The Effect of Chronic Ethanol Exposure and Thiamine Deficiency on Myelin-related Genes in the Cortex and the Cerebellum

Abstract

Background: Long-term alcohol consumption has been linked to structural and functional brain abnormalities. Furthermore, with persistent exposure to ethanol (EtOH), nutrient deficiencies often develop. Thiamine deficiency is a key contributor to alcohol-related brain damage and is suspected to contribute to white matter pathology. The expression of genes encoding myelin proteins in several cortical brain regions is altered with EtOH exposure. However, there is limited research regarding the impact of thiamine deficiency on myelin dysfunction.

Methods: A rat model was used to assess the impact of moderate chronic EtOH exposure (CET; 20% EtOH in drinking water for 1 or 6 months), pyrithiamine-induced thiamine deficiency treatment (PTD), both conditions combined (CET-PTD), or CET with thiamine injections (CET + T) on myelin-related gene expression (Olig1, Olig2, MBP, MAG, and MOG) in the frontal and parietal cortices and the cerebellum.

Results: The CET-PTD treatments caused the greatest suppression in myelin-related genes in the cortex. Specifically, the parietal cortex was the region that was most susceptible to PTD-CET-induced alterations in myelin-related genes. In addition, PTD treatment, with and without CET, caused minor fluctuations in the expression of several myelin-related genes in the frontal cortex. In contrast, CET alone and PTD alone suppressed several myelin-related genes in the cerebellum. Regardless of the region, there was significant recovery of myelin-related genes with extended abstinence and/or thiamine restoration.

Conclusion: Moderate chronic EtOH alone had a minor effect on the suppression of myelin-related genes in the cortex; however, when combined with thiamine deficiency, the reduction was amplified. There was a suppression of myelin-related genes following long-term EtOH and thiamine deficiency in the cerebellum. However, the suppression in the myelin-related genes mostly occurred 24 h after EtOH removal or following thiamine restoration; within 3 weeks of abstinence or thiamine recovery, gene expression rebounded.

Keywords: Alcohol; Cerebellum; Cortex; Thiamine Deficiency; White Matter.

Figure 1.

A. Experimental timeline for the subjects of Experiments 1 and 2. In Experiment 1, the subjects were assigned to one of four groups—PF (Pair Fed control), CET (Chronic EtOH Treatment), CET-PTD (CET combined with Pyrithiamine-Induced Thiamine Deficiency), or PTD (Pyrithiamine-Induced Thiamine Deficiency) *—which were reversed after the loss of the righting reflex but prior to seizure. In Experiment 2, subjects were assigned at random to one of six groups; PF control, CET, CET-PTD, CET+T (Chronic ethanol treatment with injections of thiamine), PTD-EAS (PTD – Early acute stage) in which subjects were reversed 1h after seizure, and PTD-MAS (PTD – Moderate acute stage)in which subjects were reversed 4h after seizure. Brain samples (frontal cortex [FC], parietal cortex [PC] and cerebellum [Cer]) were collected in different time-points: T0=during treatment in Experiment 1; T1=during treatment in Experiment 2; T2=24-h post-treatment; T3=three weeks post-treatment). B. BECs (mean ± SEM) following one month of treatment for Experiment 1. This time point signifies the end of the CET treatment for Experiment 1. C. BECs (mean ± SEM) for Experiment 2. BECs were collected after one month, three months (halfway through the CET treatment), and six months (at the conclusion of the CET treatment). *NOTE: The PTD subjects in Experiment 1 most resemble the PTD-EAS subjects of Experiment 2.

Figure 2.

A. Four steps of oligodendrocyte maturation—oligodendrocyte precursor cells, preoligodendrocytes, immature oligodendrocytes, mature oligodendrocytes—adapted from Barateiro and Fernandes (2014). During the stages of oligodendrocyte maturation, cells express different genes, which act as useful markers and indicate the cells’ current stage of maturation. B. Heat map of gene expression for Experiment 1 in the frontal cortex, the parietal cortex, and the cerebellum. Heat map of gene expression of the myelin-related genes, Olig1, Olig2, MBP, MAG, and MOG, in the frontal cortex, the parietal cortex, and the cerebellum. Gene expression was relative to the PF group (% of PF expression). The standard error of the mean is represented in parentheses. Post hoc contrasts were used to compare the PF group to the experimental conditions. Differences in gene expression are represented by color with the red schema representing an increase in expression relative to PF and the blue schema representing a decrease relative to PF (p<0.05). The darker shades of red (increase) or blue (decrease) represents p<0.01, whereas the lighter shades represents a p-value between 0.02 and 0.01.

Figure 3.

Heat map of gene expression for Experiment 2 in the frontal cortex, the parietal cortex, and the cerebellum. Heat map of gene expression of the myelin-related genes, Olig1, Olig2, MBP, MAG, and MOG, in the frontal cortex, the parietal cortex, and the cerebellum. Gene expression for the treatment effect is relative to the PF group (% of PF expression for T1, T2, and T3). A statistical difference was apparent in the PF group at T2; therefore, an ultimate PF control could not be used. For MBP, the expression levels in the cerebellum were compared to their respective time points. Gene expression for the recovery effect compared gene levels within an experimental condition across three time points, (T1=during treatment; T2=24-h post-treatment; T3=three weeks post-treatment). The standard error of the mean is represented in parentheses. Post hoc contrasts were used to compare the PF group to the experimental conditions and the different time points within each condition. Differences in gene expression are represented by color with the red schema representing an increase in expression relative to PF and the blue schema representing a decrease relative to PF. The darker shade of red or blue represents p<0.01, whereas the lighter shade represents a p-value between 0.02 and 0.01. * represents an increase from T2 relative to T1, ^ represents an increase at T3 relative to T1, & represents an increase at T3 relative to T2 when assessing the effect of recovery, # represents a decrease at T2 relative to T1, @ represents a decrease at T3 relative to T1, and % represents a decrease at T3 relative to T2 when assessing the effect of recovery (p<0.02).