Nitric oxide produced in endothelial cells affects vascular tone. To investigate the role of endothelial nitric oxide synthase (eNOS) in blood pressure regulation, we have generated mice heterozygous (؉͞؊) or homozygous (؊͞؊) for disruption of the eNOS gene. Immunohistochemical staining with anti-eNOS antibodies showed reduced amounts of eNOS protein in ؉͞؊ mice and absence of eNOS protein in ؊͞؊ mutant mice. Male or female mice of all three eNOS genotypes were indistinguishable in general appearance and histology, except that ؊͞؊ mice had lower body weights than ؉͞؉ or ؉͞؊ mice. Blood pressures tended to be increased (by approximately 4 mmHg) in ؉͞؊ mice compared with ؉͞؉, while ؊͞؊ mice had a significant increase in pressure compared with ؉͞؉ mice (Ϸ18 mmHg) or ؉͞؊ mice (Ϸ14 mmHg). Plasma renin concentration in the ؊͞؊ mice was nearly twice that of ؉͞؉ mice, although kidney renin mRNA was modestly decreased in the ؊͞؊ mice. Heart rates in the ؊͞؊ mice were significantly lower than in ؉͞؊ or ؉͞؉ mice. Appropriate genetic controls show that these phenotypes in F 2 mice are due to the eNOS mutation and are not due to sequences that might differ between the two parental strains (129 and C57BL͞6J) and are linked either to the eNOS locus or to an unlinked chromosomal region containing the renin locus. Thus eNOS is essential for maintenance of normal blood pressures and heart rates. Comparisons between the current eNOS mutant mice and previously generated inducible nitric oxide synthase mutants showed that homozygous mutants for the latter differ in having unaltered blood pressures and heart rates; both are susceptible to lipopolysaccharide-induced death.

The fundus of the guinea-pig stomach actively dilates in response to low increases in intragastric pressure. This physiological response, now called adaptive relaxation, accommodates the intake of liquid or food. It is independent of external innervation, resistant to ganglion blockade, but reflex in origin. The nerves involved are neither adrenergic nor cholinergic in nature. Non-adrenergic, non-cholinergic (NANC) nerves have now been recognized in many parts of the gastrointestinal tract and have recently been linked with release of nitric oxide (NO) on electrical stimulation. Here we show that adaptive relaxation in isolated stomach of the guinea pig is mediated by a NANC neurotransmitter substance indistinguishable from NO derived from L-arginine. This is substantiated by inhibition of adaptive relaxation by NG-monomethyl-L-arginine or N omega-nitro-L-arginine methyl ester, both inhibitors of NO synthesis, and by methylene blue, an inhibitor of soluble guanylate cyclase. There are two distinct neuronal pathways signalling NO-dependent adaptive relaxation, as evidenced by tetrodotoxin sensitivity. The first is a local reflex arc, the afferent fibres of which sense changes in intragastric pressure. The second is stimulated by an agonist for ganglionic nicotinic receptors. Thus, the functional significance of NO release from NANC nerves in the stomach is to bring about adaptive relaxation through a reflex response to increases in intragastric pressure.

The particulate enzyme, endothelial nitric oxide synthase (eNOS), produces nitric oxide to maintain normal vasodilator tone in blood vessels. In this study, we demonstrate that eNOS is a Golgi-associated protein in cultured endothelial cells and intact blood vessels. Using a heterologous expression system in HEK 293 cells, we show that wild-type myristoylated and palmitoylated eNOS, but not mutant, non-acylated eNOS targets to the Golgi. More importantly, HEK 293 cells expressing wild-type eNOS release substantially more NO than cells expressing the mutant, non-acylated enzyme. Thus, eNOS is a novel Golgi-associated protein, and Golgi compartmentalization is necessary for the enzyme to respond to intracellular signals and produce NO.