The role of ATP in excitatory neurotransmission and its possible modulating role on acetylcholine (ACh) action were examined in the longitudinal muscle of the chicken anterior mesenteric artery (AMA) using tension recording technique. Electrical field stimulation (EFS) caused frequency-dependent contractions that were sensitive to tetrodotoxin (1 μM) and completely abolished by atropine (1 μM) in 63% of the preparations but in 36% of the preparations an atropine resistant component remained and was blocked by the non-specific purinergic antagonist, suramin (200 μM). Application of suramin alone from the beginning significantly and markedly decreased the EFS- induced contraction in a dose-dependent manner. Preparations contracted to exogenous applications of ATP and 2-methythio ATP (2-MeSATP) in dose-dependent manners. Desensitization of P2X receptor with its putative receptor agonist α,β-MeATP (1 μM, for 30 min) has no or insignificant enhancing effect on the contractile response. In contrast, desensitization of P2Y receptor with its putative agonist 2-MeSATP (1 μM, for 30 min) abolished the contractile response, indicating that this response is mediated through P2Y but not P2X receptor. In atropine-pretreated preparations, the contractile response to 2-MeSATP was significantly inhibited. The obtained findings of EFS and exogenous applications together indicate that ATP produces its action, at least partly, via presynaptic enhancement of the release of ACh and this occurs through activation of facilitatory P2Y receptor; this is in addition to the direct action of ATP on P2Y receptors being expressed on smooth muscle cells of the longitudinal muscle of chicken AMA.