The longstanding quest for the anatomical basis of the Wolff-Parkinson-White syndrome has left many unanswered questions. The ultrastructural morphology of the myocytes comprising accessory atrioventricular pathways, which are capable of rapid and variable conduction, is central to understanding the development and behaviour of this congenital anomaly, but remains unknown. Examination of three surgically resected pathways was performed to determine their underlying cellular morphology and the pattern of intercellular coupling, by correlative light microscopy, electron microscopy and confocal scanning laser microscopy combined with immunohistochemical localization of the cardiac gap-junctional protein, connexin43. Two left-sided pathways were composed of myocardium of 'normal working ventricular' type. The right-sided pathway was composed almost entirely of highly abnormal myocytes characterized by aberrant myofibril organisation, with a lack of A-band material and abnormal mitochondria, but normal intact intercalated disks no different from those seen in left-sided pathways. The gap junctions of all pathways were composed of connexin43 distributed as in ventricular myocardium, and not as found in atrial or atrioventricular nodal tissues. While myocytes of abnormal structure were present in one of the accessory atrioventricular pathways examined, all pathways had morphologically normal gap junctions, the structures responsible for efficient intercellular coupling, with a pattern of distribution suggestive of working ventricular myocardium.