DESCRIPTION:
Adult male C57BL/6J mice were purchased from Jackson Laboratories (Bar Harbor, ME, USA) at 10 weeks of age. Mice were kept under a 12-hour light/dark cycle and given free access to water and standard rodent chow (Harland, Teklad, Madison, WI). Studies were designed to determine genomic responses and interactions between two different ethanol exposure models: intermittent cycles of ethanol vapor exposure in inhalation chambers (CIE), and oral consumption of 15% (v/v) ethanol in a limited access (2 h/session) paradigm. Mice were divided into 4 treatment groups (n = 12/group): the “CIE Drinking” group received inhaled ethanol in the vapor chambers followed by 2-bottle choice ethanol drinking in between vapor exposure cycles; the “Air Drinking” group received only air in the vapor chambers, but had 2-bottle choice ethanol drinking between CIE cycles; the “CIE NonDrinking” group received inhaled ethanol in the vapor chambers but only water access in between CIE cycles; and the “Air NonDrinking” group remained ethanol naïve with air exposure in vapor chambers and only water consumption between CIE cycles. Following a 2-week acclimation period, mice in the CIE Drinking and Air Drinking groups underwent 6-weeks of 2-bottle choice drinking to establish baseline drinking levels. The 2-hr limited access session to ethanol (15 v/v) vs. water started 30 min before lights off. Ethanol and water intake for each individual mouse was measured daily. Following 6-weeks of baseline drinking, mice were placed in Plexiglass inhalation chambers (60x36x60 cm) 16 hours/day for 4 days. Ethanol was volatilized with an air stone submerged in 95% ethanol. Vapor chamber ethanol concentrations were monitored daily and air flow was adjusted to ethanol concentrations within 10–13 mg/l air. This ethanol vapor concentration has been shown to yield stable blood ethanol concentrations (175–225 mg/dL) in C57BL/6J mice [25]. Before each vapor chamber session, intoxication was initiated in the CIE group by administration of 1.6 g/kg ethanol and 1 mmol/kg pyrazole intraperitoneally (i.p.) at a volume of 0.02 ml/g body weight. Pyrazole is an alcohol dehydrogenase inhibitor used to stabilize blood ethanol concentrations. All mice received the same number and timing of pyrazole injections prior to final removal from the inhalation chambers with control mice receiving saline and pyrazole (i.p.), also at a volume of 0.02 ml/g body weight, prior to being placed into control vapor chambers. Control vapor chambers delivered only air without ethanol vapor. After 4 days in the inhalation chambers, mice underwent a 72-hour period of total abstinence from ethanol. Following the abstinence period, mice in the CIE Drinking and Air Drinking groups were given 2-bottle choice drinking for 2 hours per day for 5 days. A total of 4 cycles of CIE-abstinence-drinking were performed. After the end of the 4th cycle mice were sacrificed on the 5th drinking day before receiving ethanol/water access on that day at the time they received 2-bottle choice drinking all previous drinking days (Fig 1). Affymetrix GeneChip® Mouse Genome 430, type 2 arrays were analyzed with The R Project for Statistical Computing (http://www.r-project.org/). Statistical analysis to identify significantly regulated genes was performed using the limma package for R. False discovery rates equal to or less than 0.01 were considered significant.
LABEL:
DEG C57BL/6J HPC CIE (drinkers) vs air (nondrinkers) FDR < 0.01 (gene symbol)_qvalue
SCORE TYPE:
Q-Value
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