Supraoptic nucleus (SON) neurons receive a prominent gamma-amino-butyric acid (GABA) input. This study evaluated the hypothesis, partly on the basis of recent electrophysiological data, that this innervation might arise from GABAergic neurons located in the ventral diagonal band of Broca area. For retrograde transport studies, pentobarbital-anesthetized male Long-Evans rats received 0.03-0.20-microliter injections of a suspension of rhodamine tagged latex microspheres into the SON. In two cases where such injections were confined to the SON, less than 60 retrogradely labeled neurons were detected in the ipsilateral diagonal band. In three animals where injections extended into the perinuclear zone around the SON, more than 2,000 retrogradely labeled cells were counted in the ipsilateral diagonal band. For anterograde transport studies, another group of animals received either 30% horseradish peroxidase (HRP) in 0.5% poly-L-ornithine (0.05-0.10 microliter injections) or Phaseolus vulgarus (iontophoresed from a 2% solution) into the diagonal band. After survivals of 18-24 hours (HRP) or 5 days (PHAL-L) labeled axon terminals invested the perinuclear zone above the SON. The presence of just a single fiber within the nucleus indicated a minor projection to the SON itself. The HRP-injected material was processed for ultrastructural examination and revealed dense HRP-labeled axon terminals in this perinuclear zone, most often (98%) forming axodendritic appositions. A postembedding colloidal gold technique to visualize GABA-synthesizing terminals revealed that fewer than 5% of these perinuclear HRP-labeled terminals also exhibited GABA-like immunoreactivity. Within the SON, where GABAergic axon terminals are abundant, few (less than 5%) GABAergic terminals contained HRP.(ABSTRACT TRUNCATED AT 250 WORDS)