Direct in vivo demonstration by radioautography of specific binding sites for calcitonin in skeletal and renal tissues of the rat

Abstract

An in vivo binding assay using radioautography was employed to visualize calcitonin receptors in rat tissues. At 2 min after intravenous injection of biologically active 125I-salmon calcitonin, free hormone was separated from bound hormone by intracardiac perfusion with lactated Ringer's followed by fixation with 2.5% glutaraldehyde. Various tissues were removed and processed for light and electron microscope radioautography. These were compared to tissues removed from animals that received identical amounts of labeled hormone with a large excess of unlabeled calcitonin. Among the tissues investigated, kidney and bone demonstrated labeling. In kidney, most silver grains were located over vesicles below the brush border of cells of theproximal convoluted tubules. These grains were still present after simultaneous injection of excess unlabeled hormone and most likely represented binding to sites involved with ingestion and degradation of hormone from the urinary filtrate. In contrast, grains localized to the basal surfaces of distal convoluted tubule cells were significantly reduced in number in control animals and represented sites of saturable, specific hormone binding. In bone, specific binding sites were found only at the periphery of osteoclasts. These labeled cells were located at resorption sites examined in tibia, humerus, and alveolar bone. This demonstration of the localization of 124I-calcitonin in situ provides a new approach for study the interaction of calcium-regulating hormones with their target cells.