This study in pig colon descendens circular muscle investigated the possible role of phosphodiesterases (PDEs) (1) in the control of smooth muscle activity and (2) in the signal transduction of the 5-HT4 receptors located on the cholinergic neurons. Submaximal cholinergic contractions were electrically induced in colonic circular muscle strips and the influence of the non-selective PDE inhibitor 3-isobutyl-1-methyl-xanthine (IBMX) and selective inhibitors for the 5 classic PDE families (1–5) vinpocetine (PDE1), EHNA (PDE2), cilostamide (PDE3), rolipram (PDE4) and zaprinast (PDE5) was evaluated. IBMX and cilostamide concentration-dependently reduced the amplitude of the cholinergic contractions, as good as abolishing them at 30 and 0.3 μM respectively. EHNA only reduced the contractions significantly at the highest concentration tested (30 μM). IBMX and cilostamide also concentration-dependently inhibited submaximal cholinergic contractions induced with the muscarinic receptor agonist carbachol. The 5-HT4 receptor agonist prucalopride (1 μM) significantly enhanced the electrically induced cholinergic contractions. IBMX, vinpocetine and EHNA did not influence the facilitating effect of prucalopride but rolipram tended to enhance it. When rolipram was added after prucalopride, the facilitating effect of prucalopride was significantly enhanced. These results suggest that PDE3 is the main regulator of circular smooth muscle activity and that the signal transduction of 5-HT4 receptors on the cholinergic nerves towards the circular muscle layer is regulated by PDE4 in pig colon descendens.