We next evaluated the catalytic activity, protein content, and expression of the EtOH-metabolizing enzymes, CYP2E1 or ADH, in primary human neurons after exposure to 17.5 mM EtOH (corresponding to 0.08%, legal alcohol limit in blood) for 24 h with/without 4-methylpyrazole (4-MP, inhibitor of CYP2E1/ADH). EtOH exposure resulted in a 2.8-fold induction of CYP2E1 activity in human neurons (Fig. 1A) paralleling a 1.5-fold increase in CYP2E1 protein content (Figs. 1B and C) or an enhanced CYP2E1 protein expression/localization (Figs. 1D and E) compared with respective controls. The magnitude of CYP2E activity induction by EtOH in human neurons (62 nmol/mg protein) was lower than that of human brain endothelial cells (200 nmol/mg protein). Although EtOH did not significantly induce ADH activity or protein content, it is important to note that ADH activity (>9 nmol/mg), protein content, and immunofluorescent staining for ADH expression were detected in primary human neurons (Figs. 2A-E). These results suggested that human neurons could potentially metabolize EtOH via CYP2E1/ADH, leading to reactive EtOH metabolite production.
