By applying microelectrode technic in isolated dog hearts, the nature of the so-called unidirectional conduction of the atrioventricular conduction was investigated. Retrograde conduction, i.e., conduction from the ventricle, occurred frequently in the isolated whole heart from which the sinus node was removed, or in a preparation composed of the interatrial septum, right atrium, right ventricle, interventricular septum, and atrioventricular conduction system. Retrograde conduction was blocked just as frequently. In such cases, this unidirectional block was found not to be ascribed solely to the refractory period of the atrioventricular node or of the atrial muscle fibers. The site of unidirectional block was found to be located in the domain of the ventricle, i.e., in the region adjacent to the postero-inferior margin of the membranous portion of the interventricular septum. This site is of a very limited extension. This point responded to ventricular stimuli, frequently by localized depolarization only, or in the manner of decremental conduction, whereas following atrial stimuli, it responded with full-sized action potential. By histologic examination, it was found that this point is situated in the ventricular portion of the bundle of His, just above the bifurcating point to the right and left branches.