Effects Of Maternal Ethanol Exposure Research Articles

Effects of maternal ethanol exposure during pregnancy and lactation periods on anxiety- like behaviors and hippocampal brain-derived neurotrophic factor in rat offspring

Effects of maternal ethanol exposure during pregnancy and lactation periods on anxiety- like behaviors and hippocampal brain-derived neurotrophic factor in rat offspring

CCL2/CCR2 system in neuroepithelial radial glia progenitor cells: involvement in stimulatory, sexually dimorphic effects of maternal ethanol on embryonic development of hypothalamic peptide neurons

BackgroundClinical and animal studies show that alcohol consumption during pregnancy produces lasting behavioral disturbances in offspring, including increased alcohol drinking, which are linked to inflammation in the brain and disturbances in neurochemical systems that promote these behaviors. These include the neuropeptide, melanin-concentrating hormone (MCH), which is mostly expressed in the lateral hypothalamus (LH). Maternal ethanol administration at low-to-moderate doses, while stimulating MCH neurons without affecting apoptosis or gliogenesis, increases in LH the density of neurons expressing the inflammatory chemokine C-C motif ligand 2 (CCL2) and its receptor CCR2 and their colocalization with MCH. These neural effects associated with behavioral changes are reproduced by maternal CCL2 administration, reversed by a CCR2 antagonist, and consistently stronger in females than males. The present study investigates in the embryo the developmental origins of this CCL2/CCR2-mediated stimulatory effect of maternal ethanol exposure on MCH neurons.MethodsPregnant rats from embryonic day 10 (E10) to E15 during peak neurogenesis were orally administered ethanol at a moderate dose (2 g/kg/day) or peripherally injected with CCL2 or CCR2 antagonist to test this neuroimmune system’s role in ethanol’s actions. Using real-time quantitative PCR, immunofluorescence histochemistry, in situ hybridization, and confocal microscopy, we examined in embryos at E19 the CCL2/CCR2 system and MCH neurons in relation to radial glia progenitor cells in the hypothalamic neuroepithelium where neurons are born and radial glia processes projecting laterally through the medial hypothalamus that provide scaffolds for neuronal migration into LH.ResultsWe demonstrate that maternal ethanol increases radial glia cell density and their processes while stimulating the CCL2/CCR2 system and these effects are mimicked by maternal administration of CCL2 and blocked by a CCR2 antagonist. While stimulating CCL2 colocalization with radial glia and neurons but not microglia, ethanol increases MCH neuronal number near radial glia cells and making contact along their processes projecting into LH. Further tests identify the CCL2/CCR2 system in NEP as a primary source of ethanol’s sexually dimorphic actions.ConclusionsThese findings provide new evidence for how an inflammatory chemokine pathway functions within neuroprogenitor cells to mediate ethanol’s long-lasting, stimulatory effects on peptide neurons linked to adolescent drinking behavior.

Hypothalamic CCL2/CCR2 Chemokine System: Role in Sexually Dimorphic Effects of Maternal Ethanol Exposure on Melanin-Concentrating Hormone and Behavior in Adolescent Offspring.

Clinical and animal studies show that ethanol exposure and inflammation during pregnancy cause similar behavioral disturbances in the offspring. While ethanol is shown to stimulate both neuroimmune and neurochemical systems in adults, little is known about their anatomical relationship in response to ethanol in utero and whether neuroimmune factors mediate ethanol's effects on neuronal development and behavior in offspring. Here we examined in female and male adolescent rats a specific population of neurons concentrated in lateral hypothalamus, which coexpress the inflammatory chemokine C-C motif ligand 2 (CCL2) or its receptor CCR2 with the orexigenic neuropeptide, melanin-concentrating hormone (MCH), that promotes ethanol drinking behavior. We demonstrate that maternal administration of ethanol (2 g/kg/d) from embryonic day 10 (E10) to E15, while having little impact on glia, stimulates expression of neuronal CCL2 and CCR2, increases density of both large CCL2 neurons colocalizing MCH and small CCL2 neurons surrounding MCH neurons, and stimulates ethanol drinking and anxiety in adolescent offspring. We show that these neuronal and behavioral changes are similarly produced by maternal administration of CCL2 (4 or 8 μg/kg/d, E10-E15) and blocked by maternal administration of a CCR2 antagonist INCB3344 (1 mg/kg/d, E10-E15), and these effects of ethanol and CCL2 are sexually dimorphic, consistently stronger in females. These results suggest that this neuronal CCL2/CCR2 system closely linked to MCH neurons has a role in mediating the effects of maternal ethanol exposure on adolescent offspring and contributes to the higher levels of adolescent risk factors for alcohol use disorders described in women.SIGNIFICANCE STATEMENT Ethanol consumption and inflammatory agents during pregnancy similarly increase alcohol intake and anxiety in adolescent offspring. To investigate how neurochemical and neuroimmune systems interact to mediate these disturbances, we examined a specific population of hypothalamic neurons coexpressing the inflammatory chemokine CCL2 and its receptor CCR2 with the neuropeptide, melanin-concentrating hormone. We demonstrate in adolescent offspring that maternal administration of CCL2, like ethanol, stimulates these neurons and increases ethanol drinking and anxiety, and these effects of ethanol are blocked by maternal CCR2 antagonist and consistently stronger in females. This suggests that neuronal chemokine signaling linked to neuropeptides mediates effects of maternal ethanol exposure on adolescent offspring and contributes to higher levels of adolescent risk factors for alcohol use disorders in women.

