Binding and Uptake of [125I]Iodoprolactin by Epithelial Cells of the Rat Choroid Plexus: Anin VivoAutoradiographic Analysis*

Abstract

Light and electron microscopic (EM) autoradiography were used to determine the subcellular fate of [125I] iodo-PRL that specifically binds to epithelial cells of the rat choroid plexus. Experimental animals (total binding) received an external jugular vein injection of ovine [125I]iodo-PRL, whereas control animals (nonspecific binding) received an identical dose of [125I]iodo-PRL plus a 500-fold excess of unlabeled PRL. Experimental and control animals were killed by vascular perfusion 3, 20, 40, 60, and 240 min after hormone injection. Choroid plexuses were removed, processed to Epon, and prepared for light and EM autoradiography. Quantification of silver grains at the light microscopic level revealed an intense autoradiographic reaction over the epithelial cells of experimental animals 3, 20, 40, and 60 min after hormone injection. The co-injection of unlabeled PRL resulted in a statistically significant reduction in the autoradiographic reaction throughout the initial 60 min of the test period, reflecting specific binding of PRL during this interval. Analysis of EM autoradiographs revealed that the initial binding of [125I]iodo-PRL occurred primarily at the basal and lateral plasmalemma of the epithelial cells. With time there was a dramatic reduction in the amount of radiolabel localized to the cell membrane and a concomitant increase in silver grains localized over the interior of the cells. Internalized [125I]iodo-PRL exhibited a preferential localization to small cytoplasmic vesicles, short tubules, multivesicular bodies, and dense bodies. Combined acid phosphatase cytochemistry and EM autoradiography revealed that only 11.2% of the internalized silver grains were localized over acid phosphatase-positive structures 40 min after injection. The results indicate that subsequent to the initial binding of [125I]iodo-PRL to the cell membrane, internalization and translocation occurs primarily to nonlysosomal cytoplasmic organelles. The study also demonstrates at least a partial or ultimate entrance of internalized PRL to the lysosomal system.