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The development of the corpus callosum depends on a large number of different cellular and molecular mechanisms. These include the formation of midline glial populations, and the expression of specific molecules required to guide callosal axons as they cross the midline. An additional mechanism used by callosal axons from neurons in the neocortex is to grow within the pathway formed by pioneering axons derived from neurons in the cingulate cortex. Data in humans and in mice suggest the possibility that different mechanisms may regulate the development of the corpus callosum across its rostrocaudal and dorsoventral axes. The complex developmental processes required for formation of the corpus callosum may provide some insight into why such a large number of human congenital syndromes are associated with agenesis of this structure.