Comparative assessments of the effects of alcohol exposure on fetal brain development using optical coherence tomography and ultrasound imaging

The developing fetal brain is vulnerable to a variety of environmental agents including maternal ethanol consumption. Preclinical studies on the development and amelioration of fetal teratology would be significantly facilitated by the application of high resolution imaging technologies like optical coherence tomography (OCT) and high-frequency ultrasound (US). This study investigates the ability of these imaging technologies to measure the effects of maternal ethanol exposure on brain development, ex vivo, in fetal mice. Pregnant mice at gestational day 12.5 were administered ethanol (3 g/Kg b.wt.) or water by intragastric gavage, twice daily for three consecutive days. On gestational day 14.5, fetuses were collected and imaged. Three-dimensional images of the mice fetus brains were obtained by OCT and high-resolution US, and the volumes of the left and right ventricles of the brain were measured. Ethanol-exposed fetuses exhibited a statistically significant, 2-fold increase in average left and right ventricular volumes compared with the ventricular volume of control fetuses, with OCT-derived measures of 0.38 and 0.18 mm3, respectively, whereas the boundaries of the fetal mouse lateral ventricles were not clearly definable with US imaging. Our results indicate that OCT is a useful technology for assessing ventriculomegaly accompanying alcohol-induced developmental delay. This study clearly demonstrated advantages of using OCT for quantitative assessment of embryonic development compared with US imaging.

Whole body insulin resistance in rat offspring of mothers consuming alcohol during pregnancy or lactation: comparing prenatal and postnatal exposure.

This study examined the effects of maternal ethanol (EtOH) consumption during pregnancy or lactation on glucose homeostasis in the adult rat offspring. Glucose disposal was determined by minimal model during an intravenous glucose tolerance test in rats that had a small or normal birth weight after EtOH exposure in utero and in rats whose mothers were given EtOH during lactation only. All three EtOH groups had decreased glucose tolerance index and insulin sensitivity index, but their glucose effectiveness was not different from that of controls. In addition, EtOH rat offspring that were small at birth had elevated plasma, liver, and muscle triglyceride levels. The data show that EtOH exposure during pregnancy programs the body to insulin resistance later in life, regardless of birth weight, but that this effect also results in dyslipidemia in growth-restricted rats. In addition, insulin resistance is also evident after EtOH exposure during lactation.

Effects of maternal ethanol consumption during pregnancy or lactation on intestinal absorption of folic acid in suckling rats

A fostering/crossfostering analysis of the effects of maternal ethanol exposure on jejunal and ileal folate absorption was performed. Male and female rats were randomized into two groups. In the first group, ethanol-treated rats received ad libitum 5, 10 and 15% ethanol in the drinking fluid during three successive weeks. A consumption of 20% was maintained in this group for 5 additional weeks. Ethanol-treated rats were mated. Group 2 served as the control. To study the effect of chronic alcoholism during lactation or gestation separately, at birth (2nd day postpartum) control new-borns were cross-fostered to ethanol dams (EG), and the pups issued from the ethanol treated mothers were cross-fostered to control dams (CG). Thus, three experimental groups of pups were formed: (1) control pups receiving no treatment during gestation and lactation (CG); (2) pups exposed to ethanol only during gestation (GG); and (3) pups exposed to ethanol only during lactation (LG). At 21 days postpartum the jejunal and distal ileum folate absorption was determined in the offspring rats by a perfusion technique. Milk folic acid levels were determined by an immunoluminometric assay. The results showed an increase in jejunal folic acid absorption in offsprings exposed to ethanol only during the lactation period (LG). However, in pups exposed to ethanol only during the gestation period (GG), the jejunal folic acid absorption was significantly increased only at concentrations of 0.25, 0.5 and 2.5 μM. No free folic acid absorption occurred in the distal ileum of control pups (CG) at day 21 at all assayed concentrations but in offsprings exposed to ethanol only during the gestation or lactation periods absorption did take place. Pups exposed to ethanol during the gestation period (GG) showed decreased values in ileum folic acid absorption at the lowest assayed concentration (0.25 μM) compared to values obtained for pups exposed to ethanol only during lactation (LG). Milk folic acid levels were significantly decreased in the ethanol-fed dams on day 21 of lactation. These results indicate that exposure of rats to ethanol during the lactation period affects more severely postnatal development of intestinal functions than ethanol exposure only during gestation. In summary, both the exposure to ethanol itself and the decrease in folic acid intake caused alterations in the function of the intestinal mucosa in the offspring, which in turn altered absorption time and development. However, the present results do not explain how ethanol stimulated intestinal absorption of folic acid in pups exposed to ethanol during the gestation or lactation periods. Further studies are needed.

Effects of chronic maternal ethanol exposure on hippocampal and striatal morphology in offspring.

