Background Ablation near the papillary muscle (PM) may terminate ventricular fibrillation (VF) in Langendorff-perfused rabbit hearts. Whether this finding is reproducible in large animals in vivo is unclear. Methods and Results We performed non-contact endocardial mapping and/or high density epicardial mapping in 12 dogs and 16 swine. Among them 3 swine and 2 dogs underwent global ischemia during sustained VF. The dominant frequency (DF) decreased from 10.4짹1.8 Hz at baseline to 4.3짹1.2 Hz 12 min later (p<0.001). The maximum DF was primarily (76.3%) localized to the posterior PM (PPM). Propranolol (n=11) increased VF cycle length (82.0짹29.3 ms to 109.2짹29.3 ms, p<0.0001) and the life-span of the reentrant wavefronts (RWFs) (0.06짹0.10 s to 0.18짹0.23 s, p=0.001), facilitated the anchoring of RWFs to the PPM. Purkinje potentials preceding QRS onset were recorded on the PPM both during sinus rhythm and during VF. When epicardial map revealed repetitive breakthrough patterns in the PPM area, simultaneous endocardial map showed reentry around the PPM. VF inducibility was markedly reduced (from 100% at baseline to 22% post-ablation, p<0.0001) by radiofrequency (RF) ablation at the PPM, with a linear extension of the ablation line from the PPM to the mitral valve annulus and the left ventricular apex in 7 dogs, but not in 3 dogs with anterolateral wall ablation (100% to 97%, p=NS). The application of RF energy near the PPM frequently initiated VF in pigs (100% to 100%, n=7). Conclusion PPM frequently functioned as an anchor to the RWFs during VF in both dogs and swine with acute global ischemia or during propranolol infusions. RF ablation targeted at PPM and guided by Purkinje potentials reduced the inducibility of VF in dogs.