The congenital absence of the corpus callosum, a brain anomaly frequently noted in humans, has been recently found to occur in some mice of the ddN strain in our laboratory. In the brains of these mice, the Probst's longitudinal bundle is always present on both cerebral hemispheres. In this research, the neuroanatomical features of the constituent fibers of this bundle were studied by iontophoretical injections of horseradish peroxidase into different loci in the neocortex of acallosal mouse brains. The results revealed that (1) certain cortical fibers of the Probst's bundle terminate in the ipsilateral neocortex; (2) some commissural fibers in the longitudinal bundle originate from the cells in the wide neocortical regions, and project to the opposite hemisphere in homotopic as well as heterotopic regions over the ventral hippocampal commissure; (3) the fibers from different cortical regions are arranged in a topographic manner within this bundle. The present data clearly demonstrate that a good portion of fibers in the Probst's longitudinal bundle seen in the congenitally acallosal mouse brain are corticocortical in nature, which indicates that this bundle has an ipsilateral neocortical association function.