The effect of neurotensin on canine ileal circular muscle devoid of myenteric plexus was investigated using single and double sucrose gap techniques. Similar results were obtained with microelectrode techniques. Neurotensin caused a temperature-sensitive and dose-dependent biphasic response, an initial hyperpolarization associated with inhibition of contractile activity, followed by an excitatory phase, usually consisting of spike discharge and tonic and phasic contractions, for which depolarization was not required. Neither response was affected by tetrodotoxin, phentolamine, propranolol, or atropine. The hyperpolarization was associated with decreased membrane resistance, blocked by 10−7 M apamin, and converted to tonic depolarization by apamin (10−6 M). Tachyphylaxis to neurotensin occurred when the stimulation interval was less than 20 min. After Ca2+ depletion, depolarization was observed instead of the hyperpolarization; this depolarization was not affected by nitrendipine and was gradually abolished with repetitive stimulations at 20-min intervals. When Ca2+ was present, nifedipine did not alter the hyperpolarizing phase of the response but inhibited spiking and blocked all contractions. The excitatory phase of the response was enhanced by Bay K-8644. Neuromedin N elicited a response identical with that of neurotensin. The responses of the two peptides were completely cross tachyphylactic. Inhibitory junction potentials were not affected by neurotensin tachyphylaxis. It is concluded that neurotensin and neuromedin N activate apamin-sensitive, calcium-dependent potassium channels in circular muscle, causing membrane hyperpolarization and inhibition of muscle contraction. Release of intracellular calcium is involved in the activation of these potassium channels. When opening of potassium channels was inhibited, release of intracellular calcium caused depolarization. Neurotensin also activates L-type calcium channels, resulting in muscle contraction. Neurotensin does not appear to contribute to the compound inhibitory nerve response represented by the inhibitory junction potential.Key words: neurotensin, apamin, inhibitory junction potentials, ileum, circular muscle.