AbstractThe inferior colliculus (IC) is the main ascending auditory relay station prior to the superior colliculus (SC). The morphology and origin of the connection from inferior to superior colliculus (I‐SC) was analyzed both by anterograde and retrograde tracing. Irrespective of the subregion of the IC in which they originate, the terminal fields of these connections formed two main tiers in the SC. While the dorsal one primarily involved the stratum opticum and the stratum griseum intermediale, the ventral one innervated the deep strata, although some fibers did connect these tiers. While the dorsal tier occupied almost the whole extension of the SC, the ventral one was mostly confined to its caudomedial quadrant. The fiber density in these tiers decreased gradually in a rostral gradient and the terminal fields became denser as the anterograde tracer at the injection site was distributed more externally in the cortex of the IC. Retrograde tracing confirmed this result, although it did not reveal any topographic ordering for the I‐SC pathway. Most presynaptic boutons of the I‐SC terminal field were located either inside or close to the patches of acetylcholinesterase activity. Together with previous anatomical and physiological studies, our results indicate that the I‐SC connection relays behaviorally relevant information for sensory‐motor processing. Our observation that this pathway terminates in regions of the superior colliculus, where neurons involved in fear‐like responses are located, reinforce previous suggestions of a role for the IC in generating motor stereotypes that occur during audiogenic seizures. J. Comp. Neurol. 499:716–731, 2006. © 2006 Wiley‐Liss, Inc.