The effect of nitric oxide (NO) donors on the release of mucus from a suspension of isolated gastric cells was investigated by using an enzyme-linked immunosorbent assay for rat gastric mucin. Isosorbide dinitrate (ISDN, 0.1-2 mM) produced a dose-related stimulation of mucus secretion, without affecting the viability of the isolated cells as determined by trypan blue exclusion or acid phosphatase release. In a comparable concentration range to that stimulating mucus release, ISDN elevated the guanosine 3',5'-cyclic monophosphate (cGMP) content of cell suspensions enriched with mucous cells. The nitrosothiol S-nitroso-N-acetylpenicillamine (0.3 mM), which spontaneously liberates NO, likewise stimulated mucus release, and this action was blocked by 10 microM oxyhemoglobin, which scavenges NO. Nitroprusside (1 mM), dibutyryl cGMP (0.01-1 mM), and the cGMP phosphodiesterase inhibitor M & B 22948 (0.1 mM) also increased mucus release. Thus generators of NO stimulate mucus secretion by rat gastric mucosal cells, which may reflect the elevation of intracellular cGMP. These findings, along with the presence of NO synthase in the gastric epithelial cells, suggest an effector role for NO in mediation of gastric mucus release.