Examining effects of one-day alcohol binge exposure on cell type-specific loss in the thalamic nucleus reuniens at various developmental time points in a rodent model of FASDs
Date
2024
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Publisher
University of Delaware
Abstract
Fetal Alcohol Spectrum Disorders (FASDs) is an umbrella term used to describe multiple developmental conditions stemming from prenatal alcohol exposure that manifest as deficits in growth development, cognitive impairments, and physical abnormalities. 2-5% of live births in the US are affected by FASD and result in lower brain volume, brain anomalies, and behavioral deficits including impaired executive function. Executive function (EF) defines a set of cognitive controls that aid in the formation of goal-directed movements and regulating self-control; EF has been shown to relate to medial prefrontal cortex (mPFC) and hippocampus (HPC) activity. An intermediary structure, the nucleus reuniens (Re) of the midline thalamus, facilitates communication between the mPFC and HPC and is known to be damaged by prenatal alcohol exposure. Behavioral studies in rodents indicate Re damage produces deficits in EF due to inefficient communication in the mPFC-Re-HPC circuit (Dollerman-van der Weel and Witter, 2020). Most research on Re follows an animal model as the brain connections of Re in the human brain are located in the massa intermedia, however, some humans lack the massa intermedia altogether (Dollerman-van der Weel and Witter, 2020; Rioch, 1931; Droogleever and Stefens., 1951). ☐ This study employed a rodent model of third-trimester single-day binge alcohol exposure (AE) to evaluate the neuroanatomical effects of neonatal alcohol exposure at three time points. Specifically, we examined moderate (3g/kg/day), and high (5.25g/kg/day) doses of ethanol administered on postnatal day (PD) 9 compared to a sham-intubated control group. In conjunction with histological immunofluorescence staining, this study measured the numbers of specific cell populations in Re to assess levels of cell loss at three time points after AE: 12 hours, 96 hours, and 56 days. We estimated the total number of neurons and oligodendrocytes in Re by labeling these cells with antibodies against markers NeuN (neuronal marker) and CC1 (marker for mature oligodendrocytes). ☐ Analysis of NeuN+ cells in Re on PD9 revealed a significant interaction between sex and postnatal treatment. Post-hoc analysis revealed a significant main effect of postnatal treatment on the NeuN+ cell number in female animals but not males, where AEM females had significantly more neurons in Re than SI females. AEH females had significantly fewer neurons than AEM females. No effect of sex on any parameters of the study were found when looking at CC1+ cells in Re so a One-Way ANOVA analysis was run and revealed no significant effect of postnatal treatment on the number of CC1+ cells. No significant difference in Re volume was observed between postnatal treatment groups. ☐ Statistical analysis of tissue collected on PD13 showed a significant interaction between sex and postnatal treatment effect on NeuN+ neuron counts. AEH females had significantly fewer neurons than SI females. There was no significant difference in neuron number between AEM and AEH females or SI and AEM females. For males, significant differences between SI and AEM and SI and AEH were observed. Both alcohol groups demonstrated significantly less neurons compared to SI animals. No significant difference in neuron number was observed when comparing AEM and AEH groups. Examination of CC1+ cells in Re demonstrated significantly less oligodendrocytes to be present in AEH females than AEM females. Males displayed significantly less oligodendrocytes in both AEM and AEH groups when compared to SI groups. There was no significant difference in Re volume at this time point. ☐ Two-Way ANOVA revealed a significant effect of postnatal treatment on neuron population in Re on PD65. Regardless of sex, AEM groups had significantly less neurons compared to SI groups. It was also found that AEH groups had significantly fewer neurons compared to SI groups. A One-Way ANOVA analysis of oligodendrocyte population in Re revealed no significant differences. We also saw now significant difference in Re volume at this time point. ☐ This study further illustrates the teratogenic effects of alcohol by demonstrating that a single day of alcohol exposure is sufficient to produce life-long alterations in cell population in Re. Not only is a single binge exposure detrimental to Re, but varying doses (moderate and high) have proven to hold deleterious effects on neuron and oligodendrocytes, specifically at PD13. Observations noting dose-dependent changes in cell number allows us to state less alcohol intake does not prevent the drinking-related damage on the developing brain. Gurksy and colleagues in 2022 used a similar single-day dosing paradigm but on PD7. Comparing our current results with theirs, we conclude that a single day of alcohol exposure on PD9 produces similar reductions in neuron number. However, PD7 is a more vulnerable timepoint due to significantly more pronounced reductions in all cell types and reduced Re volume.
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Keywords
Alcohol exposure, Fetal alcohol spectrum disorder, Neurons, Oligodendrocytes, Reuniens