The gastrointestinal (GI) tract responds to a variety of stimuli through local and centrally mediated pathways. Changes in the intestinal microenvironment are sensed by vagal, spinal, and intrinsic primary afferent fibers. Sensory nerve endings located close to the lumen of the GI tract respond to pH, chemical composition of lumenal contents, or distortion of the mucosa. Afferents within the muscle layers are thought to be tension sensitive, whereas those located within the myenteric plexus are also thought to respond to changes in chemical composition and humoral substances. Subpopulations of these afferent fibers are activated by capsaicin. However, the exact location of these nerves is currently not known. The vanilloid receptor (VR1) is a nonselective cation channel that is activated by capsaicin, acid, and temperature. Antibodies to VR1 make it possible to determine the location of these afferents, their morphology, and their relationships with enteric nerves and other cell types in the GI tract. VR1‐like immunoreactivity was observed on nerves within myenteric ganglia and interganglionic fiber tracts throughout the GI tract. VR1 nerves were also observed within the muscle layers and had an irregular profile, with varicose‐like swellings along their lengths. Blood vessels within the GI wall had VR1‐immunoreactive nerve fibers associated with them. VR1‐like nerves and other immunopositive cells were also observed within the mucosa. In summary, VR1‐like immunoreactivity was found in several locations within the GI tract and may provide sensory integration of chemical, physical, or inflammatory stimuli. VR1‐like fibers appear to be predominantly spinal in origin, but a few vagal VR1‐like fibers exist in the stomach. J. Comp. Neurol. 465:121–135, 2003. © 2003 Wiley‐Liss, Inc.