Drinking Pattern in Intermittent Access Two-Bottle-Choice Paradigm in Male Wistar Rats Is Associated with Exon-Specific BDNF Expression in the Hippocampus During Early Abstinence.

Abstract

Outbred rats differentially consume alcohol when having free access to it. Among others, BDNF (brain-derived neurotrophic factor) is believed to control voluntary ethanol intake in rodents. Meanwhile, expression of BDNF exons in brain regions and epigenetic mechanisms underlying alcohol intake pattern remain obscure. The main goal was to study whether voluntary alcohol drinking pattern affects expression of BDNF exons in selected rat brain regions during early abstinence. Intermittent access to 20% ethanol in a two-bottle-choice procedure (IA2BC) was used as a model of voluntary ethanol intake. Male Wistar rats (n = 24) had twenty 24-h sessions of free access to two-bottle choice (water or 20% ethanol) with 24-h withdrawal periods (water only). Control animals had access to water only (n = 11). After finishing IA2BC, the animals were divided according to the compliance of ethanol intake pattern with gradual escalation, a key feature of the paradigm. To access potential behavioral disturbances during the early abstinence, rats were consequently tested in the open field test, the elevated plus-maze, and the sucrose preference test. On the third day after the last drinking session, expression of BDNF exons and polypeptide was measured in the frontal cortex, hippocampus, striatum, and midbrain using quantitative PCR and Western blotting, respectively. Additionally, chromatin immunoprecipitation was performed to analyze enrichment of positive Ph-CREB (Ser133) and negative EZH2 transcriptional regulators as well as markers of active H3K9ac and repressed H3K27me3 chromatin at exon-specific BDNF promoters in brain regions with affected BDNF expression. During the course of the IA2BC, one part of animals demonstrated gradual escalation from low to high alcohol intake and preference of alcohol over water (a typical pattern for IA2BC) while the other one consumed alcohol at a consistently high level (an unusual pattern for IA2BC). Drinking pattern in the IA2BC does not define differences of behavior in any of the tests during early abstinence. Finally, the IA2BC rats with growing alcohol intake showed elevation of BDNF mRNA containing exon VI in the hippocampus associated with an enhanced H3K9ac occupancy at the respective promoter. Thus, rats differentially consuming alcohol in the IA2BC paradigm differ in epigenetically determined expression of BDNF exon VI in the hippocampus during early abstinence.