The distributions of sources and terminals of callosal fibers in the high-frequency representation of AI were related to binaural and frequency maps in combined anatomical and electrophysiological experiments. Sources of callosal axons were retrogradely labeled with HRP. Distributions of axon terminals were determined by autoradiographic labeling with [3H]-proline and anterograde degeneration following callosal section. Regions in which cells exhibit summation or ipsilateral dominance and suppression contain higher concentrations of sources and terminals of callosal fibers than do regions in which cells exhibit monaural contralateral responses or contralateral dominance and suppression. Callosal axon terminals aggregate into columns. In sections cut parallel to the cortical surface callosal columns take on complex forms that exhibit certain consistent features. Two prominent elongated columns separated by a narrow zone of sparse callosal innervation run in a rostrodorsal to caudoventral direction through AI crossing several octaves of the frequency representation. Ventral to these columns, along the AI-AII border, less densely labeled callosal columns are in evidence. Low frequency representations of AI are interconnected but details of their innervation patterns were not worked out. Outside of AI there are regions that contain complex configurations of callosal columns. Several morphologically distinct types of neurons, located in laminae III through VI, were retrogradely labeled following injections of HRP into the opposite AI. About 94% of callosal neurons are pyramidal cells of layers III and IV.