Maternal Ethanol Research Articles

Metabolic response to starvation at late gestation in chronically ethanol-treated and pair-fed undernourished rats

To study the role of undernourishment in the negative effects of ethanol during pregnancy and to determine whether maternal ethanol intake modifies metabolic response to starvation at late gestation, female rats receiving ethanol in their drinking water before and during pregnancy (ethanol group) were compared with animals that received the same amount of solid diet as the ethanol group rats (pair-fed group) and with normal rats fed ad libitum (control group). All animals were killed on the 21st day of gestation, either in the fed state or after 24-hours fasting. The body weight of ethanol rats was lower than that of controls but higher than that of pair-fed rats. When compared with controls, ethanol and pair-fed rats had reduced fetal body weights, whereas fetal body length was reduced only in the former. In the fed state, blood glucose concentration was lower in the ethanol and pair-fed rats and fetuses than in controls. Twenty-four-hour starvation caused a reduction in this parameter only in control and ethanol mothers. In the fed state, maternal liver glycogen concentration was lower in ethanol and higher in pair-fed mothers than in controls. Blood β-hydroxybutyrate levels were higher in ethanol-treated mothers than in the others, and 24-hour starvation increased this parameter in ethanol and control rats to a greater extent than in the pair-fed ones. Liver triacylglyceride concentration was higher in ethanol-treated mothers than in the other two groups, and starvation caused this concentration to increase in ethanol and control groups but not in the pair-fed group. Maternal liver lipoprotein lipase activity did not differ among the groups when fed, but increased significantly with starvation in controls and ethanol-treated rats. Individual values of lipoprotein lipase activity in all the starved groups correlated with liver triacylglyceride concentrations. Results indicate that maternal undernourishment in ethanol-treated rats contributes to the negative effects of ethanol on fetal development in addition to certain other changes, such as the increase in maternal redox state and in circulating and liver triglycerides, directly related to ethanol oxidation. The intense maternal hypoglycemia observed in rats from the ethanol group could impair glucose availability to the fetus, and this situation would worsen with starvation. The higher hepatic glycogen levels in pair-fed mothers and their fetuses in the fed state as compared with the ethanol group, in spite of their similar hypoglycemia, seems to indicate a different metabolic adaptation in the two groups. A relationship is proposed in the starved condition between maternal liver lipoprotein lipase activity and changes in liver triacylglyceride and circulating ketone bodies in the three groups studied. The findings seem to indicate that, in spite of the metabolic disturbances produced by ethanol intake in the mother, the response to starvation in ethanol rats was similar to the one observed in controls, and different from the one in the pair-fed animals.

Chronic maternal ethanol consumption results in decreased serotonergic 5-HT 1 sites in cerebral cortical regions from offspring

This laboratory previously demonstrated that chronic maternal ethanol consumption results in a marked deficiency of cortical serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) and of 5-HT uptake sites in the 19- and 35-day-old offspring. In order to determine whether in utero exposure to ethanol similarly affects other components of the serotonergic system we examined the influence of chronic maternal ethanol consumption on cortical, serotonergic 5-HT 1 binding sites in developing offspring. Female Sprague-Dawley rats were pair-fed, using control or 6.6% (v/v) ethanol liquid diets on a chronic basis prior to parturition. Serotonergic 5-HT 1 sites were measured in synaptosomal membranes from whole cortex and cortical regions from developing offspring. Serotonergic 5-HT 1 sites were assessed by measuring the binding of [ 3H]-5-HT to synaptosomal membranes in the presence and absence of nonradioactive 5-HT. Serotonergic 5-HT 2 sites were blocked by including 100 nM spiperone in the assay buffer. The results demonstrated that the 19- and 37-day-old offspring of ethanol-fed rats had a significant (∼10–40%) reduction in the B max for serotonergic 5-HT 1 binding sites on synaptosomal membranes from whole cortex ( p<0.025), motor cortex ( p<0.01), and somatosensory cortex ( p<0.025). However, the binding affinity (K d) for serotonin was not significantly altered ( p>0.05). These results emphasize the sensitivity of the developing cortical serotonergic system to prenatal ethanol exposure.

