EtOH-exposed Offspring Research Articles

Maternal Exposure to Ethanol During Pregnancy and Lactation Affects Glutamatergic System and Induces Oxidative Stress in Offspring Hippocampus.

Alcohol abuse during pregnancy leads to intellectual disability and morphological defects in the offspring. The aim of this study was to determine the effect of chronic maternal ethanol (EtOH) consumption during pregnancy and lactation on glutamatergic transmission regulation, energy deficit, and oxidative stress in the hippocampus of the offspring. EtOH was administered to dams in drinking water at increasing doses (2 to 20%) from the gestation day 5 to lactation day 21. EtOH and tap water intake by treated and control groups, respectively, were measured daily. Results showed that EtOH exposure does not affect fluid intake over the course of pregnancy and lactation. The toxicity of maternal exposure to EtOH was demonstrated by decreased offspring body weight at experimental age, on postnatal day 21. Moreover, maternal EtOH exposure decreased (45) Ca(2+) influx in the offspring's hippocampus. Corroborating this finding, EtOH increased both Na(+) -dependent and Na(+) -independent glial [(14) C]-glutamate uptakein hippocampus of immature rats. Also, maternal EtOH exposure decreased glutamine synthetase activity and induced aspartate aminotransferase enzymatic activity, suggesting that in EtOH-exposed offspring hippocampus, glutamate is preferentially used as a fuel in tricarboxylic acid cycle instead of being converted into glutamine. In addition, EtOH exposure decreased [U-14C]-2-deoxy-D-glucose uptake in offspring hippocampus. The decline in glucose transport coincided with increased lactate dehydrogenase activity, suggesting an adaptative response in EtOH-exposed offspring hippocampus, using lactate as an alternative fuel. These events were associated with oxidative damage, as demonstrated by changes in the enzymatic antioxidant defense system and lipid peroxidation. Taken together, the results demonstrate that maternal exposure to EtOH during pregnancy and lactation impairs glutamatergic transmission, as well as inducing oxidative stress and energy deficit in immature rat hippocampus.

Exposure to ethanol during neurodevelopment modifies crucial offspring rat brain enzyme activities in a region-specific manner.

The experimental simulation of conditions falling within "the fetal alcohol spectrum disorder" (FASD) requires the maternal exposure to ethanol (EtOH) during crucial neurodevelopmental periods; EtOH has been linked to a number of neurotoxic effects on the fetus, which are dependent upon the extent and the magnitude of the maternal exposure to EtOH and for which very little is known with regard to the exact mechanism(s) involved. The current study has examined the effects of moderate maternal exposure to EtOH (10 % v/v in the drinking water) throughout gestation, or gestation and lactation, on crucial 21-day-old offspring Wistar rat brain parameters, such as the activities of acetylcholinesterase (AChE) and two adenosine triphosphatases (Na(+),K(+)-ATPase and Mg(2+)-ATPase), in major offspring CNS regions (frontal cortex, hippocampus, hypothalamus, cerebellum and pons). The implemented experimental setting has provided a comparative view of the neurotoxic effects of maternal exposure to EtOH between gestation alone and a wider exposure timeframe that better covers the human third trimester-matching CNS neurodevelopment period (gestation and lactation), and has revealed a CNS region-specific susceptibility of the examined crucial neurochemical parameters to the EtOH exposure schemes attempted. Amongst these parameters, of particular importance is the recorded extensive stimulation of Na(+),K(+)-ATPase in the frontal cortex of the EtOH-exposed offspring that seems to be a result of the deleterious effect of EtOH during gestation. Although this stimulation could be inversely related to the observed inhibition of AChE in the same CNS region, its dependency upon the EtOH-induced modulation of other systems of neurotransmission cannot be excluded and must be further clarified in future experimental attempts aiming to simulate and to shed more light on the milder forms of the FASD-related pathophysiology.

The effects of low-moderate dose prenatal ethanol exposure on the fetal and postnatal rat lung.

