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Objective: In a model for fetal alcohol syndrome (FAS), we have previously found an alteration in NMDA receptors suggesting mediation, at least in part, of alcohol-related learning deficit. NMDA and GABA receptors interact in a multisynaptic circuit for the regulation of the inhibitory tone through the CNS. The GABA receptor subunit GABA Aα5 is involved in learning and is developmentally regulated, as it is excitatory in the perinatal brain and inhibitory in the adult. We were interested to evaluate alcohol's effect on GABA Aα5 expression to further understand alcohol-induced learning dysfunction. Study design: Timed, pregnant C57B16/Jmice were treated on gestational day 8 with alcohol (25% alcohol, 0.03 mL/kg ip) or control (saline). Embryos and brains were harvested 10 days after treatment, and brains from adult offspring were collected after evaluation in the Morris Water Maze, a well-established test for spatial learning. Gene expression included samples from at least 3 litters per timepoint, and calibrator-normalized relative real-time polymerase chain reaction (PCR) was performed to quantify GABA Aα5 with GAPDH standardization. Statistical analysis included analysis of variance (ANOVA). Results: Prenatal alcohol exposure significantly decreased GABA Aα5 expression in the embryo (P < .02) and fetal brains (P < .01) 10 days after therapy. However, in adult brains GABA Aα5 expression was increased versus controls (P < .01). As previously demonstrated, prenatal alcohol exposure resulted in deficits in adults learning the Morris Water Maze with controls learning faster (P < .05). Conclusion: Prenatal alcohol exposure alters developmental GABA Aα5 expression. This may further explain the long-lasting damage of alcohol on learning skills. Both the alcohol-induced reduction in the GABA Aα5 subunit during development and up-regulation in adult brain may be related to learning deficits resulting in decreased learning potential caused by the developmental defect and an increased inhibition of learning resulting from increased expression as an adult. In combination with our previous findings, these suggest that alcohol-induced learning impairment is likely the result of alterations of both NMDA and GABA expression and function. [ABSTRACT FROM AUTHOR]

Objective: Vasoactive intestinal peptide (VIP)-related peptides prevented the learning deficit in the offspring in a model for fetal alcohol syndrome. We evaluated whether the mechanism of the peptide protection included NR2B, NR2A, and GABAAα5. Study design: Timed, pregnant C57BL6/J mice were injected on gestational day 8 with alcohol (0.03 mL/kg), placebo, or alcohol plus peptides. Embryos were harvested after 6 hours, 24 hours, and on gestational day 18. Some of the litters were allowed to deliver, and the adult brains harvested after the offspring were tested for learning. Calibrator-normalized relative real-time polymerase chain reaction (PCR) was performed using primers for NR2B, NR2A, and GABAAα5 with GAPDH standardization. Statistic: analysis of variance (ANOVA) and Fisher PLSD, P < .05 was considered significant. Results: In the embryo, the peptides prevented NR2B rise (P < .001) at 6 hours, NR2B down- regulation (P = .002), and GABAAα5 decrease (P < .01) on gestational day 18. In the adult, the peptides prevented NR2B down-regulation (P = .01) and NR2A up-regulation (P < .001). Conclusion: VIP-related peptides prevented alcohol-induced changes in NR2B, NR2A, and GABAAα5. This may explain, at least in part, the peptides' prevention of alcohol-induced learning deficits. [ABSTRACT FROM AUTHOR]

