The distribution of axons and axon varicosities containing GABA was studied in the superior cervical ganglion of rat by light and electron microscopic immunohistochemistry. Two different polyclonal antibodies were used, which had been made against GABA conjugated by glutardialdehyde to bovine serum albumin. GABA-like immunoreactivity occurred in many axons within the cervical sympathetic trunk and in axons and axon varicosities around the principal nerve cells in the superior cervical ganglion. GABA-positive axons were intermingled with non-stained axons, except for a small group of fibers in the trunk where the staining was absent. The rostral part of the ganglion and some scattered patches were more densely innervated by GABA-positive axons than the middle and caudal parts. Within dense areas, some of the large ganglion cells were abundantly surrounded by GABA-positive nerve fibers, while the vicinity of others was devoid of any immunoreactive axon terminals. None of the principal ganglion cells contained GABA-like immunoreactivity, although a class of small cells scattered within the ganglion was stained. Transection of the cervical sympathetic trunk for 11 days caused the disappearance of GABA-like positivity from most of the fibers, and only very little GABA-like staining was revealed in some small cells, which resembled satellite cells. Ultrastructurally, the GABA-positive nerve fibers were unmyelinated. However, their terminal branches and varicosities accumulated around the perikarya and dendrites of certain principal ganglion cells were partly wrapped in glial processes. The present results provide evidence that the superior cervical ganglion of adult rat receives a significant number of GABA-positive axons from the cervical sympathetic trunk and that these axons provide an innervation which is heterogeneously distributed within the superior cervical ganglion and on ganglionic cells. The source and function of the GABA-positive axons remain to be elucidated.