Na-K-ATPase is associated with a variety of membrane populations in lacrimal acinar cells. Acinus-like structures formed by rabbit acinar cells in primary culture were incubated with horseradish peroxidase (HRP) to label basolateral and endosomal membranes and then analyzed by electron microscopy cytochemistry with the 3-3'-diaminobenzidine reaction or by fractionation and measurement of marker catalytic activities or immunoreactivities. HRP adsorbed to basolateral membranes at 4 degrees C. Fractionation showed it associated with low-density membranes enriched in acid phosphatase and TGN38 but containing only minor amounts of Na-K-ATPase. Cells internalized HRP to cytoplasmic vesicles, Golgi structures, and lysosomes at 37 degrees C. The major endosomal compartment revealed by fractionation coincided with major peaks of Na-K-ATPase and Rab6 and secondary peaks of galactosyltransferase and gamma-adaptin. Carbachol (10 microM) increased lysosomal and Golgi labeling. Thus most of the Na-K-ATPase is located in the basolateral membrane-oriented endosomal system, concentrated in a compartment possibly related to the trans-Golgi network. Constitutive and stimulation-accelerated traffic to and from this compartment may serve several exocrine cell functions.

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