Effects of Alcohol on Nitric Oxide (NO) Synthesis and Superoxide Production in Human Brain Vascular Cells

Abstract

Oxidative stress may play an important role in impaired endothelial NO synthase (eNOS)‐dependent dilation of cerebral arterioles during chronic alcohol consumption. The purpose of this study was to examine the effects of acute (3 hours) and chronic (7 days) alcohol on NO synthesis and superoxide production in cultured human brain vascular cells. Primary human brain microvascular endothelial cells (HBMEC) and human brain vascular smooth muscle cells (HBVSMC) were exposed to alcohol (5 and 50 mM) for 3 hours or 7 days. NO synthesis of HBMEC in response to L‐arginine and acetylcholine was measured by a NO microsensor. Superoxide production from HBMEC and HBVSMC was measured using lucigenin‐enhanced chemiluminescence. In addition, protein expression of eNOS in HBMEC was measured by Western blot analysis. Acute and chronic exposure of low dose alcohol did not alter NO synthesis and superoxide production. Acute exposure of high dose alcohol increased NO production in response to L‐arginine, but not acetylcholine. In contrast, chronic exposure of high dose alcohol significantly reduced NO production in response to L‐arginine and acetylcholine. In addition, chronic exposure of high dose alcohol significantly enhanced superoxide production in HBVSMC as well as in HBMEC. Furthermore, both acute and chronic exposure of high dose alcohol increased eNOS protein expression. Our findings suggest that chronic exposure of alcohol may reduce NO bioavailability via enhanced superoxide production in human brain vascular cells.