1. Inhibitory non-adrenergic, non-cholinergic (NANC) responses were studied in isolated strips from the rabbit internal anal sphincter. 2. In the presence of atropine and guanethidine, transmural field stimulation induced frequency-dependent relaxations that reached a plateau at frequencies > or = 4 Hz. 3. These relaxations were inhibited by apamin (10(-6) M) and by N omega-nitro-L-arginine (L-NOARG, 10(-4) M). With these two substances in combination, relaxations were still seen in response to field stimulation, but only at frequencies > 2 Hz. 4. In the presence of both apamin (10(-6) M) and L-NOARG (10(-4) M), responses at high frequencies consisted of a fast relaxation followed by a slow return to prestimulus tension level. alpha-Chrymotrypsin hastens the return of tension to prestimulus level after high frequency stimulation. 5. Zinc-protoporphyrin IX, an inhibitor of haeme oxygenase, had a significant inhibitory effect on relaxations induced by transmural field stimulation. It was found, however, that responses to sodium nitroprusside and to isoprenaline (both 10(-9)-10(-4) M) were reduced comparably, indicating that the effect of zinc-protoporphyrin IX was unspecific. 6. It is concluded that pharmacological analysis allows identification of at least three distinguishable components of the inhibitory NANC innervation of the rabbit internal anal sphincter. The study does not allow conclusions about the role of carbon monoxide, a recently proposed mediator of NANC responses in opossum internal anal sphincter.