Radiofrequency ablation of the slow pathway can prolong atrioventricular (AV) nodal properties and RR intervals during atrial fibrillation (AF) in many patients with AV nodal reentrant tachycardia. However, it is not well elucidated whether these changes are related to the presence of dual AV nodal pathway physiology. The aim of this study was to evaluate changes of AV nodal properties and RR intervals during AF caused by ablation of two specific areas in dogs. Assisted by fiberoptic endoscopy, linear lesions were created between the coronary sinus ostium and tricuspid valve annulus (area 1) or posterior to the ostium (area 2) in 15 dogs. Three additional dogs served as controls. The measurements were made under autonomic blockade. Catheter ablation could be assisted in all dogs by means of endoscopy. Linear lesions were confirmed at autopsy. AV nodal parameters and RR intervals showed no overall changes. Individual data showed that ablation of area 1 resulted in modification of AV nodal properties in 54.5% (facilitation in 36.3% and inhibition in 18.2%), whereas ablation of area 2 induced changes in 50% (facilitation in 10% and inhibition in 40%). The RR intervals were shortened in 33.3% and 20% and prolonged in 44.5% and 40% after ablation of areas 1 and 2, respectively. The RR intervals during AF correlated well with the Wenckebach cycle length and the AV node functional refractory period before and after ablation (r = 0.78 to 0.94, P < 0.01 for each). Ablation of the two specific areas around the coronary sinus ostium was equally effective in modifying AV nodal properties and the ventricular response during AF without dual AV nodal pathway physiology. The ventricular rate to AF after ablation correlated well with the residual AV nodal properties.