A new method was developed for specific labeling of primary vestibular afferent fibers from selected end-organs with horseradish peroxidase (HRP) applied extracellularly in the inner ear space. In 48 chinchillas, labeling was performed successfully in all animals by scratching the surface of the sensory end-organ of interest with an electrolytically sharpened needle and replacing the fluid in the vestibule with 30% HRP solution. Merely replacing the vestibular fluid (endo- and perilymph) with HRP did not label the ganglion cells or the afferent fibers in the brain stem. The specificity of labeling was verified by histological inspection of the ganglion cells and nerve fibers innervating the damaged and intact receptors. When the posterior semicircular canal and saccular receptors were scratched, labeled fibers and ganglion cells were found in the nerve and ganglion rostrodorsally and caudoventrally, respectively. Labeled ganglion cells from different superior vestibular nerve (SVN) receptors did not show as clear a segregation pattern as did labeled receptors from the inferior vestibular nerve (IVN). Once inside the brain stem, labeled fibers from the SVN receptors were rostral to those from the IVN receptors. The fibers of the vestibular root divided into an ascending and a descending branch which formed the vestibular tract. Labeled fibers from the SVN receptors divided rostrolaterally to those from the IVN receptors. In the vestibular tract, fibers coursed in different locations according to the receptor of origin. Fibers from the utriculus were lateral to those from the horizontal semicircular canal, which were lateral to those from the anterior semicircular canal. Fibers from the sacculus were lateral to those from the posterior semicircular canal.