Objective: Alcohol-related neurodevelopmental disorders are contributors to long-term learning disabilities. By using a model for fetal alcohol syndrome, we have shown that prenatal alcohol exposure results in adult learning deficits of unknown mechanisms. In the developing hippocampus, the N-methyl-D-aspartate (NMDA) receptor subunit NR2B triggers long-term potentiation, fundamental to learning and memory; this is supplemented by the less plastic NR2A subunit in the adult. To understand the mechanism of learning deficits in FAS, we evaluated NR2B and NR2A expression in embryonic and adult mice. Study design: Pregnant C57BI6/J mice were treated on gestational day 8 with alcohol or control (saline solution). Embryos were harvested at 6 hours, 24 hours, and 10 days, and brains from adult offspring were collected at 3 months (after evaluation for learning deficit). Calibrator-normalized relative real-time polymerase chain reaction was performed for NR2B and NR2A with glyceraldehyde-3-phosphate dehydrogenase standardization. Statistical analysis included analysis of variance. Results: At 6 hours, NR2B expression in the alcohol-exposed embryos was higher than in controls (P « .01). NR2A was not expressed in either group. By 24 hours there was no difference in NR2B (P .3). However, at 10 days NR2B was lower in alcohol-exposed animals (P = .02). In the adult brains there was a relative decrease in NR2B (P = .03) and an increase in NR2A (P « .01). Conclusion: Prenatal alcohol exposure during development induces NR2B expression deregulation in the embryos that persists until adulthood, when a relative increase in the less modifiable subunit NR2A occurs. This alteration in NMDA receptor subunits may underlie the learning abnormalities in fetal alcohol syndrome. [ABSTRACT FROM AUTHOR]

Objective: Previously, the novel peptides NAPVSIPQ and SALLRSIPA were shown to prevent alcohol-induced fetal death and growth abnormalities in a mouse model of fetal alcohol syndrome. This study evaluated whether these peptides could prevent long-term alcohol-induced learning abnormalities. In addition, because specific cytokines are known to effect long-term potentiation, a model of learning at the molecular level, we studied the effect of these novel peptides on tumor necrosis factor-α, interleukin-6, and interferon-γ levels. Study design: We used a well-characterized mouse model of fetal alcohol syndrome. Pregnant mice were injected on day 8 with alcohol (0.03 mL/kg) or placebo. Pretreatment with NAPVSIPQ + SALLRSIPA (20 µg) or placebo was given 30 minutes before alcohol. Embryos were removed after 6 hours, at which time cytokine, tumor necrosis factor-α, interleukin-6, and interferon-γ levels were measured with enzyme-linked immunoassays. To test spatial learning, adult offspring from litters that were treated with alcohol, control, NAPVSIPQ + SALLRSIPA then alcohol, or NAPVSIPQ + SALLRSIPA alone were evaluated for latency to find a hidden platform in the Morris water maze. Results: Alcohol treatment increased tumor necrosis factor-α levels versus control levels (50.0 ± 3.5 pg/mL vs 32.7 ± 2.4 pg/mL; P < .001). NAPVSIPQ + SALLRSIPA pretreatment prevented this increase (39.9 9 ± 2.8 pg/mL; P ⩽ .01), with levels similar to control (P = .1). Similarly, alcohol increased interleukin-6 levels versus control levels (22.6 ± 1.4 pg/mL vs 17.3 ± 0.6 pg/mL; P < .001), and NAPVSIPQ + SALLRSIPA prevented this increase (19.1 ± 1.0 pg/mL; P ⩽ .02), with levels similar to control levels (P = .2). Interferon-γ levels were not different among the 3 groups (alcohol, 14.6 ± 4.9 pg/mL; control, 17.9 ± 6.6 pg/mL; alcohol + NAPVSIPQ + SALLRSIPA, 13.6 ± 4.9 pg/mL; P = .2). In the Morris water maze, alcohol-treated groups did not learn over the 7-day trial compared with the control group (P = .001). Groups that were pretreated with NAPVSIPQ + SALLRSIPA then alcohol learned significantly, which was similar to the control group. Groups that were treated with only NAPVSIPQ + SALLRSIPA learned significantly earlier, with the shortest latency once learning commenced. Conclusion: The peptides, NAPVSIPQ + SALLRSIPA, prevented the alcohol-induced spatial learning deficits and attenuated alcohol-induced proinflammatory cytokine increase in a model of fetal alcohol syndrome. This study demonstrates the peptides' significant in vivo efficacy with long-lasting effects obtained after prenatal administration. [ABSTRACT FROM AUTHOR]

