Postnatal profiles of glycogenolysis and gluconeogenesis are modified in rat pups by maternal dietary glucose restriction.

Because glucose is an important metabolic fuel during perinatal development, the effect of restriction of maternal dietary glucose on the developmental profile of neonatal glucoregulatory pathways was investigated. Pregnant rats were fed isoenergetic diets (0, 12, 24 or 60% glucose) and offspring were killed at seven postpartum time periods: 0-2, 4-6, 12-16 and 24 h, and 3, 6 and 15 d. Failure of the most restricted pups (0%) to survive 24 h was explained by persistent hypoglycemia resulting from the following: 1) insufficient tissue glycogen reserves at birth; 2) lower liver glycogen mobilization; 3) delayed phosphorylase a induction; and 4) low phosphoenolpyruvate carboxykinase (PEPCK) gene expression, all of which occurred despite the lower insulin:glucagon ratio. Differences in liver glycogen stores, which had been exhausted in all dietary groups by 16 h, could not account for the high d 1 pup mortality in the moderately restricted (12 and 24% glucose) groups. However, a certain metabolic distress was suggested because these moderately restricted neonates had significantly higher liver PEPCK gene expression at 12-16 h but significantly lower plasma glucose at 24 h. The high d 3 mortality, confirmed by analysis of deviance, was not supported by significant differences in any of the measured glucoregulatory indices. We conclude that dietary glucose during pregnancy is required for neonatal survival; its restriction not only lowers tissue glycogen reserves, but can disrupt the normal gene expression of liver PEPCK and the neonatal profile of phosphorylase a activity. Importantly, these observations show that the development of neonatal glucoregulatory mechanisms is modified by the availability of maternal dietary glucose.

Assessing the effects of low boron diets on embryonic and fetal development in rodents using in vitro and in vivo model systems.

To date, boron (B) essentiality has not been conclusively shown in mammals. This article summarizes the results of a series of in vitro and in vivo experiments designed to investigate the role of B in mammalian reproduction. In the first study, rat dams were fed either a low (0.04 microg B/g) or an adequate (2.00 microg B/g) B diet for 6 wk before breeding and through pregnancy; reproductive outcome was monitored on gestation day 20. Although low dietary B significantly lowered maternal blood, liver, and bone B concentrations, it had no marked effects on fetal growth or development. The goal of the second study was to assess the effects of B on the in vitro development of rat postimplantation embryos. Day 10 embryos collected from dams fed either the low or adequate B diets for at least 12 wk were cultured in serum collected from male rats exposed to one of the two dietary B treatments. Dams fed the low B diet had a significantly reduced number of implantation sites compared to dams fed the B-adequate diet. However, embryonic growth in vitro was not affected by B treatment. The aim of study 3 was to define the limits of boric acid (BA) toxicity on mouse preimplantation development in vitro. Two-cell mouse embryos were cultured in media containing graded levels of BA (from 6 to 10,000 microM). Impaired embryonic differentiation and proliferation were observed only when embryos were exposed to high levels of BA (>2000 microM), reflecting a very low level of toxicity of BA on early mouse embryonic development. Study 4 tested the effects of low (0.04 microg B/g) and adequate (2.00 microg B/g) dietary B on the in vitro development of mouse preimplantation embryos. Two-cell embryos obtained from the dams were cultured in vitro for 72 h. Maternal exposure to the low B diet for 10, 12, and 16 wk was associated with a reduction in blastocyst formation, a reduction in blastocyst cell number, and an increased number of degenerates. Collectively, these studies support the concept that B deficiency impairs early embryonic development in rodents.

Pathogenesis of malformations in a rodent model for Smith-Lemli-Opitz syndrome.

The fact that Smith-Lemli-Opitz syndrome (SLOS), a syndrome comprising major malformations involving a number of organ systems, results from an abnormality in cholesterol biosynthesis, was discovered only recently. Utilizing a drug (BM 15.766) to inhibit the same step in cholesterol biosynthesis as is abnormal in those affected with SLOS, we have developed a rat model that presents with abnormalities observed as early as gestational day 12 that appear to be consistent with some of those subsequent malformations that comprise the human syndrome. Abnormalities of the brain and face include deficiency in the midline region of the upper face, narrowing of the forebrain hemispheres and of the cerebral aqueduct, and deficiency in the developing lower jaw. Associated pathogenesis, as observed on gestational day 11 in histological sections and with scanning electron microscopy, involves abnormal cell populations at the rim of the developing forebrain and in the alar plate of the lower midbrain and hind-brain. The affected cells appear abnormally rounded up, having apparently lost their normal cell contacts. The potential basis for the selective vulnerability of this cell population and the impact of its vulnerability relative to subsequent dysmorphogenesis is discussed.

Limb, genital, CNS, and facial malformations result from gene/environment-induced cholesterol deficiency: further evidence for a link to sonic hedgehog.

Low cholesterol levels produced by treating cholesterol deficient mutant mice with a cholesterol synthesis inhibitor (BM 15.766) between days 4 to 7 of pregnancy resulted in malformations consistent with those in the Smith-Lemli-Opitz syndrome (SLOS). Facial anomalies in mildly affected gestational day 12 mouse embryos included a small nose and long upper lip; in more severely affected embryos, the facial and forebrain anomalies are representative of holoprosencephaly. Additionally, abnormalities of the mid- and hind-brain were observed and included stenosis of the cerebral aqueduct at the level of the isthmus and apparent absence of the organ progenitor for the cerebellar vermis. Although not previously directly linked to cholesterol deficiency in experimental animals, limb and external genital defects were a notable outcome in this multifactorially-based cholesterol deficiency model. The results of this study provide new evidence supporting an important role for cholesterol in early embryonic development, provide additional support for the hypothesis that this role may involve the function of specific gene products, such as sonic hedgehog (shh) signaling protein, and provide a description of the pathogenesis of some of the characteristic malformations in SLOS.

