The shaping of nitric oxide signals by a cellular sink

To function as such, biological signalling molecules need to be inactivated. In the case of nitric oxide (NO), which serves as an intercellular messenger throughout the body (Moncada et al. 1991), much has been learnt during the last decade about the synthetic pathway, but how the molecule is disposed of under physiological conditions remains unknown. NO is generated in cells from l-arginine and O2 by NO synthases. Two isoforms, neuronal and endothelial (nNOS and eNOS) are constitutively expressed and typically become activated transiently as a result of a rise in cytosolic Ca2+. A third type, the inducible NO synthase (iNOS), can be expressed in many different cell types after exposure to inflammatory or proinflammatory mediators and this isoform manufactures NO continuously (Stuehr, 1999). Once produced, NO diffuses rapidly in three dimensions to elicit biological actions in neighbouring cells. Physiological NO signal transduction occurs through binding to the haem group of soluble guanylyl cyclase (sGC), leading to enzyme activation and cGMP accumulation (Waldman & Murad, 1987; Ignarro, 1991). However, NO can also contribute to tissue pathology by inhibiting mitochondrial respiration and promoting the generation of reactive free radicals (Gross & Wolin, 1995; Clementi et al. 1998; Brown, 1999; Heales et al. 1999). The rate of inactivation of NO will govern, inter alia, how far NO spreads within a tissue and at what concentrations (Wood & Garthwaite, 1994), and so is expected to be a critical determinant of whether NO acts as a physiological signal or as a toxin. The chemical reactivity of NO has been considered to be one means of disposal. NO can react with O2 (a process termed autoxidation) but this is far too slow at the concentrations existing in vivo to be of relevance (Ford et al. 1993; Kharitonov et al. 1994). A much more rapid reaction is with superoxide ions, but the resulting peroxynitrite anion is highly toxic, making it unlikely that this would serve as the primary physiological pathway (Beckman & Koppenol, 1996). Various biological mechanisms have also been proposed. Foremost among these is the reaction with haemoglobin (Hb) in circulating red blood cells, which yields nitrosylhaemoglobin and/or methaemoglobin plus nitrate ions (Beckman & Koppenol, 1996). The extent to which this reaction contributes to NO inactivation under physiological conditions remains uncertain, however, even in the case of NO produced by eNOS in blood vessels (Liao et al. 1999). We report here that mammalian cells themselves contain a powerful NO inactivating mechanism(s), or sink, that has properties that are well suited for shaping the kinetics and tissue concentrations of NO for targeting its receptor, sGC. The sink, however, is exhaustible so that during prolonged release NO rises to levels that exert pathological effects.