Objective: Fetal alcohol syndrome (FAS) is the most common nongenetic cause of mental retardation. Peptides NAPVSIPQ (NAP) and SALLRSIPA (SAL), related to activity-dependent neuroprotective protein (ADNP), prevent alcohol-induced damage in a mouse model of FAS. Our objective was to characterize ADNP in this model to relate this protein to the mechanisms of damage and peptide neuroprotection.Study Design: Timed, pregnant C57Bl6/J mice were treated on day 8. Groups were control, alcohol, peptide pretreatment, or peptide alone. Embryo and decidua were harvested at 6 and 24 hours and 10 days. To evaluate ADNP expression, real-time polymerase chain reaction was performed with results presented as the ratio of ADNP-to-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) concentration. Analysis of variance was performed for overall comparisons with P<.05 considered significant.Results: At 6 hours, there was no difference in ADNP between alcohol-exposed embryos compared with control embryos. At 24 hours, there was an increase in ADNP in alcohol-exposed embryos compared with controls (P<.001); these findings persisted at 10 days (P<.001). In the decidua at 6 hours, there was no difference between alcohol and control. At 24 hours, there was greater ADNP in alcohol-exposed decidua compared with controls (P<.001), which did not persist at 10 days (P=.97). Peptide pretreatment did not prevent the alcohol-induced increase in ADNP in embryo or decidua.Conclusion: Alcohol increased embryonic and decidual ADNP expression at 24 hours and it persisted in the embryo for 10 days. Because ADNP is a known neuroprotectant, these findings suggest that it may be released as a protective mechanism in FAS. Changes in the embryo were persistent suggesting that the embryo is more vulnerable to alcohol-induced damage than the mother. [ABSTRACT FROM AUTHOR]

Objective: Fetal alcohol syndrome (FAS) results in stillbirth, fetal growth restriction, and mental retardation with injury attributed to oxidative stress. Our objective was to identify signal transduction pathways expressed in a model of FAS and to quantify expression of c-fos, a gene in the stress signal pathway.Study Design: Timed, pregnant C57Bl6/J mice were injected on E8 with saline solution or alcohol. RNA was extracted from decidua and embryo 6 and 24 hours later. Microarray analysis was used to screen gene pathways. Differential gene expression was confirmed using real-time polymerase chain reaction with results presented as the ratio of c-fos concentration to that of glyceraldehyde-3-phosphate dehydrogenase (GAPDH).Results: Differential gene expression between alcohol and control was noted for stress signal pathway genes including c-fos. Real-time polymerase chain reaction demonstrated that c-fos messenger RNA expression was greater in the alcohol than control decidua at 6 hours after injection (P<.01). This effect persisted at 24 hours (P<.01). There was no difference in c-fos expression in embryos whose mothers received alcohol versus control after 6 hours (P=.12) or 24 hours (P=.89).Conclusion: Alcohol administration during pregnancy results in differential gene expression in the stress signal pathway, particularly in c-fos. C-fos expression in the decidua increases from 6 to 24 hours after alcohol injection, but does not change in the embryo, which may contribute to alcohol-induced damage in FAS. [ABSTRACT FROM AUTHOR]

Using a mouse model for fetal alcohol syndrome, we examined whether 2 neurotrophic peptides protect against alcohol-induced alteration in the 7-aminobutyric acid A receptor subunit GABA-β3. [ABSTRACT FROM PUBLISHER]

