Núñez Cristina, Víctor Víctor M, Tur Remedios, Alvarez-Barrientos Alberto, Moncada Salvador, Esplugues Juan V, D'Ocón Pilar
Departamento de Farmacología, Facultad de Medicina, Universidad de Valencia, Blasco Ibáñez, Valencia, Spain.
Circ Res. 2005 Nov 11;97(10):1063-9. doi: 10.1161/01.RES.0000190588.84680.34. Epub 2005 Oct 13.
It has been generally acknowledged that the actions of glyceryl trinitrate (GTN) are a result of its bioconversion into NO. However, recent observations have thrown this idea into doubt, with many studies demonstrating that NO is present only when there are high concentrations of GTN. We have explored this discrepancy by developing a new approach that uses confocal microscopy to directly detect NO. Intracellular levels of NO in the rat aortic vascular wall have been compared with those present after incubation with 3 different NO donors (DETA-NO, 3-morpholinosydnonimine, and S-nitroso-N-acetylpenicillamine), endothelial activation with acetylcholine, or administration of GTN. We have also evaluated the relaxant effects of these treatments on isolated rings of aorta following activation of the enzyme soluble guanylyl cyclase and their inhibitory action on mitochondrial respiration, which is an index of the interaction of NO with the enzyme of the electron transport chain cytochrome C oxidase. In the case of the various NO donors and acetylcholine, we detected a concentration-dependent relationship in the intensity of vascular relaxation and degree of NO fluorescence and an increase in the Michaelis constant (Km) for O2. GTN did not produce similar effects, and although clinically relevant concentrations of this compound caused clear, concentration-related relaxations, there was neither any increase in NO-related fluorescence nor an augmented Km for O2. The nature of these differences suggests that these concentrations of GTN do not release free NO but probably a different species that, although it interacts with soluble guanylyl cyclase in vascular smooth muscle, does not inhibit O2 consumption by vascular mitochondria.
人们普遍认为，硝酸甘油（GTN）的作用是其生物转化为一氧化氮（NO）的结果。然而，最近的观察结果使这一观点受到质疑，许多研究表明，只有在高浓度GTN存在时才会有NO。我们通过开发一种使用共聚焦显微镜直接检测NO的新方法来探究这种差异。将大鼠主动脉血管壁中的细胞内NO水平与用3种不同的NO供体（二乙三胺 NONOate、3-吗啉代硝酮和S-亚硝基-N-乙酰青霉胺）孵育后、用乙酰胆碱进行内皮激活或给予GTN后的水平进行了比较。我们还评估了这些处理对分离的主动脉环在激活可溶性鸟苷酸环化酶后的舒张作用以及它们对线粒体呼吸的抑制作用，线粒体呼吸是NO与电子传递链细胞色素C氧化酶相互作用的一个指标。对于各种NO供体和乙酰胆碱，我们检测到血管舒张强度和NO荧光程度之间存在浓度依赖性关系，并且氧的米氏常数（Km）增加。GTN没有产生类似的效果，尽管该化合物的临床相关浓度引起了明显的、与浓度相关的舒张，但既没有与NO相关的荧光增加，也没有氧的Km增加。这些差异的性质表明，这些浓度的GTN不会释放游离的NO，而是可能释放一种不同的物质，尽管它与血管平滑肌中的可溶性鸟苷酸环化酶相互作用，但不会抑制血管线粒体的氧消耗。
