Mice were tested for ethanol clearance by procedures adapted from [40] and described previously [41], [42] with a 0.01 mM limit of detection. Animals were habituated to the testing room for two hours. At Time 0, animals were injected with ethanol (3.6g/kg, i.p.). Approximately 20 µl retro-orbital sinus blood samples were taken at 15, 30, 90 and 180 minutes after ethanol injection from nine animals per genotype. In addition, three animals per genotype had a single blood sample taken at 180 minutes post-injection, to control for blood volume loss that could result from repeated blood sampling. Alternating eyes were sampled to minimize trauma and animals were euthanized following the procedure. Samples were centrifuged at 4°C and plasma blood ethanol concentraions determined using gas chromatography (5890A GC, Hewlett Packard). Briefly, 7 µl serum was sealed in a GC autosampler vial (National Scientific, Rockwood, TN) with 7 µl 0.05% n-propanol, as an external pippeting standard. Samples, in duplicate, were heated to 65°C for 20 min, agitated for 30 sec, and allowed to settle for 1 min prior to pressurizing for headspace extraction
