S-Nitrosoglutathione/Glutathione Disulphide/Cu 2+-Dependent Stimulation of L-arginine Transport in Human Platelets

Abstract

In this study, we have examined the effects of authentic nitric oxide (NO), NO + (NOBF 4), glutathione (GSH), glutathione disulphide (GSSG), and S-nitrosoglutathione (GSNO) in the presence and absence of Cu 2+, which thermally releases NO from S-nitrosothiols on the transport of L-arginine into the human platelet. The K M, apparent was unaffected by NO, NO +, GSH, and GSNO. However, Cu 2+ lowered K M, apparent by ∼2.85-fold. Cu 2+-dependent lowering of K M, apparent was also observed, albeit to a smaller extent when this ion was mixed with GSH (∼1.9-fold lower) and GSNO (∼2.0-fold). GSSG also lowered K M, apparent by ∼1.5-fold. The V max, apparent of L-arginine uptake was unaffected by NO, NO +, GSH, and Cu 2+. V max, apparent was stimulated by to the largest extent by GSNO (∼2.28-fold) and GSNO plus Cu 2+ (∼2.7-fold). GSSG and GSH plus Cu 2+ also increased V max, apparent by ∼1.9-fold. When these parameters are expressed in terms of transport efficiency (V max/K M) the largest effect of nearly 4.7-fold (over controls) was obtained by a combination of GSNO plus Cu 2+. These results suggest that platelet L-Arg transport is not affected either by NO or NO + but by a thiol-disulphide exchange reactions on the platelet L-Arg transporter, brought about by GSNO and GSSG. Based on these results, a GSNO/GSSG/Cu 2+ dependent regulatory mechanism for the uptake of L-arginine in human platelets has been proposed.