Little is known about whether exposure of the fetus to alcohol alters pulmonary development or function. This study aimed to determine whether low-moderate ethanol (EtOH) exposure throughout gestation alters structural and non-respiratory functional aspects of the fetal and postnatal lung. Sprague-Dawley rats were fed an ad libitum liquid diet ±6% v/v EtOH daily throughout pregnancy, achieving a plasma ethanol (EtOH) concentration of 0.03%. Gene and protein expression was determined in pulmonary tissue collected from fetuses at embryonic day (E) 20 and adult offspring. The percentage of airspace and alveolar size was measured in pulmonary tissue collected at postnatal day (PN) 1. At E20, EtOH-exposed fetuses had decreased aquaporin 5 mRNA levels and a non-significant trend for decreased epithelial sodium channel type α; expression of other pulmonary fluid homeostatic and development genes and surfactant protein genes were not different between groups. At PN1, there was no difference between EtOH-exposed and control offspring in the distal airspace percentage or diameter. At 8 months, collagen type III α1 gene expression was upregulated in EtOH-exposed male offspring; this was associated with increased collagen deposition at 10 months. At 19 months, male EtOH-exposed offspring had a 25% reduction in the protein levels of surfactant protein B. The alterations observed in male EtOH-exposed offspring suggest chronic low-moderate prenatal EtOH-exposure during development may result in increased pulmonary fibrosis. Such an alteration would decrease the respiratory capacity of the lung.

Intrauterine ethanol exposure results in hypothalamic oxidative stress and neuroendocrine alterations in adult rat offspring

Prenatal ethanol (EtOH) exposure is associated with low birth weight, followed by increased appetite, catch-up growth, insulin resistance, and impaired glucose tolerance in the rat offspring. Because EtOH can induce oxidative stress, which is a putative mechanism of insulin resistance, and because of the central role of the hypothalamus in the regulation of energy homeostasis and insulin action, we investigated whether prenatal EtOH exposure causes oxidative damage to the hypothalamus, which may alter its function. Female rats were given EtOH by gavage throughout pregnancy. At birth, their offspring were smaller than those of non-EtOH rats. Markers of oxidative stress and expression of neuropeptide Y and proopiomelanocortin (POMC) were determined in hypothalami of postnatal day 7 (PD7) and 3-mo-old (adult) rat offspring. In both PD7 and adult rats, prenatal EtOH exposure was associated with decreased levels of glutathione and increased expression of MnSOD. The concentrations of lipid peroxides and protein carbonyls were normal in PD7 EtOH-exposed offspring, but were increased in adult EtOH-exposed offspring. Both PD7 and adult EtOH-exposed offspring had normal neuropeptide Y and POMC mRNA levels, but the adult offspring had reduced POMC protein concentration. Thus only adult offspring preexposed to EtOH had increased hypothalamic tissue damage and decreased levels of POMC, which could impair melanocortin signaling. We conclude that prenatal EtOH exposure causes hypothalamic oxidative stress, which persists into adult life and alters melanocortin action during adulthood. These neuroendocrine alterations may explain weight gain and insulin resistance in rats exposed to EtOH early in life.

Glucose intolerance and resistin expression in rat offspring exposed to ethanol in utero: modulation by postnatal high-fat diet.

High-fat diet and intrauterine growth retardation may predispose to obesity, insulin resistance, and type 2 diabetes. Because prenatal ethanol (ETOH) exposure causes intrauterine growth retardation, we investigated its interactions with postnatal high-fat diet on glucose tolerance and adipocyte-derived hormones in the rat offspring. High-fat-fed offspring had increased adiposity, serum leptin, and muscle uncoupling protein-3, but decreased adiponectin mRNA, compared with corresponding chow-fed groups. ETOH-exposed offspring had normal adiponectin, but increased resistin mRNA and protein, compared with controls, regardless of postnatal diet. Skeletal muscle glucose transporter-4 content was decreased after both ETOH exposure and high-fat feeding. Glycemic and insulin responses to an ip glucose challenge were equally increased in non-ETOH-exposed high-fat-fed offspring and in ETOH-exposed chow-fed offspring, with additive effects of ETOH and high-fat diet. Pancreatic insulin content was elevated only in non-ETOH-exposed high-fat-fed offspring. The data suggest that high-fat diet worsens glucose intolerance in offspring of rats exposed to ETOH. Prenatal ETOH exposure and postnatal high-fat diet might cause insulin resistance through separate mechanisms, involving resistin and adiponectin, respectively.

