A quantitative and morphometric evaluation of 125I-alpha-bungarotoxin binding in the rat hypothalamus

The distribution of 125I-alpha-bungarotoxin (alpha-BTX), a putative nicotinic cholinergic receptor ligand was studied both in vitro and in vivo in the suprachiasmatic nucleus (SCN), anterior hypothalamic area (AHA), and supraoptic nucleus (SON) of the hypothalamus. For in vitro studies 20 micron frozen frontal sections containing SCN were incubated with either radioligand or, unlabeled alpha-BTX plus 125I alpha-BTX and tissues were processed for light microscopic autoradiography. Areas of cresyl violet stained SCN sections were measured using a Bioquant Analysis System and grain counts and distributions were determined. For in vivo investigations third ventricular infusion of either 125I alpha-BTX, or unlabeled alpha-BTX with 125I alpha-BTX was performed, and 24 hours later animals were perfused pericardially and 1 micron serial plastic sections of the SCN were processed for light microscopic autoradiography. Localization of silver grains in 1 micron serial sections was evaluated in a double blind study. In vitro and in vivo labeling patterns in the hypothalamus were the same and compared well with previously examined paraffin-processed tissues from animals which had received third ventricular infusions of the neurotoxin. We observed a distinctive and specific labeling pattern of the SCN. Grains tended to localize diffusely and uniformly in more rostral regions, but clustered densely in the dorsal and lateral mid-SCN, and dorsally in the mid-caudal SCN. Grains were localized in the SCN where larger neurons were found. In the most caudal regions of the SCN no labeling was observed. Tissues from unlabeled alpha-BTX plus 125I alpha-BTX in vitro or in vivo studies did not demonstrate grain counts above background levels.(ABSTRACT TRUNCATED AT 250 WORDS)