To characterize endothelin (ET) receptors in the lower esophageal sphincter, we measured contraction of transverse strips from the guinea-pig lower esophageal sphincter induced by ET-related peptides and binding of 125I-ET-1 to cell membranes prepared from the lower esophageal sphincter circular muscle. Visualization of 125I-ET-1 binding sites in tissue was performed by autoradiography. ET-1 or ET-2 alone did not cause contraction or relaxation in resting strips. However, in carbachol precontracted lower esophageal sphincter strips, ET-1 and ET-2 caused marked, tetrodotoxin-insensitive relaxation. The ET-1-induced relaxation was abolished by BQ-123, an ETA receptor selective antagonist, but not inhibited by BQ-788, an ETB receptor selective antagonist. ET-3 and sarafotoxin S6c, a selective ETB receptor agonist, did not cause relaxation in the carbachol precontracted muscle strips. These clearly indicate that ETA receptors mediate relaxation. On the other hand, ET-3 and sarafotoxin S6c caused tetrodotoxin and atropine-insensitive contraction in the resting strips. The sarafotoxin S6c-induced contraction was inhibited by BQ-788, but not by BQ-123. Furthermore, ET-1 and ET-2 caused contraction of the resting muscle strips after pretreatment with BQ-123. This ET-1-induced contraction was also inhibited by BQ-788. Taken together, these indicate that ETB receptors mediate contraction. Autoradiography localized 125I-ET-1 binding to the lower esophageal sphincter circular muscle as well as longitudinal muscle of the esophagus. Binding of 125I-ET-1 to cell membranes prepared from the circular smooth muscle was saturable and specific. Analysis of dose-inhibition curves indicated the presence of two classes of receptors, ETA and ETB receptors. These results demonstrate that the guinea-pig lower esophageal sphincter possesses ETA receptors mediating relaxation and ETB receptors mediating contraction.