Acetaldehyde production and transfer by the perfused human placental cotyledon.

Fetal injury associated with maternal ethanol ingestion is a major cause of congenital anomalies and mental retardation. Studies with animals suggest that acetaldehyde, the primary hepatic oxidative metabolite of ethanol, may contribute to fetal damage. It is not known, however, whether acetaldehyde reaches the human fetus, either by placental production or transfer. Studies utilizing the perfused human placental cotyledon show that the human placenta oxidizes ethanol to acetaldehyde, releasing it into the fetal perfusate. Moreover, when acetaldehyde is present in the maternal perfusate, it is transferred to the fetal side, reaching approximately 50 percent of the maternal level. These findings suggest that the human placenta may play a pivotal role in the pathophysiology of ethanol-associated fetal injury.

Alterations of fetal brain biogenic amine metabolites by maternal ethanol exposure

1. 1. Pregnant Sprague-Dawley rats treated chronically with ethanol (3 g/kg daily for the last third of pregnancy) had decreased placental weights at birth ( ca 23%). 2. 2. Whole fetal brain levels of HVA and 5-HIAA were similarly decreased 32 and 38%, respectively. 3. 3. MHPG levels were also marginally reduced (i.e. 15% decline). 4. 4. In the presence of a potent type A MAO inhibitor (harmaline, 10 mg/kg maternal weight 2 hr before fetal delivery) co-treatment with ethanol was found to result in a 60% elevation of whole fetal brain norepinephrine levels. 5. 5. Other biogenic amines and metabolites were not altered by ethanol treatment.

Mouse placental receptor for basic somatomedin following maternal ethanol administration.

We have previously reported a reduction in the basic somatomedin (B-SM) binding activity of day-15 mouse placental membranes following 3 days of acute maternal ethanol administration. In the present experiments, we have investigated the effects of acute maternal ethanol administration early in gestation on the subsequent development of the placental B-SM receptor, and its relationship to alcohol-related embryofetal growth deficits. Following administration of aqueous ethanol (0.0, 3.6, 5.5 or 7.1 ml/kg) by gavage on days 7, 8 and 9 of gestation, there was no evidence of reproductive impairment in alcohol-treated dams, but there was a significant reduction in day-15 embryonic body weight. Crown-rump length was unaffected. There was no treatment-related difference in the percentage specific binding of [125I]-B-SM by day-15 placental membranes, or in maternal serum B-SM concentrations. These observations suggest that embryofetal growth restriction following maternal ethanol administration is not mediated directly by peripheral unresponsiveness of the B-SM receptor, at least when the exposure occurs early in or prior to placental receptor ontogeny.

Maternal ethanol use and selective fetal malnutrition.

Maternal ethanol ingestion may cause fetal injury, particularly impaired somatic and brain growth, by at least two mechanisms: (1) directly, by fetotoxicity from ethanol and/or acetaldehyde; (2) indirectly, by ethanol-induced placental injury and selective fatal malnutrition. There is ample evidence that ethanol and/or acetaldehyde is fetotoxic. In addition, alcoholics are frequently in a state of poor nutrition. However, regardless of maternal nutritional status, ethanol can be placentotoxic, impairing the normal transfer of essential fetal nutrients. Ethanol and/or acetaldehyde has been shown to inhibit placental uptake and/or transfer of amino acids, zinc, and glucose. These effects have been evaluated in animal models and in human tissue. Recent use of the isolated perfused human placental cotyledon has begun to delineate the pathophysiology of ethanol-induced placental injury.

Fetal alcohol syndrome. Eye malformations in a mouse model.

