Autoradiographic localization of α-bungarotoxin binding sites in the suprachiasmatic region of rat brain

High affinity α-bungarotoxin (α-BTX) binding sites of the hypothalamus in and near the suprachiasmatic nuclei (SCN) were mapped by in vitro macroautoradiographic analysis. Adult male rats were killed at specific circadian phases. Their brains were rapidly dissected out and frozen sections were made at a thickness of 16 μm. After having been mounted on slides, the sections were incubated with iodinated α-BTX (3 nM), washed and exposed to X-ray film. Analyses of binding were performed with the aid of a digital video densitometer system. Autoradiographic loci that bound α-BTX were traced and the image of the SCN histology from the adjacent section was superimposed. In this way the exact relationships of the topography of areas that bound α-BTX and those that contained SCN cell bodies (and other hypothalamic landmarks) could be observed. Non-specific binding was tested by incubation in the presence of 3 μM unlabelled α-BTX and was found to be very low and uniform throughout the sections. Hypothalamic areas that bound α-BTX included the SCN, supraoptic, periventricular, lateral and anterior hypothalamic nuclei. In the rostral SCN, α-BTX binding coincided with the nucleus proper. Caudally this relationship dissociated so that at mid-SCN α-BTX bound dorsally and laterally both within and outside the SCN and by the most caudal portion of the nucleus, α-BTX binding was entirely outside the SCN in a vertical band dorsal to the SCN. This topography suggests that α-BTX binding may be coincident with a major output pathway that courses dorsally and caudally from the SCN.