Stimulation-associated Redistribution of Na,K-ATPase in Rat Lacrimal Gland

To test the possibility that stimulation of secretion leads Na,K-ATPase to be recruited from cytoplasmic pools and inserted into basal-lateral plasma membranes, we surveyed the subcellular distributions of Na,K-ATPase in resting and stimulated fragments of rat exorbital lacrimal gland. After a two-dimensional separation procedure based on differential sedimentation and density gradient centrifugation, we defined six density windows, which differ from one another in their contents of biochemical markers. The membranes equilibrating in window I could be identified as a sample of basal-lateral membranes; in resting preparations these membranes contained Na,K-ATPase enriched 16.6-fold with respect to the initial homogenates. Windows II through VI contained various cytoplasmic membrane populations; these accounted for roughly 80% of the total recovered Na,K-ATPase activity. Thirty-minute stimulation with 10 microM carbachol caused a 1.4-fold increase (P less than 0.05) in the total Na,K-ATPase content of window I; this increase could be largely accounted for by a 1.7-fold decrease in the total Na,K-ATPase content of density window V. Acid phosphatase activity also redistributed following stimulation, but it was recruited from a different source, and it was inserted into membranes equilibrating in windows II and III as well as into the membranes of window I.