The differential effects of acute and prolonged L-NAME exposure on A23187-induced NO-dependent dilation in human arterioles
Authors: Zinkevich Natalya , Drachuk Kostiantyn , Zhang David 
Published: May 2024

The Ca2+ ionophore A23187 induces endothelium-dependent and non-receptor-mediated vasodilation in human adipose arterioles (HAA). Preliminary data from our group suggest that while A23187 dilation mainly depends on NO as a vasodilator factor in healthy HAAs, there is a component of non-NO and likely hydrogen peroxide (H2O2)-mediated dilation, especially in arterioles from patients with coronary artery disease (CAD). The mechanisms underlying the release of two different vasodilator factors remain largely unclear. There is evidence that NO-mediated S-nitrosylation inhibits the catalytic activity of mitochondrial electron transport chain (ETC) complexes or other reactive oxygen species (ROS)-producing enzymes such as NADPH oxidases (NOX). The objective of this study was to compare the effects of acute (30 minutes) and prolonged (90 minutes) nitric oxide (NO) synthase inhibitor nitro-L-arginine methyl ester (L-NAME) exposure on A23187 dilation to elucidate whether prolonged incubation with L-NAME results in transition from NO-dependent to H2O2-dependent vasodilation in HAAs. HAAs (~150 μm ID), isolated from fat tissues obtained from subjects with and without CAD, were cannulated and pressurized at 60 mmHg for videomicroscopic measurement of vessel diameters. In non-CAD HAAs constricted with endothelin-1, A23187 (10−11 to 10−7 M) elicited concentration-dependent dilation (maximal dilation at 10−7 M: 68±5%, n=5), and this dilation was largely abolished after acute exposure to L-NAME (maximal dilation at 10−7 M: 21±7%, n=5). Prolonged L-NAME incubation restored A23187-dependent dilation (maximal dilation at 10−7 M: 74±2%, n=5), which was subsequently inhibited by hydrogen peroxide scavenger catalase (500 U/ml; maximal dilation at 10−7 M: 14±4%, n=5). HAAs obtained from CAD patients, responded to A23187 in a similar dose-dependent fashion (maximal dilation at 10−7 M: 68±3%, n=5), and this dilation was not affected by acute L-NAME exposure (maximal dilation at 10−7 M: 65±3%, n=5), but was reduced by catalase (maximal dilation at 10−7 M: 31±6%, n=6). To further determine the role of mitochondria-derived ROS in A23187-induced dilation, we treated arterioles with ETC inhibitor rotenone (1 mM). In non-CAD HAAs not pretreated with L-NAME, rotenone had no effect on A23187-induced dilation (maximal dilation at 10−7 M: 68±6%, n=5). In the presence of rotenone, however, prolonged L-NAME incubation no longer restored A23187-dependent dilation (maximal dilation at 10−7 M: 20±3%, n=5). Together, these data provide the first evidence that in healthy HAAs, prolonged L-NAME exposure changes the mechanism of vasodilation from NO to H2O2-mediated CAD-like vasodilation in response to A23187. In addition, the signaling process likely involves mitochondrial ROS. To elucidate the precise mechanisms underlying this process, future studies will be required to evaluate whether in healthy vasculature, basal NO inhibits mitochondrial ETC or other proteins such as NOXs through NO-mediated protein modification, which can be reversed by prolonged L-NAME incubation thereby promoting a switch from NO to H2O2 -mediated dilation.

This work was supported by the National Heart, Lung, and Blood Institute [R01HL096647].

This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.