediators during the third trimester were studied in 8 pregnant sheep. Results were compared to those obtained during 15 normal pregnancies. Prevention of the normal (20 ml/day) increase in maternal plasma volume was achieved by repeated haemorrhage and injections of furosemide. These treatments also reduced the rise in blood flow to the pregnant uterine horn that normally occurs during this period of gestation: at term flow was only 508 +/- 61 (SEM) compared to 838 +/- 83 ml/min in the control group (P greater than 0.01). This reduction in uterine blood flow caused a gradual fall in fetal PaO2, and rise in fetal levels of plasma renin activity, vasopressin, catecholamines and angiotensin II without change in pHa or base excess. Four to 5 days prior to delivery, the difference from control in PaO2 was -3.9 +/- 0.5 mmHg, plasma renin activity +2.9 +/- 1.7 ng/ml.h, vasopressin +4.2 +/- 1.1 pg/ml, catecholamines +957 +/- 145.3 pg/ml and angiotensin II +243 +/- 108.2 pg/ml. Furthermore, the fall in PaO2 and rise in vasoactive mediators that normally occur 3-5 days prior to the onset of labour was either absent (PaO2 and plasma renin activity) or blunted. Thus when expansion of blood volume during pregnancy is inadequate, blood flow to the uterus is adversely affected. This leads to various degrees of chronic fetal hypoxaemia and stimulation of vasoactive mediator systems. However, the normal stimulation of vasoactive mediator systems that occurs 3-5 days before delivery appears to be blunted. Experimental prevention of blood volume expansion during pregnancy produces an excellent model for the study of chronic mild fetal hypoxaemia.     

Functional anatomy of the ovaries of pregnant and lactating Cape porcupines, Hystrix africaeaustralis

The constituent cell types of the ovary of the porcupine were similar to those of New World hystricomorph rodents and accessory corpora lutea and luteal bodies were formed through the luteinization of the membrana granulosa or theca interna of antral follicles. All luteal bodies were histologically similar. The total volume of luteal tissue per female was not affected by fetal age and was unrelated to circulating concentrations of maternal plasma progesterone. Maternal plasma progesterone concentrations were correlated with fetal age. Follicular activity occurred throughout pregnancy but was not affected by fetal age or related to circulating values of oestradiol-17 beta. The formation of accessory corpora lutea during pregnancy is regarded as important in supplementing progesterone during pregnancy.     

Plasma triiodothyronines in fetal sheep: effects of illness and thyroidectomy.

Plasma 3,5,3'-triiodothyronine (T3) and 3,3',5'-triiodothyronine concentrations were measured in fetal sheep prior to death in utero and after thyroidectomy. In six fetal sheep who subsequently died in utero, plasma rT3 concentrations were elevated in all for 2 to 13 days prior to death. There were no consistent changes in plasma T4 concentrations. In two thyroidectomized fetal sheep, plasma T4 and rT3 concentrations fell to low levels. Plasma T3 concentrations remained low and there was no increase in plasma T3 in the last week prior to parturition like that which occurs in normal fetal sheep. Parturition was preceded by the normal increase in fetal plasma cortisol concentrations and occurred at the normal time. These data indicate that plasma rT3 concentrations are increased as a result of illness in fetal sheep and that such measurements may be useful as an indicator of fetal distress. The normal increase in plasma T3 late in gestation is not necessary for the late gestational cortisol surge or for normal parturition.     

Angiotensin-induced vasopressin release and activation of hypothalamic neuron in pre-term fetuses

Our previous studies have shown that central administration of angiotensin II (ANG II) causes vasopressin release in the near-term fetus in utero as evidence that the hypothalamic-neurohypophysial system has relatively matured before birth. However, it is still unknown whether the vasopressin controlling centers have been functionally developed in younger fetuses. This study determined fetal plasma vasopressin levels and hypothalamic vasopressin neuron activity in the chronically instrumented pre-term ovine fetuses. Introcerebroventricular (i.c.v.) administration of ANG II did not affect fetal plasma osmolality and sodium concentrations. However, fetal plasma vasopressin levels were significantly increased ( approximately 3-fold) in response to central injection of ANG II. Central ANG II also induced vasopressin-neuron activity marked with c-fos expression in the fetal hypothalamus at pre-term. In addition, the fetal organum vasculosum of the lamina terminalis and the subfornical organ were activated. The results suggest that hypothalamic-neurohypophysial system has been relatively intact and functional at 70% gestational age, and that central angiotensin is important in inducing fetal vasopressin release in utero.     

Hormonal factors in the control of heart rate in normoxaemic and hypoxaemic fetal, neonatal and adult sheep

The relationship of plasma levels of adrenaline, noradrenaline, arginine vasopressin (AVP) and plasma renin activity (PRA) to heart rate were studied in normoxaemic and hypoxaemic fetal, neonatal and adult sheep. The mean heart rate response of fetuses at the end of a 30 minute period of 10% oxygen delivery to the maternal ewe was tachycardia. However bradycardia, usually of a transient nature, was observed in 9 of the 12 fetuses (P less than 0.05). Multiple regression analysis was used to determine the contribution of blood gas, blood pressure and plasma hormone levels to the variance in heart rate in the perinatal sheep. 22% of the variance in fetal heart rate was provided by PRA and age from conception (P less than 0.001). Tachycardia was the invariable heart rate response of the neonates and adults to hypoxaemia. 61% of the variance in neonatal heart rate was contributed by PaO2, PaCO2, AVP, PRA and systolic blood pressure (SBP, P less than 0.001). PaO2 and plasma levels of adrenaline were significantly related to adult heart rate (P less than 0.001). Those fetuses which developed bradycardia had lower PaO2 but higher AVP and PRA during hypoxaemia than those which did not develop bradycardia. The major determinant of the area of the fetal bradycardia response was found, by multiple regression analysis, to be plasma adrenaline concentration (P less than 0.05). Thus different hormonal factors may play a role in the regulation of heart rate in normoxaemic and hypoxaemic fetal, neonatal and adult sheep.