The specific purine-receptor antagonist, arylazido aminopropionyl ATP (ANAPP3), was used to determine if purines released from nerves of the guinea-pig was deferens contribute to the neurogenic response. ANAPP3, which is a photoaffinity label, antagonized contractile responses of the in vitro vas deferens to transmural stimulation, reversibly in the presence of the compound and irreversibly after its photoactivation in the presence of the tissues. The antagonism by ANAPP3 was augmented by depletion of norepinephrine produced by reserpine pretreatment. Responses of untreated tissues were only slightly antagonized by the alpha-adrenoceptor blocker prazosin. However, neurogenic responses were markedly reduced in the combined presence of ANAPP3 and prazosin. ANAPP3 did not affect the release of tritium from tissues prelabeled with [3H]norepinephrine. The initial phasic component of the neurogenic response was preferentially antagonized by ANAPP3 whereas the secondary more tonic component of the response was preferentially antagonized by prazosin and reserpine pretreatment. Small, residual responses remaining after chemical sympathectomy produced by 6-hydroxydopamine pretreatment were potentiated, rather than inhibited, by ANAPP3 and were, unlike untreated tissues, sensitive to atropine. These and previous findings indicate that ATP or a related purine, originating from adrenergic neurons, acts as a co-transmitter with norepinephrine in this tissue.