Precise extrinsic afferent (visceral sensory) and efferent (sympathetic and parasympathetic) innervation of the gut is fundamental for gut-brain cross talk. Owing to the limitation of intrinsic markers to distinctively visualize the three classes of extrinsic axons, which intimately associate within the gut mesentery, detailed information on the development of extrinsic gut-innervating axons remains relatively sparse. Here, we mapped extrinsic innervation of the gut and explored the relationships among various types of extrinsic axons during embryonic development in mice. Visualization with characterized intrinsic markers revealed that visceral sensory, sympathetic, and parasympathetic axons arise from different anatomic locations, project in close association via the gut mesentery, and form distinctive innervation patterns within the gut from embryonic day (E)10.5 to E16.5. Genetic ablation of visceral sensory trajectories results in the erratic extension of both sympathetic and parasympathetic axons, imp...