Acute maternal ethanol administration on gestational day 7 (gastrulation stage) in C57Bl/6J mice results in a spectrum of ocular malformations. A deficiency in the anterior neural plate observable within 24 hours of exposure results in corresponding defects in the optic sulcus and subsequent optic vesicle. Deficiency in the size of the lens vesicle induced by a small optic vesicle is demonstrable as microphakia in older embryos. Delayed detachment of the lens vesicle from the surface ectoderm manifests in the live offspring as progressive corneal opacification and vascularization related to defects in corneal endothelium and Descemet's membrane. Anterior segment dysgenesis results in persistent iridocorneal adhesions, dyscoria, and abnormal formation of the anterior chamber. In contrast, ethanol exposure on day 8 of gestation did not result in eye malformations. Thus, it appears that many of the ocular abnormalities associated with fetal alcohol syndrome may result from an acute insult to the optic primordia during a very specific period that corresponds to the third week after fertilization in the human.

The role of fetal urinary excretion in the transfer of ethanol into amniotic fluid after maternal administration of ethanol to the near-term pregnant ewe.

The objective of this study was to determine whether fetal urinary excretion is a major route of ethanol transfer into the amniotic fluid surrounding the fetus following maternal administration of ethanol. Conscious instrumented pregnant ewes between 130 and 137 days' gestation (term, 147 days) with (n = 3) or without (n = 3) a catheter in the fetal bladder were administered 1 g ethanol/kg maternal body weight as a 1-h maternal intravenous infusion. Maternal blood, fetal blood, and amniotic fluid samples were collected at selected times, and fetal urine was collected continuously from the bladder-cannulated fetus during the 14-h study for the determination of ethanol concentrations. Fetal urinary excretion of ethanol occurred, and the total amount of ethanol excreted represented 0.30 +/- 0.07 (SD)% of the maternal ethanol dose. The renal clearance of ethanol by the fetus was 0.43 +/- 0.06 mL/min. The pharmacokinetics of ethanol in the maternal-fetal unit and the amniotic fluid for the bladder-cannulated fetal preparation were similar to the data for the nonbladder-cannulated preparation. The data indicate that fetal urinary excretion of ethanol is a secondary route of ethanol transfer into the amniotic fluid. It would appear that diffusion of ethanol across membranes from the maternal and fetal circulations is a major route of ethanol transfer into this intrauterine compartment.

Decreased urine production in the near-term fetal lamb after maternal ethanol infusion

Intravenous infusion of 1 gm ethanol/kg maternal body weight over 1 hour to three conscious catheterized near-term pregnant ewes decreased fetal urine production for 3 hours (overall decrease of 54% from control). This effect in the near-term fetus is opposite to the ethanol-induced diuresis that occurs in adults.

Maternal ethanol exposure with adequate protein in the diet: Influence on offspring development

Pregnant rats (n=45) were fed alcohol (29% of kcal) in a high protein (31% of kcal) liquid diet on days 6 through 19 of gestation. On day 1 (day 0 is day of birth), litters were culled to 8 pups, 4 males and 4 females when possible. Samples for blood assays, organ weights and thymus samples for histologic preparation were obtained from pups culled from the litters. Pups were randomly selected for urinary pH testing, blood assays, thymus studies and liver lipid assays on days 1, 16, 21 and 30 and for sciatic nerve myelin assay on day 21. Hematology, thymus protein and structure, liver lipids and cholesterol, and sciatic nerve myelin were unharmed by in utero alcohol exposure in the presence of a high protein diet. At day 16, body and brain weights of alcohol exposed pups were decreased and urinary pH levels were higher than those of pairfed and ad libitum controls. In utero alcohol exposure (29%) in a diet containing 31% kcal as protein did not affect hematology, liver and thymus structure or peripheral nerve myelin; but did have early affects on body, brain, kidney and thymus weights and urinary pH levels.

Stimulatory effects of ethanol on amino acid transport by rat fetal hepatocytes.

