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An Inexpensive, Scalable Behavioral Assay for Measuring Ethanol Sedation Sensitivity and Rapid Tolerance in Drosophila
Published on: April 15, 2015
Strain-specific brain metallothionein II (MT-II) gene expression, its ethanol responsiveness, and association with
Kimberly D Loney1, K Raihan Uddin, Shiva M Singh
1Molecular Genetics Laboratories, Department of Biology and Division of Medical Genetics, University of Western Ontario, London, Ontario, Canada.
Background:
Metallothioneins (MTs) are ubiquitously expressed intracellular proteins that bind heavy metals such as zinc, copper, and cadmium. Although their specific function has yet to be discovered, they are known to regulate the metabolism of these metals as well as respond to cellular stress agents, particularly oxidants.
Methods:
Brain RNA from experimental (8 g/kg 25% ethanol injection) and control (saline injection) mice from four strains (A/J, BALB/cJ, C57BL/6J, DBA/2J) that are known to differ with respect to ethanol preference was used in differential displays. This report includes molecular results on one gene (MT-II) identified.
Results:
Our results on differential displays suggest that a proportion of genes are differentially expressed across pair-wise strain comparisons. We identified MT-II as a strain-specific and ethanol-responsive gene. The level of MT-II messenger RNA (mRNA) in control mice of A/J, BALB/cJ, C57BL/6J, and DBA/2J strains was variable (0.50, 0.51, 0.90, and 0.14 times G3PDH expression, respectively). The degree of up-regulation in experimental mice was also somewhat variable among strains, ranging from 2.5 to 3.2 times expression over the matched controls. Experiments indicate that the promoter and genomic organization of the MT-II gene is identical in sequence for all four strains, and methylation studies revealed that the MT-II promoter region is unmethylated in the brains of these mice. Interestingly, MT-II expression in control mice demonstrated a positive correlation with the ethanol preference phenotype.
Conclusion:
An increase in MT-II mRNA levels after injection of ethanol is attributed to the antioxidant properties of MT-II. The differential mRNA levels of this gene among four strains are not accounted for by the genomic organization, DNA sequence, or methylation status of this gene. Furthermore, the observed correlation between MT-II mRNA levels and ethanol preference raises an interesting hypothesis about the possible role of MT-II in ethanol effects and preference in mice.