Objective: Fetal alcohol syndrome (FAS) is associated with intellectual disability and neurodevelopmental abnormalities. Neuroprotective peptides NAPVSIPQ (NAP) and SALLRSIPA (SAL) can prevent some of the alcohol-induced teratogenesis including fetal death, growth abnormalities, and learning impairment in part by preventing alcohol-induced alterations in N-methyl-D-aspartate receptor gene expression in a mouse model for FAS. We evaluated a panel of cytokines and chemokines to determine whether NAP plus SAL work through a cytokine/chemokine-mediated pathway in preventing these alterations. Study Design: Using a well-characterized FAS model, timed, pregnant C57BL6/J mice were treated on gestational day (E) 8 with alcohol (0.03 mL/g), placebo, or alcohol plus peptides. Embryos were evaluated at 2 time points: after 6 hours and 10 days later at E18. A panel of cytokines/chemokines was measured using a microsphere-based multiplex immunoassay (Luminex xMAP; Millipore, Billerica, MA). Statistical analysis included Kruskal-Wallis, with P < .05 considered significant. Results: Six hours after treatment, interleukin (IL)-6 and keratinocyte chemoattractant cytokine (KC) were not detectable in the control embryos. Alcohol treatment resulted in detectable levels and significant increases in IL-6 (median, 15.7; range, 10.1–45.9 pg/mL) and KC (median, 45.9; range, 32.5–99.1 pg/mL). Embryos exposed to alcohol plus NAP plus SAL had undetectable IL-6 and KC (both P < .003), similar to the controls. Alcohol exposure resulted in a significant increase of granulocyte colony–stimulating factor (G-CSF) (P < .003) as compared with controls, and treatment with NAP plus SAL prevented the alcohol-induced increase. IL-13 and IL-1β were decreased 6 hours after alcohol exposure, and exposure to alcohol plus NAP plus SAL did not completely ameliorate the decrease. At E18, 10 days after exposure, these alterations were no longer present. Several analytes (regulated upon activation, normal T cell expressed, and secreted, tumor necrosis factor-α, interferon-γ, and IL-4) were not detectable at either time point in any of the groups. Conclusion: Prenatal alcohol exposure acutely results in a significant elevation of IL-6, G-CSF and the KC, which are known to affect N-methyl-D-aspartate receptors. NAP plus SAL treatment prevented alcohol-induced increases. This provides additional insight into the mechanism of alcohol damage in FAS and NAP plus SAL prevention of neurodevelopmental anomalies. [Copyright &y& Elsevier]

Objective: Our objective was to evaluate whether brain-derived neurotrophic factor (BDNF) expression is affected by prenatal alcohol exposure and whether the neuroprotective effects of the vasoactive intestinal peptide (VIP)-related peptides, NAPVSIPQ (NAP) and SALLRSIPA (SAL), are mediated through BDNF. Study Design: Using a well-characterized fetal alcohol syndrome (FAS) model, timed pregnant C57BL6/J mice were treated on gestational day (E) 8 with alcohol (0.03 mL/g), placebo, or alcohol plus (NAP plus SAL). Embryos were harvested at 6 hours (E8), 24 hours (E9), and 10 days (E18) and pups at postnatal day 40. Calibrator-normalized relative real time polymerase chain reaction was performed to quantify BDNF with hypoxanthine phosphoribosyl transferase-1 standardization. Results: BDNF expression was lower in the alcohol-exposed embryos than in controls at 6 hours and higher at 24 hours and 10 days (all P < .05). Pretreatment with NAP plus SAL prevented the alcohol-induced rise in BDNF expression (P < .05) at 24 hours and 10 days after alcohol exposure. We found no difference between alcohol and control in young-adults'' brain (P > .05). Conclusion: NAP plus SAL treatment prevented alcohol-induced changes in BDNF expression 24 hours and 10 days after alcohol exposure in mouse embryos. This may explain, at least in part, the peptides'' prevention of neurodevelopmental anomalies in FAS. [Copyright &y& Elsevier]

Objective: Fetal alcohol syndrome (FAS) is the leading cause of a spectrum of preventable nongenetic learning and behavioral disorders. In adult (FAS) mice, we measured phosphorylated cyclic adenosine monophosphate response element of binding protein (pCREB) staining in hippocampal subregions to evaluate a possible mechanism underlying FAS learning deficits. Study Design: Pregnant C57BL6/J mice were treated on gestational day 8 with alcohol or control (saline). After learning assessment, the offspring were perfused for immunohistochemistry and brain sections probed using SER 133 pCREB antibody. Relative staining density was assessed using National Institutes of Health Image software. Statistical analysis included analysis of variance with P < .05 considered significant. Results: In all hippocampal subregions, pCREB staining was greater in the control animals than in the alcohol-treated group (P ≤ .0001). Conclusion: In utero alcohol exposure decreased pCREB activity in hippocampal subregions of adult mice. The dentate gyrus had the most robust cumulative decrease in pCREB staining, suggesting FAS adult learning deficits may correlate to enhanced dentate gyrus neurodegeneration. [Copyright &y& Elsevier]