Previous studies have indicated that acute, and especially chronic, maternal ethanol consumption can depress placental uptake of various amino acids. Since the fetal cell itself represents a second barrier to nutrients, one which may be altered by ethanol exposure, the effects of ethanol on amino acid net uptake by rat fetal hepatocytes was addressed. The present study determined that ethanol stimulated amino acid net uptake by fetal hepatocytes grown in monolayer culture. Fetal liver cells were grown in custom Williams' E medium (without L-arginine and with L-ornithine) and exposed to epidermal growth factor (0, 1, 2 or 5 ng per ml) and ethanol (1.7 +/- 0.1 or 3.9 +/- 0.2 mg per ml). Addition of ethanol (3.9 mg per ml) to the culture medium completely blocked measurable cell replication during a 48-hr exposure period. Fetal hepatocytes exposed to ethanol accrued both protein and water in a parallel fashion, both in excess of that by control cells. Ethanol (1.7 and 3.9 mg per ml) for 48 hr stimulated alpha-aminoisobutyric acid net uptake by fetal hepatocytes (p less than 0.05). Efflux was not affected (p greater than 0.05). The onset of this significant stimulation of net uptake was progressive and required in excess of 6 hr of contact with ethanol. This ethanol stimulation of alpha-aminoisobutyric acid net uptake persisted for at least 24 hr following ethanol withdrawal. The component(s) of alpha-aminoisobutyric acid net uptake stimulated by ethanol was independent of extracellular Na+.(ABSTRACT TRUNCATED AT 250 WORDS)

Polyamines and diamine oxidase activity in maternal, embryonal, and fetal tissues of rat after chronic ethanol consumption

The effects of maternal ethanol consumption for 4 weeks before and throughout gestation on polyamine content and diamine oxidase activity of meternal, embryonal and fetal tissues are reported. At the 12th day of pregnancy, a decrease of putrescine in the liver of the mother and marked increases in putrescine, cadaverine and spermidine in embryos were observed. At day 18, putrescine and cadaverine diminished in maternal liver and placenta, and no changes in amine content in fetal liver and brain were found. At day 12, diamine oxidase activity increased in maternal liver and placenta, whereas it greatly diminished in embryos. At day 18, enzyme activity decreased in maternal liver, placenta, fetal liver and brain. These results indicate that chronic ethanol ingestion induces alterations in polyamine concentrations and metabolism in growing and developing tissues during pregnancy that might contribute to the adverse effect of ethanol on conceptual development.

Maternal ethanol consumption: Effect on skeletal muscle development in guinea pig offspring

The effect of ethanol on developing skeletal muscle was analyzed by examining the gastrocnemius muscle from newborn guinea pigs, exposed to ethanol during the second half of gestation. Electron microscopy revealed vacuolated sarcoplasmic reticula, enlarged lipid droplets, decreased glycogen and mitochondrial abnormalities in the skeletal muscle samples from the ethanol-exposed newborn guinea pigs. None of these abnormalities were seen in the newborn controls who were born to dams which consumed the same amount of calories as the ethanol-treated dams. The ethanol-associated abnormalities, seen in this study, are similar to those seen in ultrastructural examination of skeletal muscle from chronic alcoholics.

Maternal ethanol consumption: Lack of effect on synaptogenesis in layer I of the motor cortex in 19-day-old rat offspring

Previous studies showed that in utero exposure to ethanol has an adverse effect on development of the central nervous system. Included among those abnormalities is the evidence that in utero exposure to ethanol alters the development of the cerebral cortex in terms of the arborization of pyramidal dendrites and the concentration of neurotransmitters and neurotransmitter receptors. However, limited information is available about the influence of in utero exposure to ethanol on synaptogenesis in the cortex. We examined the influence of chronic maternal ethanol consumption, prior to parturition, on synaptogenesis in layer I of the motor cortex of 19-day-old-rat offspring. Layer I (the molecular layer) was chosen because of the early timing of its development. The motor cortex was chosen because of the evidence of motor problems in children afflicted with the fetal alcohol syndrome. The results of these studies demonstrated that the 19-day-old offspring of rats that were pair-fed, using control or 6.6% (v/v) ethanol-containing liquid diets on a chronic basis prior to parturition, had a comparable density of synapses and distribution of paramembranous density lengths in layer I of the motor cortex as well as a comparable thickness of cortical layer I. Thus, evidence by this and other laboratories, of cortical neurotransmitter and dendritic abnormalities are not reflected in synaptic changes in layer I of the motor cortex at 19 days. It is possible that synaptogenesis is affected at another age or in other regions of the cerebral cortex.