The Use of a Silymarin/Phospholipid Compound as a Fetoprotectant from Ethanol-Induced Behavioral Deficits

We have replicated an earlier study in which silymarin/phytosome appeared to prevent deficits in social memory function in male rats exposed in utero to ethanol (EtOH).1 Female rats were included in the current study as well as a second behavioral test, the radial arm maze. Pregnant Sprague-Dawley rats were provided with liquid diets containing 35% ethanol derived calories (EDC). The silybin/phospholipid compound (SI) was co-administered with EtOH to the experimental group. The offspring were tested at age 90 days on the social recognition task and at 75 days on the radial arm maze. Female EtOH-exposed offspring performed more poorly on the radial arm maze than did female EtOH/SI offspring and the offspring of female controls. Male EtOH-exposed offspring were less able to form social memories than the male EtOH/SI offspring and the offspring of male controls.

The Use of a Silymarin/Phospholipid Compound as a Fetoprotectant from Ethanol-Induced Behavioral Deficits

We have replicated an earlier study in which silymarin/ phytosome appeared to prevent deficits in social memory function in male rats exposed in utero to ethanol (EtOH).1 Female rats were included in the current study as well as a second behavioral test, the radial arm maze. Pregnant Sprague-Dawley rats were provided with liquid diets containing 35% ethanol derived calories (EDC). The silybin/phospholipid compound (SI) was co-administered with EtOH to the experimental group. The offspring were tested at age 90 days on the social recognition task and at 75 days on the radial arm maze. Female EtOH-exposed offspring performed more poorly on the radial arm maze than did female EtOH/SI offspring and the offspring of female controls. Male EtOH-exposed offspring were less able to form social memories than the male EtOH/SI offspring and the offspring of male controls.

Prenatal ethanol exposure alters sensitivity to the effects of apomorphine given alone and in combination with ethanol on locomotor activity in adult male mouse offspring.

Previous studies have indicated that prenatal ethanol (EtOH) exposure alters developing catecholamine (CA) systems and acute sensitivity to the locomotor stimulant effects of EtOH. The purpose of this study was to examine whether prenatal EtOH exposure influences the effects of the direct dopamine (DA) agonist apomorphine given alone as well as in combination with a low-dose stimulant challenge of EtOH. Standard lab chow or liquid diets containing either 25% EtOH-derived calories (EDC), or 0% EDC were given to pregnant C3H/He mice on gestation days 6-18. At 90 days of age, male offspring from each prenatal treatment group were monitored for 10 min in an open field following IP injections of apomorphine (0, 0.15, 0.3, 0.6, or 1.2 mg/kg) and either EtOH (1.5 g/kg) or saline. EtOH alone increased activity by 120-143% in all three groups of offspring. In control offspring, apomorphine dose-dependently decreased activity up to 74%-78% and blocked the stimulant effect of EtOH at all doses tested. However, in prenatal EtOH-exposed offspring, higher doses of apomorphine were significantly less effective in reducing both baseline and EtOH-stimulated activity compared to control mice. This effect is most likely not due to differences in pharmacokinetics, because blood EtOH concentrations were similar across apomorphine doses and prenatal treatment conditions. As such, these results support the hypothesis that prenatal exposure to EtOH alters acute sensitivity to the locomotor stimulant effects of EtOH, particularly under conditions in which CA systems mediating those effects are additionally challenged. In addition, the results suggest that prenatal EtOH exposure results in a long-lasting perturbation of central DA receptor sensitivity.

