The proposed pathways of chick cranial neural crest migration and their relationship to the rhombomeres of the hindbrain have been somewhat controversial, with differing results emerging from grafting and DiI-labelling analyses. To resolve this discrepancy, we have examined cranial neural crest migratory pathways using the combination of neurofilament immunocytochemistry, which recognizes early hindbrain neural crest cells, and labelling with the vital dye, DiI. Neurofilament-positive cells with the appearance of premigratory and early-migrating neural crest cells were noted at all axial levels of the hindbrain. At slightly later stages, neural crest cell migration in this region appeared segmented, with no neural crest cells obvious in the mesenchyme lateral to rhombomere 3 (r3) and between the neural tube and the otic vesicle lateral to r5. Focal injections of DiI at the levels of r3 and r5 demonstrated that both of these rhombomeres generated neural crest cells. The segmental distribution of neural crest cells resulted from the DiI-labelled cells that originated in r3 and r5 deviating rostrally or caudally and failing to enter the adjacent preotic mesoderm or otic vesicle region. The observation that neural crest cells originating from r3 and r5 avoided specific neighboring domains raises the intriguing possibility that, as in the trunk, extrinsic factors play a major role in the axial patterning of the cranial neural crest and the neural crest-derived peripheral nervous system.