Reduction of the volume of the sexually dimorphic nucleus of the preoptic area by in utero ethanol exposure in male rats

Ethanol consumption during pregnancy has been demonstrated to result in alterations in central nervous system structure and function. Albino female rats were given either a liquid diet containing ethanol (5% w/v), a liquid diet which contained maltose-dextrin substituted isocalorically for the alcohol, or laboratory chow and water throughout pregnancy and for 14 days following birth. The volumes of the sexually dimorphic nucleus of the preoptic area (SDN-POA) and the nucleus accumbens septi (ACN) were measured in adult male rats exposed in utero and postnatally to these diets. Adult male rats exposed to ethanol in utero exhibit significantly smaller SDN-POA volumes ( P<0.02) when compared to those of animals which received the control diets during the gestational period and postnatally. Ethanol treatment did not significantly influence the volume of the ACN. The results indicate that maternal ethanol consumption during pregnancy retards or inhibits SDN-POA development in fetal ethanol exposed male rats when compared to the nuclear volume in rats whose mothers were not exposed to ethanol during pregnancy.

Maternal ethanol ingestion: effect on maternal and neonatal glucose balance.

Liver glycogen availability in the newborn is of major importance for the maintenance of postnatal blood glucose levels. This study examined the effect of maternal ethanol ingestion on maternal and neonatal glucose balance in the rat. Female rats were placed on the Lieber-DeCarli liquid ethanol diet, an isocaloric liquid pair-fed diet, or an ad libitum rat chow diet at 3 wk before mating and throughout gestation. Blood and livers were obtained from dams and rat pups on gestational days 21 and 22. The pups were studied up to 6 h in the fasted state and up to 24 h in the fed state. Maternal ethanol ingestion significantly decreased litter size, birth weight, and growth. A significantly higher mortality during the early postnatal period was seen in the prenatal ethanol exposed pups. Ethanol significantly decreased fed maternal liver glycogen stores but not maternal plasma glucose levels. The newborn rats from ethanol ingesting dams also had significantly decreased liver glycogen stores. Despite mobilizing their available glycogen, these prenatal ethanol exposed pups became hypoglycemic by 6 h postnatal. This was more marked in the fasted pups. Ethanol did not affect maternal nor neonatal plasma insulin levels. Thus maternal ethanol ingestion reduces maternal and neonatal liver glycogen stores and leads to postnatal hypoglycemia in the newborn rat.

Organ growth and cellular development in ethanol-exposed rats

Effects of perinatal exposure to ethanol on growth and cellular development were investigated. Alcohol was administered in liquid diets designed to provide optimal nutrition during pregnancy. Pair-fed and ad lib control groups were included. The 3 groups of females were similar in body weight during gestation and lactation, and offspring weights were similar on gestation Day 21 and at birth. By Day 9 of lactation control pups weighed more than both alcohol and pair-fed pups which were similar in body weight. Weights of brain, heart, liver and kidney were reduced in alcohol pups compared to pair-feds and controls. Decreased liver weight reflected both decreased cell size and decreased protein content, but was primarily due to decreased caloric intake. Decreased heart weight appeared to result from a direct effect of ethanol on heart protein content. Even more marked were the adverse effects of ethanol on kidney protein content and kidney DNA (reflecting a decrease in cell number). In contrast, although both absolute brain weight and DNA content were decreased in ethanol-exposed offspring, relative brain weight was increased. Finally, maternal ethanol consumption significantly increased relative placenta weights as well as placental DNA and protein content.

