Organic nitrates confer relief on Sphincter of Oddi spasm but they produce a weak relaxant effect on the gallbladder. We studied whether this is explained by a different biodegradation of nitroglycerin to nitric oxide (NO) in rabbits. Sodium nitroprusside induced relaxation of the SO is similar to that of the GB, while the relaxing effect of nitroglycerin was more prominent in the SO than in the GB. No difference in the relaxation response was seen to nitroglycerin and SNP in both tissues when they were mounted together with aortic rings. Neither baseline NO nor cGMP content increased in response to nitroglycerin, but both increased in the SO. Immunohistochemistry revealed the presence of NOS in SO and its absence in GB. Dissimilarity in nitroglycerin responsiveness of SO and GB is explained by a variance in nitroglycerin-biodegradation to NO.