Cytological transformations associated with parietal cell stimulation: critical steps in the activation cascade.
Cultured rabbit parietal cells were used to evaluate morphological responses to activators and inhibitors of HCl secretion. Immunofluorescence was used to localize the proton pump protein, H, K-ATPase, and the apical membrane-cytoskeletal linker protein, ezrin; fluorescent-labeled phalloidin was used as a marker of F-actin. Treatment of healthy control parietal cells with secretagogues resulted in exaggerated swelling of apical membrane vacuoles, presumably with the accumulation of HCl and water. Thus stimulation-associated swelling of apical vacuoles was blocked by inhibitors that work at various steps in the secretion-activation cascade. When secretion was blocked by agents that prevent the translocation of H,K-ATPase-rich tubulovesicles to apical membrane vacuoles (such as H2-receptor antagonists and protein kinase A inhibitors), the general resting morphology was maintained. ME-3407 (a functional analogue of wortmannin) was unique in preventing H, K-ATPase redistribution and effecting the delocalization of ezrin from apical membrane vacuoles. When secretion was blocked by agents that inhibit the H+ pump or induce H+ backflux, the translocation of H,K-ATPase to apical membrane vacuoles occurred but the large vacuolar swelling associated with HCl and H2O accumulation was greatly diminished. These data support the membrane recycling/recruitment hypothesis of HCl secretion in which H, K-ATPase-rich tubulovesicles are recruited from a cytoplasmic domain to the apical surface, and they are inconsistent with models proposing that the tubulovesicles, regardless of shape, are contiguous with the apical plasma membrane. These studies also demonstrate the utility of the parietal cell culture model in distinguishing a general site of action for various inhibitors and antisecretory agents.
Agnew
B J
BJ
Department of Molecular and Cell Biology, University of California, Berkeley, CA 94720, USA.
Duman
J G
JG
Watson
C L
CL
Coling
D E
DE
Forte
J G
JG
D000199:Actins / Q000032:analysis / Q000378:metabolism; D000818:Animals; D000897:Anti-Ulcer Agents / Q000494:pharmacology; D001692:Biological Transport / Q000502:physiology; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D003598:Cytoskeletal Proteins; D004789:Enzyme Activation / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017506:H(+)-K(+)-Exchanging ATPase / Q000378:metabolism*; D006851:Hydrochloric Acid / Q000378:metabolism; D007093:Imidazoles / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D009853:Omeprazole / Q000494:pharmacology; D010295:Parietal Cells, Gastric / Q000166:cytology / Q000201:enzymology / Q000378:metabolism; D010750:Phosphoproteins / Q000032:analysis / Q000378:metabolism; D054328:Proton Pump Inhibitors; D017494:Proton Pumps / Q000378:metabolism; D011725:Pyridines / Q000494:pharmacology; D011817:Rabbits; D011970:Receptors, Histamine H2 / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D013449:Sulfonamides; D013844:Thiazoles / Q000494:pharmacology; D013861:Thiocyanates / Q000494:pharmacology; D014617:Vacuoles / Q000378:metabolism; D000099149:Ezrin
D000199:Actins; D000897:Anti-Ulcer Agents; D017868:Cyclic AMP-Dependent Protein Kinases; D003598:Cytoskeletal Proteins; D004791:Enzyme Inhibitors; D017506:H(+)-K(+)-Exchanging ATPase; D006851:Hydrochloric Acid; D007093:Imidazoles; D007546:Isoquinolines; D009853:Omeprazole; D010750:Phosphoproteins; D054328:Proton Pump Inhibitors; D017494:Proton Pumps; D011725:Pyridines; D011970:Receptors, Histamine H2; D013449:Sulfonamides; D013844:Thiazoles; D013861:Thiocyanates; D000099149:Ezrin; C106473:ME 3407; C035235:Sch 28080; C024553:sodium thiocyanate; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide