Focal AF is frequently triggered by ectopic beats mostly originating from the pulmonary veins (PVs). So far, the morphological substrate for this entity is not well defined. Therefore, the distribution of myocardial cells within the PV were examined as potential target sites for RF application. The PVs (118) of 30 human autopsied hearts (age of death 63 +/- 13 years, 17 men) were dissected in their complete circumference starting 1 cm from the ostium. Tissue sections of the PV were stained with hematoxylin-eosin and with Masson's trichrome. To characterize the developmental state of the myocardial tissue in the PV, immunohistochemistry was performed with antibodies reacting with antigens which are stage specifically expressed during cardiac development (HNK1/Leu7, alpha-SMA, calponin and desmin). Furthermore, proliferative activity was assessed using antibodies against the Ki-67 antigen (MIB-1). In two hearts a left-sided common PV ostium was found. The other hearts showed four separated PV ostia. The ostium diameter of the right inferior PV (1.2 +/- 0.3 cm) was significantly smaller (P < 0.05) than remaining PV ostia (right superior 1.5 +/- 0.2, left superior, 1.5 +/- 0.3 and left inferior 1.4 +/- 0.3 cm) of the 118 specimen. There was no significant difference in the presence of myocardium in the PV 1-cm distant from the ostium comparing the right superior (78%), the right inferior (81 %), the left superior (81%), and the left inferior (81%) PV. In 54% of cases the myocardial bundles covered the complete PV circumference. In up to 38% of the small extensions of the myocardial bundles myocardial cells, characterized by distinct cross-striations and spindle shape were found. However, since these cells could not be labeled for other markers than desmin, their immature state seems unlikely. The anatomic distribution of myocardium in the PV suggests that RF applied to the entire circumference may be frequently required for its electrical isolation. Whether spindle-shaped myocytes have different electrophysiological behavior has to be further investigated.