Development of hepatic bilirubin glucuronidation as affected by prolonged maternal ethanol intake

Prolonged maternal ethanol intake by lactating mothers resulted in a 26% and 41% decrease in serum conjugated bilirubin of 6 and 10 day old suckling newborn rats, respectively. An increase of 20% and 36% was observed in the serum unconjugated bilirubin levels in suckling newborns from the ethanol-fed group compared to the corresponding controls. Newborns suckling on ethanol-fed dams showed a small, but significant hyperbilirubinemia compared to the controls. Acute administration of ethanol (2.5 g/kg) to newborn rats also resulted in a decrease in serum conjugated bilirubin and an increase in the levels of unconjugated and total bilirubin. Prolonged ethanol intake by adults also resulted in a decrease in conjugated bilirubin and an increase in unconjugated and total bilirubin levels compared to the corresponding controls. Bilirubin glucuronide formation in the liver was decreased by ethanol by about 30% and 37% in the suckling newborn, and adults, respectively. Newborns suckling on ethanol-fed dams, as well as the dams consuming ethanol showed an increase in hepatic (UDPG)/(UDPGA) levels. The activity of hepatic bilirubin-UDP-glucuronyl transferase either in the newborns suckling on ethanol-fed dams, or in dams consuming ethanol chronically, was not significantly different from their corresponding controls. Ethanol was found to inhibit the glucuronidation of bilirubin by decreasing the availability of UDP-glucuronic acid in the newborn and adult livers.

Ethanol-induced intrauterine growth retardation: correlation with placental glucose transfer.

Placental transfer of glucose and alanine analogs was studied at term in ethanol-fed and control rats. Ethanol was provided as 30% of the caloric intake throughout gestation. Control groups received isocaloric liquid diet without ethanol by pair-feeding (PF) or ad libitum (AF). On the 22nd day of pregnancy, the rats were injected with a mixture of [3H]2-deoxyglucose and [14C]alpha-aminoisobutyric acid. The ratios of fetal:maternal plasma radioactivities 1 hr later were used to compare placental transfer between the groups. Mean +/- SE body weight of EF fetuses (4.54 +/- 0.07 g) was significantly lower than that of PF (4.88 +/- 0.06 g) or AF (5.17 +/- 0.09 g) fetuses. Maternal ethanol ingestion reduced placental transfer of 2-deoxyglucose and alpha-aminoisobutyric acid by 12% and 35%, respectively. Placental transfer of both analogs was not affected in the PF controls. The weight of EF fetuses correlated (p less than 0.001) with transfer of 2-deoxyglucose to the fetus. This relationship was also found in the control groups. Fetal body weight did not show a strong correlation with alpha-aminoisobutyric acid transfer. Thus, impaired transfer of glucose to the fetus may play a significant role in the growth retardation observed in fetuses of ethanol-fed rats.

Maternal ethanol consumption: Binding of l-glutamate to synaptic membranes from whole brain, cortices, and cerebella of offspring

We examined the influence of chronic maternal ethanol consumption on the Na +- and Ca 2+-independent binding of l-glutamate to synaptic plasma membranes from whole brain as well as from cortices and cerebella of developing offspring. The maximum specific binding ( B max) of l-glutamate to the Na +- and Ca 2+-independent binding sites in synaptic plasma membranes of brain peaked at 17 days of age in the offspring of both control and ethanol-fed rats, although at that age there were significantly fewer binding sites in the brains of the offspring of ethanol-fed rats. The regional localization of this deficit is not now known. However, it appears that one major glutamatergic region (the cortex) does not reflect the transient deficiency of l-glutamate sites in brain. In fact, the concentration of l-glutamate binding sites in cortical synaptic plasma membranes was significantly increased in the 20-day-old offspring of ethanol-fed rats. In contrast to the cortex, binding to cerebellar synaptic plasma membranes was comparable in 20-day-old offspring of control and ethanol-fed rats. Despite transient alterations in the concentrations of l-glutamate binding sites in brain and synaptic plasma membranes, the affinity of the sites for l-glutamate ( K d) was consistently normal in the 14- to 26-day-old offspring of ethanol-fed rats.