VASCULAR AND ANTIPLATETLET ACTIONS OF 1,2-GLYCERYL AND 1,3-GLYCERYL DINITRATE

1 The aim of this study was to investigate whether two metabolites of glyceryl trinitrate (GTN), 1,2 and 1,3-glyceryl dinitrate (1,2-GDN and 1,3-GDN) could account for the pharmacological effects of GTN. To thi s end the formation of nitric oxide (NO) from 1,2- and 1,3-GDN in the presence of bovine aortic smooth muscle cells (SMC) or endothelial cel ls (EC) was studied. The effects of various thiols on NO formation fro m these dinitrates was also evaluated. 2 1,2-GDN or 1,3-GDN (10(-10)-1 0(-5 M) caused a dose-dependent relaxation of rabbit aortic strips den uded of endothelium and precontracted with phenylephrine. The dinitrat es were less than one tenth as potent as GTN. 3 Incubation of 1,2-GDN or 1,3-GDN (75-2400 muM) with SMC for 30 min led to a concentration-de pendent increase in nitrite (NO2-) formation but this increase was les s than that produced from GTN. Likewise incubation of 1,2-GDN or 1,3-G DN with N-acetylcysteine (NAC), glutathione (GSH) or thiosalicylic aci d (TSA) (all at 1 mM) for 30 min at 37-degrees-C produced a concentrat ion-dependent increase in NO2- formation. 4 Platelet aggregation induc ed by thrombin (40 mu ml-1) was not modified by high concentrations of 1,2-GDN or 1,3-GDN (175-700 muM). However, aggregation was inhibited when platelets were exposed to 1,2-GDN or 1,3-GDN (700 muM) in the pre sence of SMC (0.24-1.92 x 10(5) cells) or EC (0.8-3.2 x 10(5) cells). These effects were abrogated by co-incubation with oxyhaemoglobin (Oxy Hb, 10 muM) indicating that they were due to NO release. The concentra tions of the dinitrates required to inhibit platelet aggregation by 50 % were about 15 times higher than for GTN in the presence of the same numbers of SMC or EC. 5 When NAC or TSA (both at 0.5 mM) were co-incub ated with platelets for 3 min in the presence of 1,2-GDN or 1,3-GDN, a concentration-dependent inhibition of platelet aggregation was observ ed. These anti-platelet effects were abolished by co-incubation with O xyHb (10 muM). Glutathione had no potentiating effects. 6 Thus the din itrate metabolites of GTN are metabolized to NO by SMC or EC and are a cted upon by thiols to form NO at concentrations about 10 times higher than those of GTN. In vivo, after oral or intravenous GTN, GDN levels are reached which are more than 10 times higher than those of GTN. Th ese data support the notion that part of the effects of GTN are due to the generation of NO from 1,2-GDN and 1,3-GDN by the cells of the vas cular wall.