Elasmobranch rectal gland cell: autoradiographic localization of [3H]ouabain-sensitive Na, K-ATPase in rectal gland of dogfish, Squalus acanthias.
Specific binding of radiolabeled inhibitor was employed to localize the Na-pump sites (Na,K-ATPase) in rectal gland epithelium, a NaCl-secreting osmoregulatory tissue which is particularly rich in pump sites. Slices of gland tissue from spiny dogfish were incubated in suitable [3H]ouabain-containing media and then prepared for Na,K-ATPase assay, measurement of radiolabel binding, or quantitative freeze-dry autoradiography at the light microscope level. Gross freezing or drying artifacts were excluded by comparison with additional aldehyde-fixed slices. Characterization experiments demonstrated high-affinity binding which correlated with Na,K-ATPase inhibition and half-saturated at approximately 5 microM [3H]ouabain. At this concentration, the normal half-loading time was approximately 1 h and low-affinity binding to nonspecific sites was negligible. Autoradiographs from both 1- and 4-h incubated slices showed approximately 85% of the bound [3H]ouabain to be localized within a 1-micrometer wide boundary region where the highly infolded basal-lateral cell membrane are closest to the mitochondria. These results establish that most of the enormous Na,K-ATPase activity associated with rectal gland epithelium is in the basal-lateral cell membrane facing interstitial fluid and not in the luminal membrane facing secreted fluid. Moreover, distribution along the basal-lateral membrane appears to be nonuniform with a higher density of enzyme sites close to mitochondria.
D000818:Animals; D001345:Autoradiography; D001693:Biological Transport, Active; D004284:Dogfish / Q000378:metabolism*; D006651:Histocytochemistry; D008854:Microscopy, Electron; D010042:Ouabain / Q000378:metabolism; D012491:Salt Gland / Q000201:enzymology* / Q000378:metabolism / Q000648:ultrastructure; D012754:Sharks / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D014316:Tritium
D014316:Tritium; D010042:Ouabain; D012964:Sodium; D000254:Sodium-Potassium-Exchanging ATPase