Maternal dietary carbohydrate restriction influences the developmental profile of postnatal rat brain indoleamine metabolism.

Dietary glucose restriction during pregnancy can retard fetal brain development, lower term brain glycogen levels and adversely affect the serotonergic neurotransmitter system in the fetus. To study if the postnatal profile of brain indoles continues to respond to these diet-induced changes, pregnant rats were fed graded levels (0, 12, 24, 60%) of glucose from impregnation to day 15 postpartum, and neonatal brain measurements were made. A steady decrease in tryptophan levels, a steady increase in 5-hydroxytryptamine (5-HT) levels and a U-shaped change in 5-hydroxyindoleacetic acid (5-HIAA) were observed during the first 15 postpartum days. Superimposed on these development profiles was a temporary surge in the concentrations of all three indoles 24 h after birth, which was dramatic for tryptophan and more modest for 5-HT and 5-HIAA. The level of carbohydrate in the maternal diet significantly influenced the magnitude of this increase in tryptophan, 5-HT and 5-HIAA at 24 h: the values were significantly higher in the carbohydrate-restricted (12 or 24%) rat pups when compared with control or carbohydrate-free (0% glucose) offspring. No effects of dietary treatment were apparent by day 6. However, the reemergence of a significant difference in brain 5-HT content at day 15 postpartum indicates that even when energy intake is adequate the level of carbohydrate in the maternal diet may continue to play a role in modulating serotonergic neurotransmitter levels later in development.

Glucose-restricted diets alter milk composition and mammary gland development in lactating rat dams.

To examine the effects of chronic restriction of maternal dietary glucose on lactational performance, pregnant dams were fed one of four isoenergetic diets containing graded levels of glucose (0, 12, 24 and 60%) from d 2 of pregnancy to d 15 of lactation. Dams fed the 0% glucose diet produced colostrum with higher protein and lower lactose and fat concentrations than normal, but all pups born to these dams failed to survive more than 24 h postpartum. Dams fed glucose-restricted diets (12 and 24%) had significantly lower d 15 milk fat concentration than dams fed the control diet, but there were no differences in milk protein, lactose and glucose concentrations. On d 15, pups suckling dams fed the 12% glucose diet had significantly lower body weights than pups of dams fed 24 and 60% glucose diets. Restriction of dietary glucose to 12% resulted in significantly smaller mammary gland cell size but failed to produce any significant differences in mammary gland composition (protein, fat and glycogen). The results indicate that the level of glucose in the maternal diet is an important determinant of milk composition and that > 24% glucose is needed for optimal milk fat concentration, whereas 12% dietary glucose is sufficient to sustain normal milk lactose concentration. The altered lactational performance in response to dietary glucose restriction was not mediated through changes in mammary gland composition, but in part by the reduced mammary gland size.

Restriction of maternal dietary carbohydrate decreases fetal brain indoles and glycogen in rats.

In spite of evidence that dietary carbohydrate can increase brain tryptophan and 5-hydroxytryptamine in adult rats, the possible influence of maternal dietary carbohydrate on fetal brain indoles has received little attention. We studied the effect of graded levels (0, 4, 12 and 60%) of maternal dietary fructose or glucose fed throughout pregnancy on fetal brain glycogen and indoles. The diets were isoenergetic and met the NRC energy requirements for pregnant rats. The results demonstrated that low maternal dietary carbohydrate, with adequate energy intake, reduced fetal brain weight and concentrations of glycogen, tryptophan, 5-hydroxytryptamine and 5-hydroxyindoleacetic acid. There were no significant differences between glucose and fructose feeding at any dietary carbohydrate level for any fetal brain measurements, showing that it was the level and not the type of dietary carbohydrate that was important. Significant correlations between fetal brain 5-hydroxytryptamine and brain glycogen, and between fetal brain 5-hydroxytryptamine and brain weight, suggested that lowered brain 5-hydroxytryptamine was only one symptom of disrupted brain development in fetuses of dams fed low levels of carbohydrate. The results show that dietary carbohydrate restriction during pregnancy can have adverse effects on fetal brain development, glycogen levels, and neurotransmitter synthesis even when maternal dietary protein and energy intake are adequate.

Placental composition does not respond to changes in maternal dietary carbohydrate intake in rats.

In this study we investigated whether placental glycogen reserves and protein and DNA content could be manipulated by altering the level of glucose in the maternal diet. Pregnant rat dams were fed isocaloric diets containing graded levels of glucose (0, 12, 24 and 60%), and placentas were analyzed for glycogen, protein and DNA content on gestational days 18.5 to 21.5. Regardless of the level of glucose in the maternal diet, there was a significant increase in placental size with advancing age, which was characterized by protein accretion but not by an increase in cell number of glycogen content. Restriction of glucose in the diets of pregnant dams failed to produce statistically significant reductions in placental protein, DNA and glycogen and did not retard placental growth, even though intrauterine growth retardation was observed. Fetal weight, plasma glucose, and liver and heart glycogen were positively correlated with placental weight and inversely correlated with placental glycogen and DNA concentrations; by contrast, no significant correlations were calculated between maternal and placental variables. Our study indicates that the placenta is not affected by a specific dietary glucose restriction and that changes in placental weight or glycogen content do not account for the growth retardation observed in fetuses of dams fed glucose-restricted diets.