Fallopian tube smooth muscle contractions are physiologically related to transport of the ovum within the oviduct. Nitric oxide (NO) has proved to be a mediator of tubal contractility. The main pathway by which NO exerts its relaxing effect on tubal contractions has not been fully elucidated. NO-mediated effects may be cyclic guanosine monophosphate (cGMP)-dependent or cGMP-independent. The objective of the present study was to investigate whether a NO-cGMP pathway is present in the Fallopian tube and, if present, to examine whether this pathway is involved in tubal contractility. Tubal smooth muscle strips were mounted in organ baths for measurement of tissue cGMP and for isometric recording of contractile activity. Following administration of the NO donor spermine NONOate a more than three-fold increase in tissue levels of cGMP was measured. Pretreatment with inhibitors of cGMP production prior to administration of spermine NONOate resulted in similar levels of cGMP as found in strips exposed to only plain buffer solution. Administration of spermine NONOate to muscle baths resulted in a significant inhibition of contractile activity, while pretreatment with inhibitors of cGMP production almost eliminated the relaxing effect of the NO donor. This study showed that a NO-cGMP pathway is present in the Fallopian tube and that the pathway is involved in Fallopian tube contractility.