Ethanol enhances basal and flow-stimulated nitric oxide synthase activity in vitro by activating an inhibitory guanine nucleotide binding protein

The aim of this study was to determine the effect of ethanol on endothelial nitric oxide synthase (eNOS), the enzyme responsible for the production of the important vasoactive agent nitric oxide. The effect of ethanol (0.8-16 0 mM) on both basal and flow-stimulated eNOS activity was determined using cultured bovine aortic endothelial cells (EC), In "static" EC ethanol dose- dependently increased basal eNOS activity with a maximum response (similar to 2.0-fold increase) achieved at 40 mM in the absence of any effect on cel l viability or nitric oxide synthase protein expression. Pertussis toxin (P TX) pretreatment significantly inhibited the ethanol-induced increase in ba sal eNOS activity. EC exposed to steady laminar flow exhibited a flow- and time-dependent increase in eNOS activity. Ethanol significantly enhanced th e laminar flow-induced eNOS response from 0.62 +/- 0.1 to 1.06 +/- 0.06 pmo l [C-14]citrulline/mg/min, a response that was inhibited by PTX, PTX-cataly zed ribosylation of Gi alpha substrates, an index of G-protein functional a ctivity, was increased in laminar flow-exposed EC compared with static cont rols and was further enhanced by ethanol treatment. Likewise, EC exposed to low (similar to 0.5 dynes/cm(2)) and high (similar to 12 dynes/cm(2)) puls atile flow demonstrated increased eNOS activity, an effect that was associa ted with increased PTX-catalyzed ribosylation of Gi alpha substrates, Ethan ol enhanced the low flow response in a PTX-sensitive manner. These data dem onstrate a stimulatory effect of ethanol on basal and flow-stimulated eNOS activity, mediated in part by a mechanism involving a PTX-sensitive G prote in.