Reduced sensitivity to the effects of clonidine on ethanol-stimulated locomotor activity in adult mouse offspring prenatally exposed to ethanol

Prenatal exposure to ethanol (EtOH) alters developing catecholamine (CA) systems and acute sensitivity to the locomotor stimulant effects of EtOH. As an extension of previous work involving CA agents, this study addressed whether prenatal EtOH exposure influences central norepinephrine (NE) systems by examining the motoric effects of the direct α 2 adrenoreceptor agonist clonidine given alone and in combination with a low-dose stimulant challenge of EtOH. Standard lab chow or liquid diets containing either 25% EtOH-derived calories (EDC), or 0% EDC (pair-fed group) were given to pregnant C3H/He mice on gestation days 6–18. At 90 days of age, male offspring from each prenatal treatment group were monitored for 10 minutes in an open field following IP injections of clonidine (0, 0.0125, 0.025, or 0.05 mg/kg) and either EtOH (1.5 g/kg) or saline. In control offspring, clonidine suppressed locomotor activity and attenuated the stimulant response to EtOH in a dose-dependent fashion. In contrast, clonidine given alone did not suppress, but appeared to stimulate, activity in prenatal EtOH-exposed offspring. Furthermore, the ability of clonidine to attenuate the locomotor stimulant properties of EtOH was greatly reduced in prenatal EtOH-exposed animals. Taken together, these results indicate a shift to the right in the doseresponse function for clonidine in prenatal EtOH-exposed offspring relative to control mice. Further, the results suggest that prenatal exposure to EtOH may result in long-lasting alterations in developing central NE systems, particularly presynaptic α 2 adrenoreceptor sensitivity.

Patterns of growth deficiency in rats exposed in utero to undernutrition, ethanol, or the neuroteratogen methylazoxymethanol (MAM)

Children and experimental animals exposed to ethanol (EtOH) in utero commonly have low birthweights, and many remain small at maturity. Low body weight or small stature in adulthood may reflect an inability to recover from in utero growth retardation, or it may reflect a separate, postnatal growth deficiency. In this study, daily body weights (postnatal days 1 to 60) were compared among the offspring of the following groups of Long Evans rats: dams fed liquid diet containing 35% EtOH-derived calories; their pair-fed and chow-fed controls; and dams exposed to methylazoxymethanol (MAM) in two previous studies, in which offspring exhibited reduced numbers of growth hormone releasing factor (GRF) neurons. All treatments produced a number of offspring with weight deficits beginning after birth and persisting into maturity. Three distinct patterns of growth deficiency were observed: (1) weight loss relative to controls in the first weeks of life, seen in offspring exposed to EtOH, pair feeding, or MAM on gestation day 13 (G13); (2) a delay in the onset of the prepubertal growth spurt, seen in all EtOH-exposed offspring and in G13 MAM-exposed dwarfs; and (3) failure to sustain the prepubertal growth spurt, seen only after exposure to MAM on G14. The results of this study support the view that prenatal EtOH exposure is capable of affecting postnatal growth specifically; moreover, the pattern of growth deficiency seen in EtOH-exposed offspring was distinct from that of the undernourished offspring of pair-fed dams.

Changes in the reproductive system following acute prenatal exposure to ethanol or methylazoxymethanol in the rat: I. Effects on immunoreactive LHRH cell number

It has long been recognized that ethanol (EtOH) interferes with the hypothalamo-pituitary-gonad axis in adults of many species, and recent studies have provided evidence for similar effects after prenatal EtOH exposure. Since EtOH is capable of injuring dividing cells, we investigated the possibility that a single acute in utero EtOH exposure during the period of LHRH neuron formation might change the number of immunoreactive LHRH cells in the hypothalamus. Final LHRH cell division in Long-Evans rats was determined by [3H]thymidine autoradiography to take place over a short period between gestation days 12 and 13. Subsequently, pregnant rats were treated acutely with either EtOH or methylazoxymethanol (MAM), a known neuroteratogen, and the numbers of immunoreactive LHRH cells were counted. On gestation day 22, LHRH-positive cell numbers were significantly fewer than control numbers in both EtOH- and MAM-exposed offspring. On postnatal day 60, cell numbers in EtOH-exposed offspring did not differ from control numbers, whereas cells in MAM-exposed offspring remained significantly reduced. In controls, there were 40% fewer LHRH-positive cells on postnatal day 4 than in late gestation or at maturity. We conclude that 1) acute exposure to a high dose of EtOH at a critical time in early gestation can alter the expression of LHRH in late gestation; 2) exposure to MAM in the same period alters LHRH expression before birth and in the adult; and 3) in the early postnatal period, LHRH expression decreases profoundly.