Astrocytes and serotoninergic neurons play a role in central nervous system (CNS) development, probably through serotonin (5HT) stimulation of the glial 5HT(1A) receptor. Activation of 5HT(1A) receptors causes the release of S-100 beta, a glial derived growth factor. In vitro, astrocytes are profoundly altered by chronic maternal ethanol exposure (CMEE). CMEE is also associated with reduced 5HT brain levels and abnormal development of the serotoninergic system. In the present study we analyzed the hippocampal and striatal serotoninergic innervation and astroglial cells in the offspring of CMEE mothers. Female Wistar rats were orally exposed to ethanol 6.6% (v/v) ad libitum for 6 weeks before breeding and during gestation. After parturition, rat mothers continued receiving ethanol until pups reached 21 days old. The control group received water ad libitum. Rat offspring brains were processed by immunocytochemistry using antibodies directed to GFAP, serotonin transporter (5HTT), or S-100 beta protein. Hippocampus and striatum were studied by computer-assisted image analysis. Cell area of GFAP(+) astrocytes, surface of 5HTT(+) fibers per area unit, and relative optical density (ROD) of S-100 beta(+) astrocytes were measured and statistically processed. Our results show that astroglial GFAP was increased (astrocytes were hypertrophied) and 5HTT(+) fibers were increased in both the hippocampal CA-1 area and the striatum. On the other hand, S-100 beta ROD was increased only in the hippocampal CA-1 area but not in the striatum. The different response of the studied regions is an interesting result considering evidence of a close 5HT/astroglial relation during CNS development. These differences could be due to different gradients of development in the studied areas and/or different responses of those areas to the effect of maternal ethanol exposure since the first stages of embryonic development.

Effects of maternal chronic alcohol administration in the rat: lactation performance and pup's growth.

A fostering/crossfostering analysis of the effects of maternal ethanol exposure on lactation performance and offspring growth was performed. Wistar rats were kept under one of the three experimental nutritional treatments: alcohol-treated (EG), pair-fed-treated (PFG) (as a nutritional control of alcohol-associated malnutrition), and control or normal diet (CG). Rats from the EG group were accustomed to increased amounts of ethanol (5% during the first week to 20% in the fourth week). The 20% ethanol level was maintained throughout three additional weeks and during gestational and lactational period. Daily food intake, fluid consumption, body weight and gestational parameters were studied in control (CG), pair-fed (PFG) and ethanol dams (EG). At birth, half the litters were fostered to other dams of the same treatment (GLG) and half were cross-fostered to dams of the opposite treatment (GG, LG). No cross-fostering analyses were performed on the pair-fed group. Offspring body weight was controlled throughout lactation. Liver, kidney and spleen weights as well as milk consumption were also studied at the end of lactation period. In dams, a significant reduction of body weight was described throughout the suckling period. No ethanol detrimental effects were observed on body weight at birth, but in spite of a normal birth weight, alcohol during lactation was responsible for a growth deficit. Milk consumption was significantly reduced in offspring exposed to ethanol during gestation and/or lactation. Curiously, prenatal alcohol exposure affects adversely the suckling behaviour in pups at the time of weaning. In our study, alcohol treatment and malnutrition affects liver and spleen weights. However, malnutrition decreases spleen weights more than alcohol treatment. In the case of the kidney weights the alcohol decreases kidney weight more than malnutrition. Collectively, the data from the present study show similar effects following pre/postnatal and postnatal alcohol exposure. The findings suggest that chronic alcohol administration during gestation and/or lactation adversely affects pup growth at weaning as indicated by its effect on milk consumption, pup and organ weight.

The effects of maternal ethanol exposure on neurotransmission and second messenger systems: a quantitative autoradiographic study in the rat brain

The effects of maternal ethanol exposure on neurotransmission and second messenger systems were examined in rats using histochemistry and in vitro autoradiography. Thirty % ethanol was administered to pregnant rats from gestational day 7 to the day of delivery. Quantitative autoradiography was used to map muscarinic cholinergic, dopamine D 2, adenosine A 1 and inositol 1,4,5-trisphosphate binding sites, as well as to localize adenylate cyclase and protein kinase C. We found no difference in the patterns of staining with acetylcholinesterase and Timm's stain between control and prenatally ethanol-exposed rats on postnatal day (PN) 30. In the ethanol-exposed rats, [ 3H]forskolin binding sites were increased during early development in the CA1 subfield of the hippocampus and the occipital cortex; [ 3H]phorbol ester binding sites were increased in the cortex, striatum, and hippocampus; hippocampal muscarinic cholinergic sites were increased on PN4 and 30; adenosine A 1 binding was reduced on PN10 in most regions examined, but was increased in the CA1 subfield on PN30; dopamine D 2 receptor levels were significantly reduced on PN30 in the striatum; and IP 3 receptors were decreased in most regions studied, but particularly in the cerebellum. Thus, some of these changes were transient and others were long-lasting. Although histophathological abnormalities were minimal, the alterations of binding sites in the cerebellum (the coordination center) and in the hippocampus (related to memory and learning) that were detected may contribute to the behavioral and mental deterioration seen in the fetal alcohol syndrome.