Recent reports have suggested that the smooth muscle relaxant effects of drugs such as nitroprusside and nitroglycerin are mediated by increases in tissue levels of cyclic GMP. This hypothesis was examined by comparing the effects of nitroprusside, nitroglycerin, hydroxylamine and sodium azide on tension and cyclic GMP levels in rat vas deferens, rat myometrium and guinea-pig taenia coli. All four of the agents were capable of increasing cyclic GMP levels in these tissues but there did not appear to be a good correlation between cyclic GMP elevation and muscle relaxation in any of the tissues studied. For example, nitroprusside markedly elevated cyclic GMP levels in rat vas deferens and myometrium but had no relaxant effect on either tissue. Nitroglycerin was less effective than nitroprusside in elevating cyclic GMP levels, but was an effective relaxant in both tissues. Significant increases in myometrial cyclic GMP were seen with 0.1, 1 and 5 mM concentrations of hydroxylamine but only the highest concentration had a definite relaxant effect on the muscles. A similar concentration of nitroprusside produced a greater elevation of cyclic GMP than did hydroxylamine, but had no relaxant effect in this preparation. In guinea-pig taenia coli, significant increases in cyclic GMP levels were obtained with concentrations of sodium azide and hydroxylamine which had no effect on the contractile activity of the preparations. These results, together with previous results from this laboratory, suggest that the relaxant effects of this group of drugs in rat vas deferens, rat myometrium and guinea-pig taenia coli are not mediated by increases in tissue levels of cyclic GMP. Further experiments are necessary to determine whether a causal relationship exists between cyclic GMP elevation and relaxation in other types of smooth muscle.