Effects of prenatal ethanol exposure on later sensitivity to the low-dose stimulant actions of ethanol in mouse offspring: possible role of catecholamines.

The purpose of this study was to examine whether prenatal ethanol (EtOH) exposure alters later sensitivity to the low-dose stimulant effects of EtOH. Because the locomotor stimulant effects of EtOH are thought to be mediated, at least in part, by activation of brain monoamine systems, and because prenatal EtOH exposure has been shown to alter brain monoamine activity, it was hypothesized that prenatal EtOH exposure may alter sensitivity to the stimulant actions of EtOH. To test this hypothesis, sensitivity to the locomotor stimulant effects of various challenge doses of EtOH was examined in male and female offspring from prenatal alcohol (A), pair-fed (PF), and lab chow (LC) groups at different ages. In addition, to address the hypothesis further, sensitivity to the catecholamine synthesis inhibitor alpha-methyl-p-tyrosine (AMPT) was examined in these offspring, as well. Results indicated that male offspring prenatally exposed to EtOH exhibited reduced baseline activity and a blunted stimulant response to all challenge doses of EtOH (0.75-1.5 g/kg) in comparison with control offspring at 30 days of age, but these effects appeared to "normalize" at 70 days of age. Female EtOH-exposed offspring also exhibited a reduced baseline level of activity relative to control offspring, as well as a blunted stimulant response to the lowest challenge dose of EtOH (0.75 g/kg) at 30 days of age, and these effects persisted into adulthood. The stimulant response to higher doses of EtOH did not significantly differ among prenatal treatment groups in young or adult female offspring. However, because baseline activity was significantly lower in female EtOH-exposed offspring than control offspring, the stimulant response to these doses of EtOH (1.125 and 1.5 g/kg) was relatively greater than that for PF and LC offspring. Importantly, none of the differences in performance among the prenatal treatment groups could be attributed to an alteration in EtOH pharmacokinetics, because blood EtOH levels measured immediately following the 10-min test session were similar for all prenatal treatment groups across all of the EtOH test doses. Further, a similar response profile as that observed following EtOH challenge at 70 days of age was obtained following phenobarbital challenge (10-40 mg/kg). Finally, whereas AMPT (50-400 mg/kg) dose-dependently antagonized the stimulant effects of EtOH in all prenatal treatment groups, this effect of AMPT was significantly greater in mice prenatally exposed to EtOH in comparison with control offspring.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of prenatal ethanol exposure in C57BL mice on locomotor activity and passive avoidance behavior

The purpose of this study was to examine the long-term behavioral effects of prenatal ethanol exposure in C57BL mice. Pregnant mice received free access to a liquid diet containing 25% ethanol-derived calories (EDC) from gestation days 6 to 18. Control animals were pair-fed an isocaloric 0% EDC diet during the same period of time. An additional control group was included that was maintained on standard lab chow and water throughout pregnancy. At 30 days of age, female offspring were tested for spontaneous locomotor activity in an open field under two lighting conditions (dim or bright illumination). Male offspring were tested in a passive avoidance task at 25 days of age. The activity results demonstrated that the 25% EDC female progeny were more active than controls. This hyperactivity was observed under both lighting conditions, despite the fact that all groups evidenced suppressed activity when tested under bright lights. With regard to passive avoidance behavior, male EtOH-exposed offspring required a greater number of trials to reach criterion than controls. Additionally, they exhibited shorter latencies to enter the shock-associated chamber after receiving a single shock. Taken together, these results confirm our previous findings and demonstrate that C57BL mice are sensitive to both the deleterious behavioral and morphological consequences of prenatal ethanol exposure.