For the microarray experiment, 2 × 106 LCLs from each of 21 alcoholics and 21 non-alcoholics were seeded in 10 ml of RPMI1640 medium supplemented with 15% FBS, 2 mM glutamine, 50 U/ml penicillin, and 50 μg/ml streptomycin. Cultures were maintained in tightly capped flasks to minimize evaporation. Alcoholics were defined as meeting DSM-IV criteria for alcohol dependence (American Psychiatric Association, 1994) at age 18 years or younger. Non-alcoholics were defined as having taken at least one drink of alcohol and not meeting any of four definitions of alcohol dependence: DSM-IV (American Psychiatric Association 1994), DSM-IIIR (American Psychiatric Association, 1987), ICD-10 (World Health Organization, 1993), or Feighner definite alcoholism (Feighner, 1972); none were dependent on any illicit drug. Each phenotypic group (alcoholic or non-alcoholic) contained 12 males and 9 females. Growth of ethanol treated and untreated cells was parallel by 22 h even up to 100 mM ethanol; we chose 75 mM to be within this range and to offer a good possibility of discerning effects. Cells were cultured in the absence or presence of 75 mM ethanol for 24
