We hypothesized that pituitary adenylate cyclase-activating polypeptide (PACAP) activates intracardiac postganglionic parasympathetic nerves and has a different effect than cervical vagal stimulation. We measured effective refractory period (ERP) and conduction velocity at four atrial sites [high right atrium (HRA), low right atrium (LRA), high left atrium (HLA), and low left atrium (LLA)] and minimum atrial fibrillation (AF) cycle length at 12 atrial sites during cervical vagal stimulation and after PACAP in 26 autonomically decentralized, open-chest, anesthetized dogs. PACAP shortened ERP to a similar extent at all four sites (HRA, 58 ± 2.0 ms; LRA, 60 ± 6.3 ms; HLA, 68 ± 11.5 ms; and LLA, 60 ± 8.3 ms). Low- and high-intensity vagal stimulation shortened ERP at the HRA, but not in the other atrial sites (low-intensity stimulation: HRA, 64 ± 4.0 ms; LRA, 126 ± 5.1 ms; HLA, 110 ± 9.5 ms; and LLA, 102 ± 11.5 ms; high-intensity stimulation: HRA, 58 ± 4.2 ms; and HLA, 101 ± 4.0 ms). Conduction velocity was not altered by any intervention. Minimum AF cycle length after PACAP was similar in both atria but was shorter in the right atrium than in the left atrium during vagal stimulation. After atropine administration, no interventions changed ERP. These results suggest that PACAP shortens atrial refractoriness uniformly in both atria through activation of intrinsic cardiac nerves, not all of which are activated by cervical vagal stimulation.