Anterograde tracing of retinohypothalamic afferents with Fluoro-Gold

The anterograde neuronal tracing properties of Fluoro-Gold (FG) were characterized in this study by its ability to label the retinohypothalamic tract (RHT) upon pressure injection of the substance into the vitrous body of the eye in the Djungarian hamster, Phodopus sungorus. Tracing was compared to the anterograde neuronal transport of cholera toxin B subunit (CTB), Fast blue (FB), Phaseolus vulgaris leucoagglutinin (PHA-L) and biocytin. After survival times that ranged from 24 h to 4 weeks, a major projection was found to the bilateral hypothalamic suprachiasmatic nuclei (SCN). Labeling was also found in the anterior medial preoptic nucleus and, in relatively sparse amounts, in the lateral geniculate nucleus, superior colliculus and lateral habenular nucleus. Similar results were obtained upon injection of CTB or FB, respectively, into the eye, whereas the application of PHA-L or biocytin did not label retinal afferents. The combined injection of FG and CTB or FB into the same eye labeled retino-afferent fibers only when FG was applied three days before the injection of the other tracers. Retrogradely labelled neurons were seen occasionally in the hypothalamus which may provide a sparse retinopetal projection. Additional experiments combining FG tracing and the immunofluorescent detection of the neuropeptides substance P (SP) or vasoactive intestinal polypeptide (VIP) in the SCN showed that FG-containing punctae were accumulated in the vicinity of immunoreactive cell bodies. Our data demonstrate that FG may be used as an anterograde axonal tracer of the retinohypothalamic pathway.